Jjst magazine 2015 final

JAMAICAN JOURNAL OF SCIENCE & TECHNOLOGY, Volume 26 ISSN: 1016-2054 |

JJST ISSN: 1016-2054

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Published by: The Scientific Research Council, Information Services Division, Hope Gardens Complex, Kingston 6, Jamaica W.I.

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JAMAICAN JOURNAL OF SCIENCE & TECHNOLOGY, Volume 26 ISSN: 1016-2054

Table of Contents

3 | From the Editor 4 | Editorial Page

5 | Research Briefs THE FORUM 7 | The Experiences of the Scientific Research Council (SRC) in Providing Cost Effective and Sustainable Wastewater Management Options (Julia Brown)

MAIN ARTICLES 20 | A Review of Dibenzyl Trisulfide an Anti-Cancer Polysulfide Isolated from Petiveria alliacea (Lawrence A.D. Williams, Harald Rosner and Wolfgang Kraus) 26 | Potential of Two Jamaican Essential Oils against Mosquitoes and Bacteria (Chenielle M. Delahaye, Sylvia Mitchell and Dwight Robinson) 34 | Dietary Intake of Trace and Heavy Metals from Consumption of Instant Coffees on the Jamaican Market (Johann M.R. Antoine, Leslie A. Hoo Fung and Charles N. Grant) 46 | Assessment of Cadmium, Mercury and Lead Concentrations in Jamaican Conch, Lobster, and Tilapia (Leslie A. Hoo Fung, Vaughn R. Rattray and Gerald C. Lalor)

The Experiences of the SRC....?

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Review of Dibenzyl Trisulfide......

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Potential of Two Jamaican Essential Oils...

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Dietary Intake of Trace and Heavy Metals from...

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RESEARCH IN PROGRESS [Young Scientists’ Abstracts] 71 | Production of Solanum tuberosum L Microtuber using Temporary Immersion System (Shishca Higgins) 73 | Guide for Authors

Assessment of Cadmium, Mercury...

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From the Editor

A paramount challenge for Jamaica today is to enhance productivity – particularly through expanding innovative, globally competitive goods and services. These will originate in large part from tourism, agriculture, the creative arts and music; sports, shipping and mining may also have a role, as well as integrative activities, cutting across these categories. Science and Technology will feature prominently in this latter group, providing basic Research & Development (R&D) to drive knowledge acquisition and on-going innovation in order to move our products up the value chain. In the area of agriculture, our high endemicity, rich ethnobotanical folklorical knowledge, and an environment which seems to imbue even our imported crops (coffee, cocoa, ginger, marijuana) with special, desirable properties, has spawned over many years a vigorous enterprise in natural products research which continues to yield innovative discoveries and inventions, some patented, which have yet to be fully exploited agriculturally or industrially. And at the root of all our agricultural efforts, will be the effective management of our water resources which have come under increasing pressure over recent years.

In our current 2015 issue, the FORUM article focuses on the problem of wastewater management. Recent disputes over burning dump sites and clogged drainage, indicate clearly the economic consequences of the mismanagement of wastes. Dr Julia Brown of the SRC reports on the valiant efforts of the SRC over the years to improve the management of wastewater, both domestically and industrially, through the development of patented methodologies aimed at mitigating the threat of pollution to our important groundwater and riverine water resources. These innovations, however, are yet to be widely embraced, although the need to adopt them becomes increasingly cogent.

Two articles focus on health-related extracts from local plants. Dr Lawrence Williams of the SRC and his collaborators, report that dibenzyl trisulphide (DTS) the active principle in extracts from Guinea Hen Weed (Petiveria alliaceae) is a potent anticancer agent with a wide therapeutic window, targeting one of the two major families of cell-signalling proteins, the RTK (receptor tyrosine kinase) family. Drs Chenielle Delahaye, Sylvia Mitchell and Dwight Robinson (UWI) investigate the potential of essential oils from the bottlebrush plant (Callistemon viminalis) and from lemongrass (Cymbopogon citratus) as mosquito repellants and antibacterial agents, the importance of which is obvious in the face of the ravages of the chikungunya virus (Chick-V) and the zika virus (Zik-V).

Two reports from the International Centre for Environmental and Nuclear Research (ICENS) highlight the importance of understanding the nature of the foods we consume and export. Monitoring content and establishing and/or adhering to standards for concentrations of trace and heavy metal in coffee (first article) and of heavy metals in our sea foods such as conch, lobster and fish (second article), are critical in protecting our consumers and sustaining the markets they represent.

A Research in Progress report from Shischa Higgins and SRC researchers, looks at improving the efficiency of microtuber propagation of irish potatoes – a technique which might be more acceptable than tissue culture, to the farming community, and have economic importance.

But we must celebrate not only research directed at immediate economic outcomes. A vibrant scientifically literate society thrives upon fostering rigorous observation, analysis and synthesis of new ideas. It is through promotion of such habits of dealing with problems that we may derive radical new ways of parsing and understanding the world and the universe about us. We no longer live in a world in which the proven realities are simply those which are tangible on the surface.

We hope you will find useful reading amongst these articles and invite and encourage your responses, comments and suggestions, which may be sent to ronald.young05@gmail.com or to prinfo@src-jamaica.org.

Ronald E. Young

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JAMAICAN JOURNAL OF SCIENCE & TECHNOLOGY, Volume 26 ISSN: 1016-2054

EDITIORIAL PAGE Chief Editor Professor Ronald E. Young Emeritus Professor, c/o School for Graduate Studies & Research Regional Headquarters, University of the West Indies, (UWI) Mona

Editorial Board Deputy Chief Editor Dr. Lawrence Williams Research Consultant, SRC

Professor Wayne McLaughlin Head, Dept of Basic Medical Sciences UWI (Mona)

Professor Tara Dasgupta Chief Editor Emeritus Department of Chemistry, UWI (Mona)

Professor Robert Lancashire Department of Chemistry, UWI (Mona)

Professor Bertram Fraser-Reid President, NPG Industries, Raleigh North Carolina, USA (Formerly Professor of Chemistry, Duke University, Durham, North Carolina) Professor Trevor Jackson Emeritus Professor Lecturer, UWI (St Augustine)

Dr. Monty Patrick Jones Executive Director Forum for Agriculturale Research in Africa, FARA Secretariat Mrs. Swarna Bandara Open Access Consultant, Jamaica Professor Albert Sasoon Senior Consultant UNESCO

Panel of Referees (8) About the Jamaican Journal of Science and Technology (JJST) The Jamaican Journal and Science and Technology (JJST) (ISSN: 1016-2054) is published by the Scientific Research Council of Jamaica (SRC) since 1990. It is a continuation of the Journal of the Scientific Research Council of Jamaica and beginning with this current volume. JJST will publish two issues for the year in electronic format as well as printed format.

SRC as the public sector agency charged with fostering and coordinating of scientific research and promotion of the application of research results in Jamaica, among other activities, also publishes the JJST to achieve its objectives. Therefore, JJST aims to publish scientific papers based on original data on research of interest and relevance to Jamaica, review papers on current scientific and policy issues. It also publishes research notes, research in progress, abstracts of projects and papers, book reviews, current events in science, profiles of scientists and technologists and perspectives. Manuscripts could be submitted in all disciplines of Natural Science, including Geology, Agriculture and Computer Science.

JAMAICAN JOURNAL OF SCIENCE & TECHNOLOGY, Volume 26 ISSN: 1016-2054 |

JJST RESEARCH BRIEFS THE EXPERIENCES OF THE SCIENTIFIC RESEARCH COUNCIL IN PROVIDING COST EFFECTIVE AND SUSTAINABLE WASTEWATER MANAGEMENT OPTIONS Over the last twenty years the agency has focused on providing appropriate technological and economic alternatives to the agricultural, municipal and industrial sectors in order to reduce and treat wastewater.

Pioneering research and developmental activities has been completed in order to adapt anaerobic technology to local conditions in an environmentally friendly and cost effective manner. The paper presents the experiences of the SRC in providing cost effective solutions to waste producers in Jamaica and will enable the delivery of ‘green’ products and services to meet local and international standards.

A REVIEW OF DIBENZYL TRISULFIDE A ANTI-CANCER POLYSULFIDE FROM PETIVERIA ALLIACEA One of the greatest challenges of the modern pharmaceutical industry is the need to find safe broad spectrum anti-cancer agents. Natural products and their derivatives have been a source of major anti-cancer agents; many have toxic side effects which make their application as drugs limited. The paper examines among other things; the binding action of Dibenzyl Trisulfide (DTS) to serum albumin, it’s in vivo anticancer activity and use as a synthetic prototype for the synthesis of broad spectrum cytotoxic agents as well as its mode of action.

POTENTIAL OF TWO JAMAICAN ESSENTIAL OILS AGAINST MOSQUITOES AND BACTERIA The essential oils of Callistemon viminalis (bottlebrush) and Cymbopogon citratus (lemongrass) were investigated to scientifically prove the ethnomedicinal claims of being antibacterial and insecticidal. Bottle brush and lemongrass are two medicinal plants grown in Jamaica and other parts of the world; the former is a small tree native to Australia and the latter a coarse grass and perennial herb cultivated as an ornamental and medicinal plant. The extraction was carried out via steam distillation; the essential oils were tested against three pathogenic bacteria and both showed to be very effective inhibitors of microbes. The insecticidal potential of both oils indicated high repellency.

DIETARY INTAKE OF TRACE AND HEAVY METALS FROM CONSUMPTION OF INSTANT COFFEES ON THE JAMAICAN MARKET Coffee is one of the most popular beverages in the world. Instant coffee is increasingly consumed, with roughly half of the world consumption now being consumed in emerging markets. Eleven brands of instant coffee sold on the Jamaican market were collected and analyzed; thirty-one trace elements were determined using highly specialized apparatus. The results showed that generally, instant coffees contributed a very low percentage of the Provisional Tolerable Weekly Intake for all potentially toxic elements examined; the dietary intake of most essential elements from instant coffees is also negligible; and moderate consumption would contribute some amount of specific trace elements as per the RDI.

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JJST RESEARCH BRIEFS ASSESSMENT OF CADMIUM, MERCURY AND LEAD CONCENTRATIONS IN JAMAICAN CONCH, LOBSTER AND TILAPIA Of the samples examined, tilapia is most consumed on the island. Conch and lobster are largely exported because of their high value; this is also the case in other Caribbean territories. The findings indicate that with relatively low levels of consumption it is unlikely that any of these would contribute significantly to the dietary intake of cadmium, mercury and lead. While the authors know of no cases in which trace metal concentrations have led to human illness in Jamaica; these studies are new to the Caribbean and they suggest that any problems may be location specific and as such, much more data would need to be gathered on the elemental composition of foods. The authors do recommend that there be periodic monitoring of larger ocean fish.

JAMAICAN JOURNAL OF SCIENCE & TECHNOLOGY, Volume 26 ISSN: 1016-2054 |

1. The Experiences Of The Scientific Research Council (SRC) In Providing Cost Effective And Sustainable Wastewater Management Options Julia Brown Scientific Research Council, Kingston, Jamaica, West Indies

ABSTRACT: Jamaica, an island state in the Caribbean, is one of the places in the world that heavily depends on its water resources to sustain the economic livelihood and social well being of its people. Tourism and agriculture are two of the main economic activities from which Jamaica earns most of its foreign exchange. Both sectors are poised for significant growth in the immediate future with the planned development of several thousand new hotel rooms, some of which are already under construction, and an expanded agroparks programme. The latter will include an additional six agroparks. Fishing from the Caribbean Sea, inland artificial aquaculture pond systems, and rivers are other important means of livelihood for many of its residents. These activities, among others, directly impact the water resources of the country.

Among the implications of this great demand is the need to manage these resources in a sustainable, efficient and effective manner to ensure long-term availability for use. There is also an urgent demand to support public and private industry in compliance with national and international environmental regulatory framework.

The availability of local water resources is threatened by environmental pollution caused by the inadequate management of these resources for domestic, agricultural and industrial use and reuse.

The Scientific Research Council (SRC) of Jamaica has, for over the past twenty (20) years, focused on providing appropriate technological and economic alternatives to the agricultural, municipal and industrial sectors, in order to reduce and treat wastewater. The SRC has completed pioneering research and development activities to adapt anaerobic technology to local conditions, in the treatment of wastewater in an environmentally friendly and cost effective manner. The process generates: treated water that is suitable for irrigation purposes, or for discharge to local water bodies; organic fertilizer/soil conditioner; and biogas, which is an alternative source of fuel generated in a closed system from the breakdown of entrained solid organic waste, by indigenous bacteria.. The process generates energy and requires minimal intervention and maintenance, thus reducing operating and maintenance costs.

This paper will present the experiences of the SRC in providing cost effective solutions to waste producers in Jamaica; and will enable the delivery of “green� products and services that will satisfy local and international environmental regulations and standards.

INTRODUCTION Water is the universal solvent and there are very few activities that can be done in nature or man made systems without it. It therefore sustains the terrestrial, marine and riverine ecosystems of the world and is a fundamental input in agricultural, industrial, residential and commercial developments.

In addition to its economic importance water is also vital for health care, hygiene and sanitation and a vast number of human activities. For this reason it is critically important to support sustainable development. Its protection, preservation and conservative use cannot be overemphasized.

Consequent on Climate Change, which is now approaching catastrophic proportions, several economies

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and societies have become more sensitive to the need for a more constant and timely supply of water to meet demand. This has been underscored by deep and protracted droughts and severe flood events.

Caribbean, is poised for significant growth in the immediate future. Invariably this will increase the demand for water to meet these needs. The country's economy is largely dependent on tourism, agriculture, the minerals processing industry and manufacturing.

In many instances this has threatened food security, agricultural production in general , induced desertification In view of the economic and social development trends and severely disrupted the livelihoods of farmers, their existing in Jamaica and the necessity for pollution confamilies and the communities in which they live. trol, wastewater treatment technologies should be sustainable, cost effective and environmentally sound.

One way of managing and sustaining water supply is to These technologies should combine a high efficiency prevent the pollution of ground water stored in our aq- with simplicity in construction and operation and maxuifers and carefully manage them as natural water storage imize the opportunities for efficient removal of pollureservoirs. tants whilst producing useful by-products.

The development and use of efficient, effective and costeffective wastewater management technologies, is among the methods which may be successfully applied for this purpose. The Scientific Research Council of Jamaica, through intensive R&D wastewater management projects has developed state of the art cutting edge, world class technologies in this area. It has earned patents and applied this technology on a wide scale in agriculture, industry, housing, commerce and health care institutions. This goes hand in hand with several benefits while protecting groundwater resources and generating treated effluent of the highest quality. The treated effluent produced may be, recycled and used in various applications. In this regard it must also be borne in mind that for every litre of wastewater which is recycled or reused, one litre of raw water or potable water becomes available for other uses.

In developing countries like Jamaica, there are many competing demands on the limited resources available for development. Waste management, although important for public health, generally gets a lower level of recognition than for example a safe and reliable water supply system. As will be shown in the following chapters, the two are inextricably linked, and the need for efficient and cost effective solutions has become a priority.

Anaerobic technology will be presented as an efficient and cost effective option. The anaerobic technology is not new to Jamaica. It has been in use over two decades as the conventional biogas plants utilising animal manure to generate energy for farm and household purposes.

The technology, however, has advanced to what is now called the Biodigester Septic Tank (BSTTM) for the onsite treatment of domestic sewage in single households, housing complexes and institutions, as well as the Upflow Anaerobic Sludge Blanket (UASB) reactors for the treatment of agro-industrial waste such as coffee and food processing wastewater.

The inadequate treatment and disposal of sewage in the Kingston Metropolitan Area has led to pollution of groundwater resources making it unsuitable for drinking because of its high nitrate content. This has also resulted in the closure of several wells and sequestered several million cubic liters of water.

The SRC has, for over the past twenty (20) years helped several sectors which generate significant quantities of With the use of the SRC wastewater management tech- wastewater to avoid and reduce wastewater generation nologies this could have been avoided. It is therefore through treatment and recycling. These sectors are listed important that this technology be considered among the below. options for treatment of wastewater in these geographic Sectors regions and nationally in the future.

Sugar estates Municipal – Residential, commercial and InstituThe economy of Jamaica, which is a mid-developed tions country (MDC) and Small Island State located in the Agro-proccessors (food processing)

JAMAICAN JOURNAL OF SCIENCE & TECHNOLOGY, Volume 26 ISSN: 1016-2054 | The Experiences Of The Scientific Research Council (SRC) In Providing Cost Effective….

Farms Distillers Dairy

The present “Onsite” systems as well as most of the “offsite” systems of sewage disposal do not provide much in the way of treatment so that most of the potentially polluting influent load escapes into the surrounding COST EFFECTIVE AND SUSTAINABLE ground or surface waters.

WASTE MANAGEMENT SYSTEMS The following factors highlight the need to improve or There are many definitions cited for cost-effectiveness substitute the present practices in the way of waste water but in simple terms as experienced in the waste sector, a treatment and discharge: cost-effective system is one, which includes the following:  The observed negative impact of wastewater discharge on economically important natural resources Low capital cost and ecosystems e.g. aquifers, beaches, offshore waLow operating cost ters and coral reefs Low maintenance cost Short payback period  The risk to health posed by inadequate handling and Offers useful byproduct/s improper discharge of sewage A sustainable waste management system is one, which is  effectively fulfilling the demands of the particular situation in which it is placed. Essentially producing the required results for the design lifespan and beyond.  Why the need for Cost Effective and Sustainable Waste management Systems?

Present Scenario with wastewater treatment in Jamaica



The domestic sewage generated across the island is handled in the following ways (Betts, 1995): 

The negative effect of household sewage on surface and ground waters The present ‘on-site’ systems of sewage disposal e.g. soak away pits do not provide much in way of treatment The high cost of high tech systems such as activated sludge, oxidation ditches -membrane filtration to name a few The potential use of the generated byproducts such as energy and the treated water.

Central Sewerage System » 30 % of the total sewage generated (this figure is due to the fact that Solutions put forward by the Scientific Research the full capacity is not operational in places like Soapber- Council ry. If total capacity is realized the figure will be about 40 %).

The Scientific Research Council (SRC) along with the now Ministry of Science, Technology, Energy and MinPit Latrines - » 30 % of the population ing (MSTEM) has since 1970 been involved in the use of the biodigester system for the digestion of animal waste Soak-away - » 20 % of the population and the subsequent production of biogas as a source of energy. The use of the technology has over the years Septic Tank/Soak away - » 15 % of the population been successfully anchored in Jamaica with the operation of over five hundred (500) systems. Since 1993, and with Other soil absorption systems - » 1 % of the population the increased need for low-cost, on-site treatment and environmentally sound, cost-effective alternatives, the

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biodigester system has been expanded to the use of the BSTTM. There has since been the construction of over two hundred (200) BSTTM systems in single households, housing complexes and institutions. The performance of the BST has been very successful with over 90 % removal of organic matter and other biodegradable solids. The SRC has also embarked on the implementation of the Upflow Anaerobic Sludge Blanket (UASB) reactor for the treatment of Agro-industrial wastewater.

idly in popularity worldwide (Lettinga et al., 1988) and as a result of its cost-competitiveness, has evolved into a mature technology for waste treatment (Pfeiter et al., 1986). With the present state of technological development and basic insight into the process, only a few of the presumed drawbacks such as the slow rate of bacterial activity remain, while all its principal benefits such as the production of energy and the low production of sludge over conventional aerobic methods are still relevant (Lettinga, 1995; Lettinga et al., 1988; Lettinga, 1996)

ANAEROBIC TECHNOLOGY Digestion Process The use of technology for the anaerobic treatment of waste water has been in existence for decades. According The anaerobic digestion process is done by five major to McCarty (1981) it has existed as a technology for over groups of putrefactive bacteria, which break down the one hundred years. Anaerobic treatment processes re- organic material under anoxic conditions. These are: quire no mechanical energy as input to facilitate or mainfermentative bacteria; tain the raw biological processes, and require little hydrogen-producing acetogenic bacteria; maintenance and attention (Schellinkhout et al., 1985). hydrogen consuming acetogenic bacteria; This has favoured the development of anaerobic syscarbon dioxide-reducing methanogens; tems, even as similar aerobic systems have been subject aceticlastic methanogens to increasing cost of energy; along with the increasing cost of operation and maintenance (Schellinkhout et al., 1985). Anaerobic digestion processes occur naturally in many places where organic material is available and either where anoxic conditions persist or the electrochemical reduction of oxygen is not favoured. Such places include the stomach of ruminants, in marshes, sediments of lakes and ditches, municipal landfills and also sewers (Alaerts et al., 1990).

For a long time these anaerobic processes have been used for the stabilization of wastes and for the production of methane, as a valuable source of energy (Alaerts et al., 1990). Votchen et al., (1987)observed that the utilization of the anaerobic technology gradually evolved from an airtight cesspool and septic tank to a temperature controlled, completely mixed digester and finally to a high rate reactor containing a large quantity of highly active biomass.

Anaerobic processes can be profitably applied for all types of waste that are organic in nature. Successful fullscale facilities have been constructed and operated for the treatment of dilute wastewater such as municipal sewage and for very concentrated effluents such as rum stillage (dunder). Anaerobic treatment has increased rap-

Figure 1 Shows a schematic illustration of the bacterial activities on the various processes of anaerobic degradation. Schematic Outline of BST

JAMAICAN JOURNAL OF SCIENCE & TECHNOLOGY, Volume 26 ISSN: 1016-2054 | The Experiences Of The Scientific Research Council (SRC) In Providing Cost Effective‌.

Biopolymers like proteins, carbohydrates, nucleic acids and fats contained in agro-industrial waste/wastewaters are first hydrolyzed to mono- and oligo-mers, and then fermented to products which can be used by methanogens directly (acetate, hydrogen, formate) and to reduced organic compounds such as propionate, butyrate, longchain fatty acids, alcohols, lactate and succinate (Gujer & Zehnder 1983). Acetogenic bacteria oxidize higher fatty acids anaerobically to acetate, CO2, H2 and formate (Gujer & Zehnder 1983). Acetogenic bacteria can also oxidize lactate and ethanol to acetate, but fermentation of these compounds to e.g. acetate and propionate is also possible (Laanbroek et al. 1982). Succinate can be decarboxylated to propionate (Schink & Pfennig 1982).

Acetate, formate and H2 are converted by methanogens to methane and CO2 (Schink 1997). Volatile fatty acids are known to be important intermediates in the degradation of organic matter under methanogenic conditions.

About 70% of the reducing equivalents formed in the anaerobic digestion process are transferred via acetate to methane (Gujer & Zehnder 1983). Since there are 4 moles of H2, but only 1 mole of acetate is required to produce 1 mole of CH4, the contribution of H2 to methanogenesis during anaerobic degradation of carbohydrates can maximally be 33% of the total CH4 formed (Conrad 1999). Propionate and butyrate are key intermediates in the mineralization of complex organic matter.

The complete oxidation of propionate and butyrate can account for 20 to 43% of the total methane formation, depending on the type of digester and the nature of the organic compounds (Mackie & Bryant 1981). Four different phases can be distinguished in the overall conversion process; these are Hydrolysis, Acidogenesis, Acetogenesis and Methanogenesis.

Hydrolysis

alcohol, carbon dioxide, hydrogen and ammonia.

Acetogenesis The short-chain fatty acids (other than acetate) are converted to acetate, hydrogen gas and carbon dioxide.

Methanogenesis Methanogenesis occurs from carbon dioxide reduction by hydrogen and from decarboxylation of acetate to produce methane.

The biogas produced in anaerobic digestion consists mainly of methane (CH4, 60 - 70 %), carbon dioxide (CO2, 30 - 40 %), and traces of hydrogen sulphide (H2S, 0.5 - 1 %).

Advantages of anaerobic digestion The advantages of anaerobic treatment include: Low production of biological sludge High treatment efficiency Low capital cost No oxygen requirement Methane production (potential source of fuel) Low nutrient requirement Low operating cost One of the major reasons for the promotion of the anaerobic technology for the treatment of waste is the benefit it offers over the conventional treatment systems.

These benefits are outlined as follows: The method is simple in construction and operation, and consequently less expensive

First, complex polymeric materials such as polysaccharides, proteins and lipids (fats and grease) are hydrolysed by extracellular enzymes to soluble products of a size small enough to allow their transport across cell membranes.

The method generally does not require mechanical energy to facilitate or maintain the biological process

Acidogenesis

This decentralised application dispenses with the need for a sewerage network, leading to significant savings

The method can be applied at small and at large scales, facilitating a decentralised application

These relatively simple, soluble compounds are fermented or anaerobically oxidised to short-chain fatty acids, In the case of the BST there is no sludge development

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due to complete degradation of organic solids within the The Biodigester Septic Tank (BST) system The BST is an on-site sanitation unit, which provides for One of the major drawbacks of the anaerobic process the disposal of toilet (black) wastewater as well as of for the treatment of waste is the fact that the method kitchen and bathroom sullage (grey water). It should be cannot accomplish complete removal of faecal coliforms stressed that the BST is not an alternative or a replaceand nutrients (nitrogen and phosphorus) and therefore ment of the centralised treatment systems but for the as is the case with the aerobic processes, tertiary treat- present ineffective on-site methods of disposal such as ment is required. The tertiary or advanced treatment is soak-away pits. responsible for the removal of the nutrients and bacteria as well as the remaining organic matter from the anaero- Figure 2 show a schematic representation of a BSTTM bic process which is the secondary stage of the treatment process.

The use of the BST has increased in importance due to the following reasons: SYSTEMS ADAPTED AND OPTIMIZED The negative impact of wastewater discharges on ecoThe systems adapted, patented (as is the case with the nomically important natural resources and ecosystems BSTTM ) and being propagated by the SRC for the use of waste management in Jamaica are as follows: The risk to health (especially of young children) posed by inadequate management of sewage.  The Biodigester Household sewage causes serious pollution of surface  The Biodigester Septic Tank and ground waters and ultimately coastal water..  The Upflow Anaerobic Sludge Blanket reactor  The Bio-latrine The present ‘on-site’ systems of sewage disposal do not provide much in the way of treatment – most of the inThe Biodigester and the Biodigester Septic Tank fluent’s polluting load escapes into surrounding ground and surface waters.

The Biodigester System The biodigester system is a concrete tank (see Figure 2) which is located below the ground. The gas produced comes off at the top of the tank via pipelines, while the residual waste comes out automatically and flows into another tank (compensation tank). From this second tank the bio-fertilizer can be dried or sent as a slurry directly to crops or to the soil. The liquid from the drying beds is sent to a polishing pond to clean the water for irrigation purposes.

The high cost to operate and maintain high-tech systems (activated sludge, oxidation ditches etc.)

The generation of biogas which can be treated for use as a replacement for liquid petroleum gas (LPG) and firewood hence reducing fossil fuel use and deforestation.

The general need for low cost environmentally friendly alternatives.

The biodigester system can be used to treat the following Upflow Anaerobic Sludge Blanket Reactor types of organic waste: The UASB is a high rate anaerobic system used for the treatment of different types of wastewater. The system Dung from cattle, pigs, goats and other livestock was developed in the Netherlands in the early seventies.

Vegetable matter, Green plants and plant waste The UASB process is characterised by an active sludge Agro-industrial waste and wastewater blanket/bed at the bottom of the reactor that degrades the incoming wastewater. The bacteria may spontane-

JAMAICAN JOURNAL OF SCIENCE & TECHNOLOGY, Volume 26 ISSN: 1016-2054 | The Experiences Of The Scientific Research Council (SRC) In Providing Cost Effective….

Figure 2

A Schematic Outline of BST

ously agglomerate to form granules. These granules have good settling properties and are not susceptible to being washed from the system under practical conditions. Retention of active sludge, whether granular or flocculent within the UASB reactor enables good treatment at high organic loading rate. The maintenance of a high sludge concentration in the reactor is one of the most important conditions of a UASB process.

entering the settling zone, where they create turbulence and consequently hinder the settling of sludge particles.

Some characteristics of the UASB that make it popular and so widely used are listed below.

High organic loading rates Short hydraulic retention time Low energy demand A schematic representation of the UASB reactor is No need for support media shown below in Figure 3. The most characteristic device Simple reactor construction of the UASB reactor is the phase separator. This device Substantial full scale experience is placed at the top of the reactor and divides it into a lower part, the digestion zone, and an upper part, the By-products of the Anaerobic Process settling zone. The presence of a settler on top of the digestion zone enables the system to maintain a large There are three major by-products of the anaerobic prosludge mass in the reactor, while discharging an effluent cess – biogas, nutrient rich effluent and a stabilised basically free of suspended solids. The biogas bubbles sludge. However in the BST there are only two (biogas rise up to the liquid–gas interface under the phase sepa- and effluent) since there is almost complete degradation rator. This interface may be at the same level as the wa- of solid, as was previously stated, in a BSTTM system. ter–air interface in the settler or at some lower level if a hydraulic seal pressurises the biogas. Sludge flocs with Biogas adhering gas bubbles may rise up to the interface in the gas collector, but will settle when the gas bubbles are Biogas consists of about 60 % methane (CH4) and 40 % released to the gas phase at the interface. Baffles, placed carbon dioxide (CO2). It also contains small amounts of beneath the apertures of the gas collector units, operate other substances, including up to 1 % hydrogen sulphide as gas deflectors and prevent the biogas bubbles from (H2S).

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Figure 3

Typical cross section of conventional UASB reactor showing sludge blanket/Bed and Gas Liquid - Solids separation (van Haandel and Lettinga, 1994)

It is lighter than air and has an ignition temperature of ap- Consumption of Biogas proximately 700oC which is higher than diesel oil (350 oC) or petrol and propane (about 500oC). The temperature of Table 2 outlines the amount of biogas that is consumed by the flame is 870oC. various types of equipment.

Biogas Utilisation

Table 2

Biogas can be used in the same way as any other combustible gas. The biogas can be used in a modified gas stove, refrigerator, water heater, diesel engine, gas lamps and indeed any equipment that uses liquid petroleum gas (LPG).

The biogas can be piped to the various pieces of equipment via PVC pipes, used as gas lines. The gas cannot be economically liquefied. The calorific value of biogas is about 6 kWh/m3. This corresponds to about a litre of diesel oil.

Biogas has a very high level of efficiency as outlined below in Table 1.

Table 1: Biogas for cooking (practical values from India - Sasse, 1988).

Amount Cooked

Time (min)

Gas (L)

1 L water

10

40

5 L water

35

165

500 g rice

30

140

1000 g rice

37

175

350 g pulses

60

270

700 g pulses

70

315

Utilization and Consumption of Biogas Equipment

Amount of Biogas

Household burners

200 - 450 L/h

Industrial burners

1000 - 3000 L/h

Refrigerator 100 L depending on outside temperature

30 - 75 L/h

Gas lamp, equiv. to 60 W bulb

120 150 L/h

Biogas/diesel engine per bhp

420 L/h

Generation of 1kwh of electricity with biogas/diesel mixture

700 L/h

The major disadvantage in the use of biogas is the presence of hydrogen sulphide which has a very pungent odour. The Hydrogen Sulphide combines with water vapour to form corrosive acids. Water heating appliances, utensils and re-

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frigerators are particularly at risk. However biogas can be rid of the sulphur using iron oxide filters based upon the following reactions: NB: If gas is not used it should always be flared because non-combusted methane is harmful to the environment (it is a potent greenhouse gas).

The Public Health Act addresses the perspective of human health while the NRCA Act focuses on the natural environment as a resource. The NRCA Act has significant powers related to the management of the environment, and specifically for the regulation of effluent discharges (Sections 9(4) and 12). The National Water Commission (NWC) Act of 1980 delegates to the NWC the responsibility for public water supply systems and public sewerage and sewage treatment. The Water Resources Act was established to facilitate the establishment of the now Water Resources Authority whose responsibility is to regulate, control and conserve Jamaica’s local water resources.

Regulations/Standards

Effluent The effluent from the anaerobic process is rich in nutrient and an excellent source of liquid fertilizer. However, if it is to be discharged or utilized for irrigation purposes, it must be further treated to reduce the high nutrient content and to remove harmful bacteria. Tertiary treatment which utilizes natural systems such as a constructed wetland, or a chemical based system such as Dissolved Air Flotation (DAF), may be employed to remove the bacteria and nutrients. This is done in order to comply with the National Sewage Effluent Discharge Standards as well as the Trade Effluent Standards for safe reuse.

The NEPA since April 2015 has released the newly enacted Sludge and Sewage Regulations which works in tandem with the National Sewage Effluent Discharge Standard (see table 5 below) as well as the Trade Effluent Discharge Standards.

The Public Health Act in Section 14, allows the Minister to make regulation s in relation to air, soil and water pollution. In Section 7(p) it also allows the Local Board of Health to make regulations for the sanitary collection and disposal of solid waste and any other waste matter.

The National Water Commission has adapted various regulations under the National Water Commission Act, and is mainly concerned with the setting and collection of charges The sludge produced by the anaerobic process is mineral- for water supply and sewerage services. ized/stabilized biomass and is fit for reuse as fertilizer and soil conditioner.

Table 3 - National Sewage Discharge Standards Sludge

POLICIES AND ACTIVITIES IMPLEMENTED TO ADDRESS WASTEWATER IN JAMAICA Legislation and Regulation With regards to wastewater management the most important statutes are: The National Resources Conservation Authority (NRCA) Act, 1991 The Public Health Act 1974, amended in1985 The National Water Commission Act, 1963, amended in 1965, 1973 and 1980 The Water Resources Act, 1995

PARAMETERS

EFFLUENT LIMIT

BOD5

20 mg/L

COD

100 mg/L

TSS

20 mg/L

Total Nitrogen

10 mg/L

Total Phosphates

4 mg/L

pH

6-9

Faecal Coliform

200 MPN/100 ml

Residual Chlorine

1.5 mg/L

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Government Policy

NWC, and the strong involvement of Environmental Non-Governmental Organizations (ENGO) and ComThe NEPA (then NRCA) and the Planning Institute of munity Based Organizations (CBO).

Jamaica (PIOJ), with technical assistance from the World Bank, have jointly published the ‘Jamaica National Envi- Relevance of policies and activities to Small Island ronmental Action Plan’ (JANEAP - 1995). The stated States purpose of JANEAP is: The Small Island Developing States (SIDS) are all very ‘to document the major environmental problems facing the country similar in many respects. Their commonalities generally and to formulate the appropriate policy framework, institutional include: arrangements, legal instruments, strategies, programmes and projects to address and mitigate these problems’.  Climatic conditions  Dependence upon tourism This document is significant because it explicitly recog-  Beaches nizes the need to pursue the goal of sustainable develop-  Inadequate infrastructure ment and the role which ‘the Polluter Pays’ principle  Cultural practices must play in order to achieve that goal. It contains the  Inadequate waste treatment and disposal practices commitment that the Government must establish (?) standards for trade effluent, sewage effluent, ambient These areas among others are very similar for most if water quality, potable water, irrigation water and recrea- not all of the SIDS therefore enhanced wastewater treattional water (pool and beaches). The JANEAP is updat- ment solutions, sewage regulations as well as sewage and ed annually. trade effluent standards are very important for survival and sustainable development of key industries such as Institutional and Organizational Arrangement for tourism as well as improving well being and health of the Wastewater Management people.

With regards to wastewater management, the following BENEFITS AND COSTS: ECONOMICAL, ENagencies play a significant role: VIRONMENTAL AND SOCIAL OUTCOMES RESULTING FROM THE TREATMENT  the National Environment and Planning Agency MEASURES IMPLEMENTED  the Environment Health Unit of the Ministry of Health Economic Implications  the National Water Commission  the Water Resources Authority The anaerobic technology being propagated and implemented by the SRC for the treatment of wastewater has Other national public agencies having involvement in significant economic benefits. This is because the prowastewater management are: cess produces two useful by-products:    

The Scientific Research Council The Urban Development Corporation National Housing Trust Ministry of Agriculture

 

biogas for energy generation, and treated nutrient rich effluent, suitable for irrigation.

Providing wastewater solutions will minimize the impact on existing water resources, and will preserve its potenLocal involvement in wastewater management has im- tial for use for economically important activities such as proved significantly with the establishment of bodies and tourism and recreational sports and for domestic such as the Northcoast Wastewater District by the purposes.

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Environmental Implications The main impacts of inadequate wastewater management in Jamaica are those associated with the pollution and degradation of the natural environment and particular surface and ground water resources.

The management of water in all its manifestations is among the biggest environmental challenges in Jamaica today. According to the World Bank (Jamaica - Economic Issues for Environmental Management, 1993):



The pollution of ground water. Ground water provides approximately 80 % of the potable water in Jamaica, and is extensively used in industry and agriculture. The sources of ground water pollution are principally: agro-industries (especially from sugar factories and distilleries), sewage (especially from soakaway pits), leachate (from waste dumps sites) saline intrusion (mainly from over-pumping), and the bauxite/alumina industry. The Liguanea aquifer is monitored by the UWI and the WRA and has been shown to be acutely affected.

“Jamaica’s largest problems, from the point of view of affecting the  The pollution of Kingston Harbour by untreated (or largest number of people’s lives and livelihoods, are related to water. poorly treated) sewage. It is also affected by polluPollution of surface and seawaters threatens human health and tants carried by surface watercourses entering the tourism revenues. Clean water for domestic purposes has frequently harbour. The pollution from Kingston Harbour is been short in Kingston Metropolitan Area (KMA) and is presentthought to affect part of the south coast of the isly facing a significant deficit. Due to degradation of watersheds, land, even as far as Negril. Water Qaulity monitoring flooding is becoming more frequent and more severe, and pollution data has indicated that the continued pollution of of ground water is raising cost of infrastructure, eroding agricultural the harbour has caused the persistence of eutrophic productivity and posing health risks. Topsoil loss in the watershed conditions which has lead to the growth of algal areas is harming agricultural productivity and causing marine siltablooms and consequent fish kills. tion. Although a number of other environmental problems do exist, the water issue is by far the most serious. It must be recognized that  With all island countries, there is close interaction action is required on a number of fronts and is likely to be of fairly between terrestrial and marine ecosystems, and Jalong duration, with significant improvements only likely in the late maica is no exception. Some of the coral reefs have 1990s. Realizing that these problems will be solved in short order been badly affected by land based sources of polluimplies making interim arrangements as well as planning for pertion, as have some of the fisheries. manent and sustainable solutions”.

The effects of water pollution are now being appreciated Interestingly however, a significant number of Jamaicans by an increasing number of Jamaicans who have recogregard solid waste management as a higher priority for nized the connection between pollution and the well action than wastewater management and water pollution being and economic prosperity. The implementation of control. This is likely due to the fact that solid waste is a the cost effective and sustainable options can reduce the more visible problem than water pollution, and appears pollution caused by soakaway pits and ineffective to touch the average Jamaican’s daily life more directly. wastewater treatment systems.

Betts(1995) identifies sewage as the single largest source Social Implications of water pollution. The effects of water pollution can found throughout Jamaica. Some of these effects in- The health hazards as well as the environmental degradaclude: tion created by improper disposal and handling of wastewater, has the potential to result in a decrease in  The contamination of most surface waters. The Rio quality of life for Jamaicans especially in the lower inCobre, for example, which flows through Spanish come strata and informal settlements.

Town and discharges into the Kingston Harbour, is monitored the UWI and has been demonstrated to There is a decrease in economic opportunity and therebe polluted. fore the quality of life that is created by the pollution of

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the receiving water bodies as well as the possibility of exposure to gastro instentinal and some skin diseases by  people living in areas with improper waste disposal systems (ref., PIOJ).

CONCLUSIONS

viduals; Recognizing that wastewater is a resource and the treatment should form a link to the water company and major industrial and agricultural consumers including major electricity generating companies (wastewater treatment systems paying for itself); Having legislation in place and the systems to enforce it;

The anaerobic/biogas technology has become a proven cost-effective and sustainable alternative for wastewater Having regulations enforced; treatment in Jamaica. It is anticipated that the technology will: Having the commitment of the public and private sector to wastewater management best practices  Improve hygiene and protect the environment Overcoming resistance to new technologies  Avoid pollution of ground water resources  Provide a supplementary supply of fuel for cooking, Lessons Learnt refrigeration, lighting, heating and in diesel engine  Provide effluent for irrigation purposes and reduce the quantity of potable water used for irrigation pur- Some of the lessons learnt in trying to develop and implement cost effective and sustainable wastewater manposes  Reduce the impact of wastewater effluent on beach- agement in Jamaica areas follows: es and will protect the resources for local coastal  The approach must be to engage all major organizarecreational activities and tourism tions and individuals in the process  Provide a system with lower maintenance and operating costs than similar aerobic systems.  Legislation and regulations should be clear and focussed Challenges and Lessons Learnt Challenges



The challenges which demand an integrated wastewater  management framework for Jamaica are as follows: 

Legislation and regulations should be enforced New technologies and processes are not easily implemented. Every effort should be made by the government to facilitate their utilization

Recognition of the need for cost effective and sustainable approaches to wastewater management by key public and private sector organizations and indi-

REFERENCES 1.

McCarty, P.L. (1981). One hundred years of anaerobic treatment. In: Hughes, D.E. et al. Eds. Anaerobic Digestion 1981. Amsterdam. Elsevier. 1981, 3.

2.

Schellinkhowl et al 1988

3.

Alaerts, G.J., Veenstra, S., Bentvelsen, M., van Duijl, L.A. et al.. (1990). Feasibility of anaerobic sewage treatment in sanitation strategies in developing countries. Delft, The Netherlands. International Institute for Hydraulic and Environmental Engineering. IHE Report Series 20, 130p.

JAMAICAN JOURNAL OF SCIENCE & TECHNOLOGY, Volume 26 ISSN: 1016-2054 | The Experiences Of The Scientific Research Council (SRC) In Providing Cost Effective‌.

4.

Votchten, P., Schowanek, D. & Verstraete, W. (1988). Aerobic vs. anaerobic wastewater treatment. Proceedings of the 5th International Symposium on Anaerobic Digestion. Bologna, Italy. Hall ER, and Hobson PN, Eds. P91 - 103.

5.

Lettinga, G., de Man, A.W.A., and van der Last, A.R.M. (1988). The use of the EGSB and UASB anaerobic systems for low strength soluble and complex at temperature ranging from 8 to 30oC. Proc. Of the 5th International Symposium on Anaerobic Digestion. Bologna, Italy. Hall ER, and Hobson PN, Eds. P197 - 208.

6.

Pfeiter et al 1986.

7.

Lettinga, G (1995). Anaerobic digestion and wastewater treatment systems. Antoine van Leeuwenhoek. 67:3 - 28.

8.

Lettinga, G (1996). Sustainable integrated biological wastewater treatment. Water Science and Technology, 33 (3): 85 – 98.

9.

Conrad, R. (1999): Contribution of hydrogen to methane production and control of hydrogen concentration in methanogenic soils and sediments. FEMS Microbiol. Ecol. 28: 193-202.

10.

Gujer W & Zehnder AJB (1983): Conversion processes in anaerobic digestion. Wat. Sci. Technol. 15: 127-167.

11.

Laanbroek HJ, Abee T & Voogd IL (1982): Alcohol conversions by Desulfobulbus propionicus Lindhorst in the presence and absence of sulphate and hydrogen. Arch. Microbiol. 133: 178-184.

12.

Schink B & Pfennig N (1982) Propionigenium modestum gen. nov., sp. nov., a new strictly anaerobic nonsporing bacterium growing on succinate. Arch. Microbiol. 133: 209-216.

13.

Schink B (1997) Energetics of syntrophic cooperation in methanogenic degradation. Microbiol. Mol. Biol. Rev. 61: 262-280.

14.

Van Haadell and Lettinga 1994.

15.

Sasse, 1988.

16.

The National Resource Conservation Authority Act, 1991.

17.

The Public Health Act, 1985.

18.

The National Water Commission Act, 1980

19.

The Water Resources Act, 1995.

20.

Jamaica - Economic Issues for Environmental Management, World Bank Country Study, 1993.

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JAMAICAN JOURNAL OF SCIENCE & TECHNOLOGY, Volume 26 ISSN: 1016-2054 A Review Of Dibenzyl Trisulfide An Anti-Cancer...

2. A Review Of Dibenzyl Trisulfide An Anti-Cancer Polysulfide Isolated From Petiveria alliacea Lawrence A.D. Williams1, Harald Rosner2 and Wolfgang Kraus3 1. The Natural Products Research Unit, Scientific Research Council, Kingston 6, Jamaica, W.I. 2. Institute of Zoology, University of Hohenheim, Garbenstrasse, Stuttgart, Germany.

3. Institute of Bio-organic Chemistry, University of Hohenheim, Garbenstrasse, Stuttgart, Germany.

ABSTRACT: Dibenzyl trisulfide (DTS) is a broad-spectrum anti-cancer agent with IC50 values ranging from 0.34 µM to 18.84 µM on cancer cells. DTS is weakly cytotoxic on non-cancerous cells e.g. the HOFA fibroblast cell line. The cytotoxic activity of DTS is increased by 2400 fold when it is conjugated to albumin on the SH-SY5Y neuroblastoma cells in vitro. The binding of DTS to albumin is via the tyrosine amino acid residues in the protein.

The mode of action elucidated for DTS is an attenuator of the dephosphorylation of tyrosyl residues of mitogenactivated protein, extracellular regulated kinases 1 and 2 (MAP kinases ERK 1 and ERK 2), with selective disruption activity on microtubules.

INTRODUCTION The need to find safe broad spectrum anti-cancer agents is one of the greatest challenges of the modern pharmaceutical industry. Natural products (plant secondary metabolites) and their derivatives have been a source of major anti-cancer agents, examples include vinblastin, vincristine and the toxoids.1 However, these natural products anti-cancer agents have been associated with a wide range of toxic side effects which make their application as drugs limited. One compound which has shown promise in recent times as an anti-cancer agent and drug prototype is dibenzyl trisulfide (DTS).2,3 (Figure 1)

degenerative diseases treatment based on mode of action studies done on the molecule.4 Thus, DTS seems to be a poly-pharmaceutical agent, i.e. one drug treating many diseases with very few side effects.5 The search for polypharmaceutical agents is one of the major foci of leading drug companies today.

The Broad-Spectrum In Vitro Anti-Cancer Activity of Dibenzyl Trisulfide (DTS)

Dibenzyl trisulfide (DTS) has been shown to inhibit the proliferation of a broad-spectrum of cancer cell lines.

These cell lines include IC50 values in µM are in parentheses, IC50 value is the concentration required for inhibiting the proliferation of cancer cells by 50% relative FIGURE 1 Structure of Dibenzyl Trisulfide to the control); Jurkat leukaemia (0.35) , Ovarian (A2780) (0.40), Ovarian (OVCAR4) (1.40), Fibrosarcoma (HT 1080) (1.90), Non-small cell lung (H460) (5.10), Breast (MCF7) (6.60), Breast (M231) (2.40), Adenocarcinoma (HeLa) (2.50), Neuroblastoma (SH-SY5Y) (0.43), Melanoma (IPC) (2.90), Primary bladder carcinoma (18.84), Miapaca (pancreatic cancer) (0.34), Prostate (DU145) (0.59) and PC-3 (0.63), Small lung cell carcinoma (A549) (0.84), Breast cancer MDA-MB-231 (0.38).6,3,7,8 The IC50 values were determined according to the methodologies used by An et al3, Williams et al6 Dibenzyl trisulfide not only has anti-cancer activity, but and Lowe et al8 the compound seems to be interesting as an anti-

JAMAICAN JOURNAL OF SCIENCE & TECHNOLOGY, Volume 26 ISSN: 1016-2054 | A Review Of Dibenzyl Trisulfide A Anti-Cancer...

Williams et al.6 revealed that the non-cancerous fibroblast HOFA cell line treated with DTS at concentration > 8.9µM did not detach from the extracellular matrix and proliferated for the duration of the experiments over seven days. After seven days in culture DTS had minuscule anti-proliferative/cytotoxic effect on the noncancerous HOFA fibroblasts. Thus, the culturing period was extended for another seven days and the antiproliferative action of DTS re-examined on the fibroblast revealing a 30 % inhibition at 8.9µM. DTS caused dramatic inhibition of the proliferation of cancer cells at 72 - 96 hours at doses lower that 8.9µM, except for the primary bladder carcinoma. This suggests that DTS is more active on pathological cells than on healthy cells.

Serum Albumin-Binding Cytotoxic Enhancement Effects of Dibenzyl Trisulfide (DTS) In vitro experiments revealed that DTS has a strong affinity for serum albumin e.g. bovine serum albumin or human serum albumin, and when bound, the cytotoxic activity is increased by 2400 fold on SH-SY5Y neuroblastoma cells (Figure 2).9 Thus, we are of the view that when DTS is absorbed from the intestine and bound to human serum albumin, anti-cancer activity should be increased significantly, and this could explain the potent anti-cancer activity of the molecule in vivo.

In vivo Anti-Cancer Activity of Dibenzyl Trisulfide (DTS) In the mouse Lewis lung cancer model DTS showed an average tumour inhibition rate of 67.05%, 51.34% and 45.21% for doses of 50, 25 and 12.5 mg/kg body wt (relative to the normal saline controls).10 For tumour inhibition in the mouse sarcoma S180 model DTS gave inhibition rate of 63.30%, 54.68% and 48.69% at doses of 50, 25 and 12.5 mg/kg body wt. (relative to the normal saline controls).10

FIGURE 2 ID NMR interaction of BSA with DTS in deuterium oxide (D20)-phosphate buffer saline pH 6.8 (1:9, v/v). (A) DTS (2.8 mM) in BSA (25 mg/ml), new signals at 3.4 ppm (*) and 6.66.8 ppm (**); (B) BSA (25 mg/ml); (C) carrier solvent (D20-phosphate buffer saline) (1:9, v/v) with 10% (v/v) methanol; (D) 1D NMR profile of 2.8 mM DTS in deuterium oxide-phosphate buffer saline (Ph 6.8) (1:9, v/v), with 10% (v/v) methanol

safety profile on the following cancer cell lines at micromolar doses: PC3 (prostate cancer), Bcap-37 (breast cancer), NCI-69 AR (small cell lung carcinoma), SKOV3 (ovarian cancer), SH-SY5Y (neuroblastoma), A549 (nonsmall cell lung carcinoma), MV522 (non-small cell lung carcinoma), HT29 (colon cancer) and Panc-1 (pancreatic cancer).10 Fluorapacin is now in Phase II clinical trial in Dibenzyl Trisulfide as a Synthetic Prototype for the China for a wide range of cancers.11 Synthesis of Broad-Spectrum Cytotoxic Agents Binding of Dibenzyl Trisulfide (DTS) to Tyrosine An et al.3 have reported on several bisaryl trisulfide derivatives of DTS with potent cellular anti-tumour activi- It is now accepted that malfunction of receptor tyrosine ty. The synthetic derivative of DTS, bis(4-fluorobenzyl) kinases (RTKs) regulate various diseases such as cancer, trisulfide (BFBTS) also known as Fluorapacin, has cell death (apoptosis), developmental effects, ageing and demonstrated good anti-cancer activity with excellent diabetes.13 Dibenzyl trisulfide (DTS) has a strong bind-

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FIGURE 3 Effects of 1 µM DTS on the expression of tubulin (a), GAP-43 (b), and bFGF (100 ng/ml)induced tyrosine phosphorylation (c-f) in SH-SY5Y neuroblastoma cells. (a) Lanes: 1, control; 2, DTS, 2 days; 3, DTS, 4 days; 4 DTS, days, then without DTS, 2 days; 5, DTS, 4 days, then without DTS, 4 days; 6, DTS, 4 days, then without DTS, 7 days, (b) Lanes: 1, control; 2, bFGF, 4 days; 3, DTS, 4days; 4, bFGF plus DTS, 4 days. (c) Pattern of tyrosine phosphorylation detected with mAb PY99 (see Section 2). Lanes: 1, control; 2, bFGF, 5 min; 3, DTS, 15 min, then bFGF, 5 min; 4, DTS, 5 min. (d) Tyrpsine phosphorylation of MAP kinase (erk1/erk2). Lanes: 1, control; 2, DTS, 5 min; 3, bFGF, 5 min; 4, DTS, 2 h, then bFGF, 5 min. € Tyrosine phosphorylation of MAP kinase (erk1/erk2). Lanes: 1, control; 2, bFGF, 5 min; 3, bFGF, 30 min; 4, bFGF, 90 min; 5, DTS, 2 h, then bFGF, 90 min. (f) Percentages of the signals of bands 1 and 3-7 € as compared to band 2 (bFGF, 5 min = 100%). Lane 8, DTS, 2 h. ing affinity to tyrosine amino acid residues as revealed by Rosner et al.2 and Williams et al.12 The above studies were conducted using bovine serum albumin and tyrosine amino acid in one-dimensional NMR experiments (Figure 2). The binding of DTS to tyrosine amino acid residues was one of the first indicators and, along with its kinases signaling properties, suggested that DTS could be a very interesting biologically active molecule.

selective disruption activity on microtubules (Figure 3).

Rosner et al.2 have revealed that DTS has no effects on actin or the molecular biological processes such as growth-associated protein-43 (GAP-43) which regulate actin polymerization (Figure 4). However, DTS strikingly disassembles microtubules, which are dynamic polymers of α-tubulin and β-tubulin heterodimers and are important in cellular process such as mitosis (Figure 5).2 Staining SH-SY5Y neuroblastoma cells with the m-phase Mode of Action of Dibenzyl Trisulfide (DTS) -specific marker KE 67 after DTS treatment resulted in an m-phase-synchronized arrest of cells. 12 The DTS Rosner et al.2 elucidated the mode of action of DTS as derivative BFBTS was also found to be an anti-mitotic an attenuator of the dephosphorylation of tyrosyl resi- agent inducing mitotic arrest.10 dues of mitogen-activated protein, extracellular regulated kinases 1 and 2 (MAP kinases ERK 1 and ERK 2), with

JAMAICAN JOURNAL OF SCIENCE & TECHNOLOGY, Volume 26 ISSN: 1016-2054 | A Review Of Dibenzyl Trisulfide An Anti-Cancer...

FIGURE 4 No effect of DTS on the expression and redistribution of F-actin in SH-SY5Y neuroblastoma cells. Cells were cultured in the presence of 1 µM DTS for 1 day, cholinergically stimulated to form lamellar protrusions for 6 min, fixed, and F-actin stained with phalloidin-TRITC. No effect of DTS either on F-actin stress fibres (a,b) or on the polymerization of F-actin at the ‘leading edges’ of cholinergically induced lamellar protrusion (c,d).

FIGURE 5 Reversible disassembly of microtubules in Wistar 38 human lung fibroblasts (a-c) and SH -SY5Y neuroblastoma cells (d-f) cultured in the presence of 1 µM DTS for 1 day. (a,d) Controls; (b,e,e,) DTS-treated cells; (f) reappearance of microtubules in a SH-SY5Y cell 1 day after removing DTS.

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Non-toxic Nature of Dibenzyl Trisulfide (DTS) in Mice DTS when injected at 11 - 34 mg/kg body wt and 10 50 mg/kg body wt in mice shows no toxic effects.14, 10 The above doses are effective anti-cancer doses in animals. In fact, a dose of 11 mg/kg body wt stimulates bone marrow activity and the proliferation of thymic cells in aged mice by about 50 % both effects being

important cancer chemotherapy parameters.14 In addition, the conjugate of DTS and serum albumin when injected intravenously shows no toxic effects in mice.

This is important as the conjugate represents an enhancement in the anticancer activity of DTS. Presently, this is a new area of research dealing with anti-cancer agents and so there is no new data available. In addition, not many anti-cancer agents show strong affinity to bind to albumin.

REFERENCES 1.

Hamburger, M., Marston, A. and Hostettmann, K. Search for New Drugs of Plant Origin, Advance in Drug Research. Academic Press 1991, 20: 167 - 215.

2.

Rosner, H., Williams, L.A.D., Jung, A. and Kraus, W. Disassembly of Microtubules and Inhibition of Neurite Outgrowth, Neuroblastoma Cell Proliferation, and MAP Kinase Tyrosine Dephosphorylation by Dibenzyl trisulphide. Biochim Biophys Acta , 2001, 1540: 166 – 177.

3.

An, H., Zhu, J., Wang, X., and Xu, X. Synthesis and Anti-tumor Evaluation of New Trisulfide Derivatives.

Bioorg. Med. Chem. Lett. 2006, 16: 4826 - 4829.

4.

Williams, L. A.D., Rosner, H., Levy, H. G. and Barton E. N. A Critical Review of the Therapeutic Potential of Dibenzyl trisulphide Isolated from Petiveria alliacea L (Guinea Hen Weed, anamu). West Indian Med. J. 2007, 56 (1):17 - 21.

5.

http://www.researchandmarket.com/publication/mnj1u9s/

6.

Williams, L. A. D., Rosner, H., Moller, W. and Kraus, W. Anti-proliferation/cytotoxic Action of Dibenzyl trisulphide, a Secondary Metabolite of Petiveria alliacea. Jam. J. Sci. Tech. 2004, 15: 54 - 60.

7.

Williams, L.A.D., Rosner, H. and Kraus, W. Molecules with potential for cancer therapy in the developing world: Dibenzyl trisulfide (DTS). In genomic application for the developing world. Editors Karen E. Nelson and Barbara Jones-Nelson, Springer, New York, 2012, 273 – 278.

8.

Lowe, H. I.C., Facey, C. O. B., Toyang, N. J. and Bryant, J. L. Specific RSK kinase inhibition by dibenzyl trisulfide and implication for therapeutic treatment of cancer. Anticancer Res. 2014, 34: 1637 – 1642.

9. Williams, L.A.D. and Levy, G. H. Composition and method for treating cancer. US Patent Application US2008/058002. Publication No. wo/2008/118847, Class 514 12 Code A1, International application No. PCT/US2008/058002, 2008.

10.

Xu, X., Xu, X. An, H. and Wang, X. Substituted organosulfur compounds and methods of using thereof.

US Patent application number: 20090192146, USPC Class: 5142252, 2009.

11.

www.granlen.com/upload/files/2015/4/17113733437.pdf

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12.

Williams, L.A.D., Rosner, H., Conrad, J., Moller, W., Beifuss, U., Chiba, K., Nkurunziza, J. P. and Kraus, W. Selected secondary metabolites from the Phytolaccaceae and their biological/pharmaceutical significance.

Recent Research Development in Phytochemistry, 2002, 6: 13 – 68, Research Signpost.

13.

http://receptorkinase.gsc.riken.jp

14.

Williams, L.A.D., The T. L., Gardner, M., Fletcher, C. K., Naravane, A., Gibbs, N. and Fleishhacker, R. Immunomodulatory activities of Petiveria alliaceaL. Phytother. Res., 1997, 11: 251 - 253.

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3. Potential of Two Jamaican Essential Oils against Mosquitoes and Bacteria Chenielle M Delahaye McKenzie, Sylvia Mitchell and Dwight Robinson University of the West Indies, Jamaica

ABSTRACT: The antimicrobial and insecticidal properties of Callistemon viminalis (bottlebrush) and Cymbopogon citratus (lemongrass) have been studied. The results are encouraging Ca. viminalis and Cy. citratus essential oils were extracted using a steam distiller apparatus. The essential oils were separated from the essences and tested against three pathogenic bacteria (Staphylococcus aureus, Streptococcus pyogenes and Escherichia coli) using the disc diffusion and tube dilution assay.

Both essential oils were very effective inhibitors of microbes producing inhibition zones of > 20 mm and MIC of 0.008 mg/ml against S. aureus. The insecticidal potential of both essential oils was investigated using the olfactometer bioassay with mosquitoes. A spatial repellency assay was conducted using an olfactometer. The mosquitoes had a high repellency percentage to olfactory cue created by the essential oils of Ca. viminalis and Cy. citratus with a spatial activity index ranging from 0.27 to 0.77. Products being produced from these extracts should be in great demand in Jamaica and the wider world based on these findings.

INTRODUCTION Callistemon viminalis commonly known as bottlebrush and Cymbopogon citratus commonly known as lemongrass are two medicinal plants grown in Jamaica and other parts of the world. Bottlebrush is a small tree native to Australia1 and mainly used as an ornamental in Jamaica. Lemongrass in contrast is a tall coarse grass and a perennial herb cultivated as an ornamental and medicinal plant2. In this study Ca. viminalis and Cy. citratus essential oils were investigated to scientifically prove the ethnomedicinal claims of being antibacterial and insecticidal.

Ca. viminalis and Cy. citratus have been cultivated for their medicinal benefits all over the world. Their uses and benefits have some similarities as well as differences. Both medicinal plants are used as a folk remedy for coughs and consumption. However, Cy. citratus is also used to treat elephantiasis, malaria, ophthalmia, pneumonia and vascular disorders2. Researchers have also concluded that lemongrass possesses antidepressant, antioxidant, antiseptic, astringent, bactericidal, fungicidal, nervine and sedative activity3. Ca. viminalis on the other hand, is used to treat

acne, bronchitis, diarrhoea, and skin disorders4. Researchers have concluded that bottlebrush has bactericidal, fungicidal, antiseptic and disinfectant properties5.

Callistemon viminalis when extracted with methanol, hexane, ethyl acetate and water are all active antibacterial agents against Staphylococcus aureus, Methicillin resistant S. aureus (MRSA), Streptococcus pyogenes, Bacillus cereus, Escherichia coli, Pseudomonas aeruginosa, Salmonella enteritidis and Shigella sonnei5.

Ca. viminalis water and ethanol extracts were found to contain anti-quorum sensing activity against the bacteria Chromobacterium violaceum and Agrobacterium tumefaciens6. Cy. citratus essential oil, on the other hand, was an effective antibacterial agent against S. aureus, S. pyogenes, E. coli, B. subtilis and Klebsiella pneumonia. The lemongrass oil was, however, more effective against gram positive bacteria2.

Callistemon viminalis essential oil was tested against the insects Acanthoseelides dotectus and Callosobruchus maculates adults

by Ndomo et al7. The research concluded that using a fumigatory process, the essential oil of Ca. viminalis caused high inhibition of F1 progeny production of both insects7.

Cymbopogon sp. essential oil was also tested against eight

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stored-grain insect pests (Sitophilus oryzae, Tribolium costaneum and Rhyzopertha dominica by Lee et al8, the results showed potent fumigant activity against all the insects. Ca. citratus essential oil insect repellency was also studied by Baldacchino et al9 in France against mosquito (Diptera: Culicidae) and house flies (Diptera: Muscidae). The study concluded that lemongrass essential oil was an effective repellent against these insects9.

water mixture of 30 g sugar to 300 ml of water was placed in the bottle and yeast added. To prevent mosquitoes from drowning, a mesh partition was layered directly above the mixture but below opening of the inverted top half of bottle. The inverted top half of bottle was then taped to the bottom half, wrapped in dark paper/ plastic bag at the base and placed in a dark mosquito infested areas.

Preparation of different concentrations of essential oils Insects are often carriers of diseases and infections often caused by bacteria. Having a product that can ward off both bacteria and insects should reduce outbreaks significantly as a result of the synergistic effects. Bacterial and insecticidal products are usually sold separately. Products that can inhibit and repel these nuisances have an added benefit of increasing effectiveness and reducing the cost to consumers. This study explores the effectiveness of several formulations of the two essential oils (lemon grass and bottle brush) and the prospective combination of the two oils as antibacterial and insect-repellents, in a single product.

MATERIALS and METHODS

Extraction of Bottlebrush and Lemongrass Essential Oils Lemongrass and bottlebrush essential oils were extracted using the steam distillation method. Leaves of both plants were collected, washed and dried at room temperature.

About 500 g of the dried leaves was placed in the steam distiller with 2000 L of water. The steam distillation process was allowed to run for approximately 6 to 8 hours after which the essences and essential oils were collected and separated using a separating funnel.

Test organisms - (Bacteria and Insects) The bacteria used in this study were Staphylococcus aureus, Streptococcus pyogenes and Escherichia coli. These organisms were obtained from the culture collection of the microbiology department of the University of the West Indies, Mona, Jamaica.

The mosquitoes used in this study were collected using a plastic bottle mosquito trap with a 24 hour worn sock, yeast and sugar mixture. A 2 litre plastic bottle was cut in half and cover removed. The top half of the bottle was inverted and placed inside the bottom half of the bottle. A sugar and

Essential oils were dissolved in chloroform to obtain different concentrations (v/v) of bottlebrush and lemongrass essential oils. Concentrations produced included 10%, 20%, 30%, 40% and 50%.

Antibacterial Assay - (Inhibition Zones) The Agar well diffusion method as described by NCCLS (2000)10 was used in the testing of the antibacterial activity of bottlebrush and lemongrass essential oils against the bacteria S. aureus, S. pyogenes and E. coli. The well diffusion method used in this study was a modified form of the disc diffusion method as described by Bauer et al11. Muller Hinton agar in petri-dishes was used to test for bacterial susceptibility12 to the essential oils of bottlebrush and lemongrass.

1 ml of bacterial cell suspension was dissolved in peptone water and spread on agar plates using a sterile cotton swab.

Six wells of approximately 5 mm in diameter each were made in the MH agar petri-dishes equidistance apart, using the back of sterile glass droppers. A sterilized micro-pipette was used to dispense 25 µl of each concentration of the essential oils to the wells. Only a single concentration of the essential oil was placed in the six wells of each petri-dish.

The essential oils were allowed to diffuse in the wells 15- 20 minutes before being incubated at (37±1°C) for 24 hours.

A calliper was used to measure the inhibition zones. This experiment was repeated three times for each concentration.

Antibacterial Tube Dilution Assay - Minimum Inhibition Concentration (MIC) and Minimum Bacterial Concentration (MBC) Muller Hinton broth was used to create dilutions of the essential oils of bottlebrush and lemongrass. Cultures of the test bacterial strains were prepared and incubated overnight at (37±1°C). 25 µl bacterial cell suspension cultures were prepared and added to each set of serial dilutions of the

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essential oils to determine the MIC. The MIC was determined by the lowest dilution of the essential oil to inhibit the growth of micro-organisms (visual observation of microbial growth) and the MBC was the lowest dilution that killed the micro-organisms (no growth observed when liquid extract was plated out on agar). Each test tube used contained 10 ml sterile MH broth with a different concentration of bottlebrush and lemongrass essential oil ranging from 0.50% to 0.008% (v/v) MH broth. 0.5% (v/v) chloroform without oil was used as negative control, and gentamicin as a positive control. Test tubes with extracts and test organisms were incubated at (37±1°C) for 24 hours.

Aliquots of 1.5 ml of the lemongrass essential oil and the solvent DMSO were applied evenly to 11 x 25 cm (275 cm2) Whatman No. 1 paper with a pipette. The filter papers after being placed in the flask were allowed to sit for 15 minutes before the first test replicate was initiated to ensure the solvent was completely evaporated, leaving the lemongrass essential oil only on the filter paper13.

The test tubes were observed macroscopically after incubation of 24 hours for turbidity. The MIC was recorded as the lowest concentration at which no bacterial growth is visible. The tubes were then plated out on MH agar petridishes and incubated for a further 24 hours at 37±1°C. The highest concentration that showed no bacterial growth was recorded as the MBC. Each assay was repeated thrice.

Groups of about 20 mosquitoes were introduced into olfactometer cylinder and allowed to acclimatize for 2 minutes. Damaged or dead mosquitoes were recorded and a correct initial sample size recorded. Vapour of the treatment flask was allowed to circulate in the cylinder and observation recorded after 10, 30 and 60 minutes. The direction and distance in which the mosquitoes moved away from their point of entry was recorded. The mortality was also recorded after 2 hours. All the apparatus of the olfactometer was disconnected and washed at the end of each evaluation. Successive replicates of each condition were done twice.

Spatial Repellency Assay (Olfactometer Test)

Statistical Analysis

An Olfactometer apparatus was set up to determine the spatial repellence of mosquitoes by lemongrass and bottlebrush essential oils. A central clear cylinder was connected to a conical flask at each end, one with the control substance (dimethyl sulfoxide (DMSO)) and one (treatment, T) with a concentration of lemongrass essential oil (C-T arrangement). The center of the cylinder was the point of entry of the mosquitoes. A concentration gradient was created using a pressure pump that was attached between the two ends to distribute the air, prior to the entry of the mosquitoes. Foil paper was wrapped around the flask (containing essential oil) to stimulate darkness13.

Data recorded from the antibacterial and spatial repellency assay were analyzed using SPSS Analysis of Variance (ANOVA). Conclusions were drawn from the analysis that determined if the essential oils were effective and what concentrations they were most effective.

As the negative control, filter paper treated with dimethyl sulfoxide (DSMO) (the diluent) was placed at both ends of the olfactometer test unit (C-C arrangement) simultaneously. This ensured an even, 13 distribution of mosquitoes for the test conditions of temperature and humidity. Multiple assays were run at the same time using different flasks labeled accordingly T1, T2, T3, T4, and T5 for the different concentrations of lemongrass essential oil. Note, the same concentration of DMSO (undiluted) was used for all tests, however for each test a different container of DMSO was used.

RESULTS Ca. viminalis and Cy. citratus were both active antibacterial agents against S. aureus, S. pyogenes and E. coli (Table 1). The bioactivity against the bacteria however increased as the concentration of the extracts increased. Only inhibition zones greater than 10 mm were considered effective and are represented in Table 1 by plus signs. Table 1 also shows the activity of the negative control (diluent) chloroform and the positive control gentamicin. Chloroform had no bactericidal effect; this suggests that chloroform had no added effect on the dilutions of the essential oils.

Gentamicin, on the other hand, is a known antibacterial, and when tested against the bacteria was very effective (Table 1). The inhibition zones created by gentamicin were used to compare the effectiveness of the essential oils.

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Table 1 Antibacterial activity of Lemongrass and Bottlebrush essential oils against three bacteria

Plus signs indicated the size of the inhition zones created by the essential oils against the bacteria, (-) <10 mm, (+) < 15mm, (++) < 20 and (+++) > 20 mm. Inhibition zones less than 10 mm are not considered to be effective and, therefore, are represented by a (dash). N= 18, 6 replicates for 3 repeated experiments

Higher concentrations of essential oils (50) were significantly more effective against the microbes than the lower concentrations (20%). The 50% bottlebrush and lemongrass essential oils were significantly more active, with inhibition zones greater than 20 mm in diameter, than the 20 % concentrations of the essential oils. 10% lemongrass essential oil was not effective against E. coli; the inhibition zones created were less than 10 mm in diameter, Table 1. The10% lemongrass oil was however very effective against S. aureus and S. pyogenes, having inhibition zones at about 20 mm in diameter, Figure 1C. The 20 % lemongrass oil was not as effective against E. coli when compared to the 20% bottlebrush oil. Figure 1A shows the large inhibition zones created against E. coli by the 20% bottlebrush essential oil, unlike the 20% lemongrass oil that had zones less than 15 mm. The 20% lemongrass oil had inhibition zone against S. aureus and S. pyogenes that were not significantly different from that of 20% bottlebrush oil; inhibition zones were about 20 mm in diameter, Table 1. Concentrations of 30%, 40% and 50% lemongrass oil and 50% bottlebrush oil were all effective against the test microbes, all of which had inhibition zones greater than 20 mm, Table 1 and Figure 1B, E and F. Figure 1F shows inhibition zones that were so large that they fused into each other. The measurements, in this case, were done at the outer end of the zones using a caliper. At higher concentrations (40% and 50% lemongrass oil and

50% bottlebrush oil), no microbial growth occurred on the plates. These experiments were repeated three times with only three wells, the same inhibition of growth on the entire petri-dish occurred. Inhibition zones were recorded as being greater than 40 mm in diameter.

Figure 1. Petri dishes showing inhibition zones created by bottlebrush and lemongrass essential oils. A- 50 % BB against E. coli; B- 30 % LG against S. pyogenes; C- 10 % LG against S. aureus; D- 20 % BB against S. aureus; E- 40 % LG against E. coli; and F- 30 % LG against S. aureus. The caliper shown in A and C was the instrument used to measure the inhibition zones.

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Mosquito Repellence

sent in the treated and untreated zones (0 : 32 and 2 : 18, respectively).

Table 2. Spatial repellency of bottlebrush and lemongrass essential oils against mosquitoes using olfactometer, observing attraction or repellency to the zone treated and untreated with essential oils.

Figure 2 Olfactometer to test the repellency of mosquitoes to bottlebrush and lemongrass essential oil. A - the conical flask attached with no treatment, B- the conical flask with filter paper at base soaked with oil, C- the point of entry of mosquitoes and attachment of pressure pump, D- pressure pump. Circles around the cylinder represent a scale used to measure the distance moved from the point of entry Figure 2 shows the olfactometer used to test the spatial repellency of the essential oils against mosquitoes. A and B show conical flasks in which, respectively, control and treatment solutions are soaked onto filter paper. C shows the point of entry of mosquitoes, and where the pressure pump is attached using a clamp. Lemongrass essential oil was a better repellent than bottlebrush essential oil. Table 2 shows the median distance of the mosquitoes from the treated and untreated zones with the essential oils. These results indicate that significantly fewer mosquitoes remained in the treated zone of all the essential oils.

Lemongrass oil had one or no mosquito in the treated zone for the entire observation period, Table 2. The potency of the lemongrass essential oil caused the mosquitoes to move further away from the treated zone when compared to the bottlebrush essential oil. There was no significant difference between the mosquitoes in the treated and untreated zones when DMSO was used as the treatment. On the other hand, when the known mosquito repellent OFF was used as a treatment there was a significant difference between the number of mosquitoes pre-

There was no significant difference between the mosquitoes in the treated and untreated zones when DMSO was used as the treatment. On the other hand, when the known mosquito repellent OFF was used as a treatment there was a significant difference between the number of mosquitoes present in the treated and untreated zones (0 : 32 and 2 : 18, respectively).

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DISCUSSION This study provides clear evidence that both bottlebrush and lemongrass essential oils possess antibacterial and mosquito repellent activities against test organisms. The well diffusion and tube dilution assays supported similar observation by Cimanga et al14, that the gram-positive bacteria were more sensitive to the essential oils than the gramnegative bacteria. All the concentrations of the lemongrass and bottlebrush essential oils were antibacterial, except 10 % lemongrass against E. coli. This may be due to the low concentration that possessed less of the active compound to penetrate the lipopolysaccharide membrane of the gramnegative bacteria E. coli15. El-Ahmady et al16 studied the essential oil of C. viminalis and found that S. aureus was more susceptible with an average inhibition zone of 18 mm than to the gram-negative Pseudomonas aeruginosa. In addition to being antibacterial agents, the essential oils were also found to be mosquito repellents at all concentrations tested. All concentrations of lemongrass and bottlebrush oil were active; however lemongrass showed similar activity to the known mosquito repellent brand product “OFF”.

His study provides clear evidence that both bottlebrush and Cimonga et al,13 that the gram-positive bacteria were more sensitive to the essential oils than the gram-negative bacteria. All the concentrations of the lemongrass and bottlebrush essential oils were antibacterial, except 10% lemongrass against E. coli. This may be due to the low concentration that possessed less of the active compound to penetrate the lipopolysaccharide membrane of the gram-negative bacteria E. coli (Adwan and Abu-Hassan15. El-Ahmady et al16 studied the essential oil of C. viminalis and found that S. aureus was more susceptible with an average inhibition zone of 18 mm than to the gram-negative Pseudomonas aeruginosa. In addition lemongrass essential oils possess antibacterial and mosquito repellent activities against test organisms. The well diffusion and tube dilution assays supported similar observation antibacterial agents, the essential oils were also found to be mosquito repellents at all concentrations tested. All concentrations of lemongrass and bottlebrush oil were active; however lemongrass showed similar activity to the known mosquito repellent brand product “OFF”.

Lemongrass and bottlebrush essential oils inhibited the growth of the test organisms at lower concentrations in the tube dilution assay when compared to the well diffusion

assay. This may be due to the difference in microbial growth, the exposure of the microorganisms to the oil (broth provided a larger surface area), the solubility of the oil, the compounds in the oil, the way in which the oil is being used, and the solvent used to dilute the oil17;18.

Oyedeji et al19 reported similar antibacterial activity of C. viminalis essential oil, reporting inhibition zones of 23.0 mm - 26.3 mm against S. aureus when 0.5 mg/ml essential oil was used and MIC of 0.3- 2.5 mg/ml against P. aeruginosa. 50% C. viminalis and Lemongrass essential oil resulted in inhibition zone greater than 20 mm against all micro organisms. These results shows that the oils are as active as the extracts (ethyl acetate, methanol, hexane and water) of C. viminalis against the test organisms used (such as Staph. aureus, Strep. pyogenes and C. albicans,5;6.

The spatial repellent activity of both lemongrass and bottlebrush essential oil was also observed. The mosquitoes moved away from the lemongrass oil more than that of the bottlebrush oil. In a review, Sahelian20 suggested that lemongrass provided short-lived protection against mosquitoes while the oil-of-eucalyptus products provided longer lasting protection. C. viminalis is from the same family Myrtaceae as eucalyptus and contains similar compounds such as eucalyptol 21 in its oils. Therefore, similar repellent activity was expected; however, this was not the case. The mosquitoes were not attracted to bottlebrush or lemongrass oil; instead they avoided or were repelled by the vapours of these oils over a 30 mins period. The lemongrass showed a greater spatial repellent activity than bottlebrush, but similar to that of the brand ‘OFF’. In a review done by Sahelian20 of the lemongrass and eucalyptus oil the mosquito repellent assay was done differently.

Human subjects were treated topically with the oils and placed in a mosquito-infested chamber. This study used an olfactometer and the movement of the mosquitoes towards or away from the oils observed. Similar results were achieved with the olfactometer in this study compared to the human chamber used by Sahelian20.

Analysis of lemongrass essential oil by Yousef24 and bottlebrush essential oil by Srivastiva et al21 indicated that similar antibacterial compounds such as myrcene, 1, 2-cineole, geranial, menthyl acetate, citronellyl acetate, α-pinene, limonene, and linalool. These compounds were present in both oils at different concentrations21,24. The presence of these compounds in both oils may be the reason both oils

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are antibacterial and a mosquito repellent. Geranial is one of the main compounds responsible for repelling mosquitoes, and this was present in both oils. According to Yousef24, a mixture of geranial and neral produces a compound called citral which is the main compound causing lemongrass to be such an effective repellent against mosquitoes.

oils may be a skin irritant, the dilutions used in this study indicate that the oils are still very active at lower concentrations and therefore can be mixed into various formulations. This investigation shows the possibility of incorporating essential oils into products.

Lemongrass and bottlebrush essential oils are natural, nontoxic, effective mosquito repellent and antibacterial agents that are economical feasible.

In conclusion, the essential oil of lemongrass and bottlebrush can be utilized for both their antibacterial and mosquito repellent activity. Although the neat of the essential REFERENCES 1.

Spencer R. D., and Lumley, P. F. 2007. New South Wales Flora Online: Callistemon viminalis Royal Botanic Gardens & Domain Trust, Sydney, Australia

2.

Naik, MI, Fomda, BA, Jaykumar, E and Bhat, J. 2010. “Antibacterial activity of lemongrass (Cymbopogon citratus) oil against some selected pathogenic bacteria.” Asian Pacific Journal of Tropical Medicine. Pg 535-538

3.

McGuffin M, Hobbs C and Upton R. 1997 “American herbal products association botanical safety handbook.” Boca Raton: CRC press.

4.

Wallace, R. J. 2004. Antimicrobial properties of plant secondary metabolites. Proc. Nutr. Soc. 63, 621-629.

5.

Delahaye, Chenielle, Lois Rainford, Allison Nicholson, Sylvia Mitchell, John Lindo, Mohammed Ahmad. 2009.

“Antibacterial and antifungal analysis of crude extracts from the leaves of Callistemon viminalis.” Journal of Medical Biological Science. 3(1): 1-7.

6.

Adonizio, AL., Downumk K, Bennett BC, and Mathee K. 2006. “Antiquorum sensing activity of medicinal plants in Southern Florida.” Journal of Ethnopharmacy 105(3):427-35. DOI: 10.1016/j.jep.2005.11.025

7.

Ndomo, Agnès Flore, Leon A. Tapondjou, and Leonard Tinkeu Ngamo. 2009. “Insecticidal activities of essential oil of Callistemon viminalis applied as fumigant and powder against two bruchids.” Journal of Applied Entomology 134 (4).

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Lee, ByungHo, Annis P.C., Tumaalii F., Choi WonSik, 2004. Fumigant toxicity of essential oils from the Myrtaceae family and 1,8-cineole against 3 major stored-grain insects. Journal of Stored Products Research, 40(5):553-564. http:// www.sciencedirect.com/science retrieved Jan 2015

9.

Baladacchino F, Muenworn V, Desquesnes M, Desoli F, Charoenviriyaphap T and Duvallet G. 2013. “Transmission of pathogens by Stomoxys flies (Diptera, Muscidae): a review.” Parasite 20, 26 http://www.parasite-journal.org/articles/ parasite/pdf/2013/01/parasite130035.pdf

10.

NCCLS. 2000. Performance standards for antimicrobial susceptibility testing: twelfth informational supplement M100S12. NCCLS, Wayne, Pa.

11.

Bauer AW, Kirby WMM, Shevis JC and Turck M. 1966. “Antibiotic susceptibility testing by a standardized single disc method.” American Journal of Clinical Pathology; 45:493-6.

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NCCLS. 2002. Performance standards for antimicrobial susceptibility testing: twelfth informational supplement M100S12. NCCLS, Wayne, Pa.

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13.

World Health Organization 2013. “Guidelines for efficacy testing of spatial repellents.” Control of neglected tropical diseases WHO pesticide evaluation scheme. WHO library-cataloguing-in-publication Data pp 7-10.

14.

Cimanga K, Tona L, Apers S, Bruyne Tde, Hermans N, Totte J, et al. 2002. Correlation between chemical composition and antibacterial activity of essential oils of some aromatic medicinal plants growing in the Democratic Republic of Congo. J Ethanopharmacol; 79(2): 213-20.

15.

Adwan, Kamel and Nael Abu-Hasan. 1998. Gentamicin resistance in clinical strains of Enterobacteriaceae associated with reduced gentamicin uptake. Folia Microbiology 43:438-40.

El- Ahmady Sherweit., Mohamed El-Shazly, and Rola Milad. 2013. “The Synergetic Efficacy of the Combination of Amphotericin B and Certain Essential Oils against selected Fungal Clinical isolates.” Journal of Applied Pharmacological Science. 3 (04): 026-030.

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17.

Hili P, Evans CS, Veniss RG. Antimicrobial action of essential oils: the effect of Dimethyl sulphaoxide on the activity of Cinnamon oil. Let Appl Microbiol 1997; 24: 269-75.

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Janssen AM, Scheffer JJC, Baerheim SA. Antimicrobial activity of essential oils: a 1976-1986 literature review. Aspects of the test methods. Plt Med 1987;53: 395-8.

19.

Oyedeji, Opeoluwa O., Oladipupo A Lawal, Francis Shode, and Adebola O Oyedeji. 2009. “Chemical composition and antibacterial activity of the essential oils of Callistemon citrinus and Callistemon viminalis from South Africa.”

Molecules. 14 (6).

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Sahelian, R. 2014 “Mosquito natural repellant.” Complement Ther Med. File://C:/Users/User/Desktop/ Mosquito.html (retrieve 05 Oct. 2014)

21.

Srivastava, S. K., Ateeque Ahmad , K. V. Syamsunder, K. K Aggarwal, and S. P. S. Khanuja. 2003. “Essential oil composition of Callistemon viminalis leaves from India.” Flavour and Fragrance Journal. 18 (5): 361-363. www.interscience.wiley.com (retrieved July 2, 2013).

22.

Eifert, JR. 2004, ‘The effectiveness of lemongrass as a natural Mosquito Repellent.” California State Science Fair. Project Number J1408

23.

Maia, MF and Moore, SJ. 2011. “Plant-based insect repellents: a review of their efficacy, development and testing.”

Malaria Journal Vol 10; suppl 1 www.ncbi.nlm.nih.gov/pmc/articles/PMC3059459 (retrieved 04 Oct. 14)

24.

Yousef SAA. 2013. “Antibacterial activity of volatiles from lemongrass (Cymbopogon citratus) and peppermint (Mentha piperita) oils against some respiratory pathogenic species of Aspergillus.” International Journal of Current Microbiology and AppliedSciences 2(6) pp 261-276.

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4. Dietary Intake of Trace and Heavy Metals from Consumption of Instant Coffees on the Jamaican Market Johann M.R. Antoine, Leslie A. Hoo Fung, Charles N. Grant International Centre for Environmental and Nuclear Sciences, University of the West Indies, Mona Kingston, Jamaica, West Indies ABSTRACT: Eleven brands of instant coffee sold on the Jamaican market were collected and analyzed. Al, As, Ba, Br, Ca, Cd Ce, Co, Cr, Cs, Cu, Eu, Fe, Hg, K, La, Mg, Mn, Mo, Na, Ni, P, Pb, Rb, Sb, Sc, Se, Sm, Sr, Th and Zn were determined using Graphite Furnace Atomic Absorption Spectrophotometry, Inductively Coupled Plasma Optical Emission Spectroscopy and Instrumental Neutron Activation Analysis. Contributions to the Provisional Tolerable Weekly Intakes for potentially toxic elements were estimated for aluminium (0.26-0.34%), arsenic (0.02-0.03%), cadmium (0.06%), lead (0.03%) and mercury (0.33 â&#x20AC;&#x201C; 0.54%). Contributions to recommended daily intakes were also estimated for macronutrients calcium (0.21-0.40%), magnesium (1.6-2.6%), phosphorus (0.7-0.9%), potassium (1.5-1.7%) and sodium (0.004-0.056%) and micronutrients copper (0.009-0.013%), chromium (0.001-0.002%), iron (0.3-0.9%), manganese (1.7-4.5%), molybdenum (1.8-2.4%), selenium (0.4-0.8%) and zinc (0.06-0.10%). Instant coffees contribute a very low percentage of the Provisional Tolerable Weekly Intake for all potentially toxic elements examined. The dietary intake of most essential elements from instant coffees is also negligible, although moderate consumption would contribute some amount of potassium, magnesium, manganese and molybdenum to the Recommended Dietary Intake of those elements.

INTRODUCTION Coffee is one of the most popular beverages in the world.

It is also one of the most valuable agricultural commodities for the developing world1. Jamaica is a net exporter of coffee with Jamaican Blue Mountain coffee commanding a premium price on the retail market, as high as US$95 per kilogram. While roasted and ground Jamaican coffees retail at high price point, instant coffees are also growing in popularity, with new brands being introduced in recent years in order to gain a piece of the market2.

of instant coffee is not simply a local phenomenon but may reflect the global growth of the soluble coffee market which according to the Global Coffee Report has risen by annual growth rates of 7 to 10 percent in the past decade4.

Instant coffee is increasingly consumed, with roughly half of world consumption now being instant coffee5, in part because of rising consumption in emerging markets6. It is possible that the lower price point of soluble coffee in comparison to roasted and ground coffee as well as the convenience and lower cost of preparation are important factors in the growth of local consumption.

Recently there has been a significant rise in the consumption of soluble coffee by Jamaicans. Importation of regular instant coffee was 16,872 kg in 2010, 19,867 kg in 2011, 23,569 kg in 2012 and 36,419 kg in 2013. During the years 2010-2013, importation of instant coffee premixes was 113,630 kg, 119,552 kg, 105,379 kg and 228,045 kg respectively. Total imported soluble coffee products more than doubled from 130,502 kg in 2010 to 264,464 kg in 2013.

During this four-year period importation of regular roasted coffee occurred only in 2011 and 2012 and was 2,867 kg and 371 kg respectively3. This increased consumption

Diet is, in general, the primary method of exposure to heavy metals and trace elements for the nonoccupationally exposed population. In order to assess this exposure and the potential risk that may be posed the elemental content of various foods has to be known as well as consumption data. Because of the islandâ&#x20AC;&#x2122;s geochemistry Jamaicans may already be exposed to a variety of potentially toxic elements through the consumption of locally grown crops7,8. With the marked expansion of the soluble coffee market in Jamaica an evaluation of the elemental food safety of these beverages is important. When one

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considers that a significant portion of the soluble coffee consumed is imported the assessment of trace elements and heavy metals becomes even more pertinent. This study was therefore undertaken to determine the trace elements and heavy metals of all soluble coffee brands, both local and imported on the Jamaican market at the time of sampling and with this data to estimate the dietary intake of these elements based on typical serving sizes.

METHODS

Samples Eleven brands of instant coffee sold on the Jamaican market were collected. Of these, 6 were Jamaican instant coffee (5 regular, 1 decaffeinated) and 4 were imported (3 regular, 1 decaffeinated). The remaining sample is processed by a Jamaican manufacturer using imported robusta coffee. Elemental analysis of these samples was performed using Instrumental Neutron Activation Analysis (INAA), Inductively Coupled Plasma Optical Emission Spectroscopy (ICP-OES) and Graphite Furnace Atomic Absorption Spectrophotometry (GF-AAS).

Samples were prepared for Inductively Coupled Plasma Optical Emission Spectroscopy (ICP-OES) and Graphite Furnace Atomic Absorption Spectrophotometry (GFAAS) analysis by microwave-assisted acid digestion. 10 mL of HNO3 (Trace Metal Grade) was added to 0.5 g of sample in an EasyPrep Teflon vial and allowed to stand for 1 h before digestion using a CEM MARS5 microwave system (CEM Corporation, NC, USA). After cooling, samples were made up to 25 mL using deionized water.

Analyses were performed using a PerkinElmer Optima 7000DV ICP-OES (PerkinElmer, MA, USA) and PerkinElmer AAnalyst 600 GF-AAS (PerkinElmer, MA, USA).

Calibration standards for ICP-OES and GF-AAS analyses were prepared using Certiprep solutions (SPEX Certiprep, NJ, USA) in 2% HNO3. An internal standard containing 0.2 mg/L yttrium was also used.

Instrumental Neutron Activation Analysis (INAA) was performed using the SLOWPOKE-2 nuclear reactor. For the determination of short-lived radioisotopes, approximately 0.5 g of sample was weighed out into pre-cleaned double polyethylene bags and heat sealed in pre-cleaned 7 cm3 polyethylene vials9. Each sample was irradiated for 3 min at a neutron flux of 5 x 1011 n cm-2 s-1 and allowed decay periods of approximately 5 and 60 min before

counting. For intermediate and long lived radioisotopes approximately 1 g of sample was weighed out in precleaned polyethylene capsules which were then heat sealed in 7 cm3 polyethylene vials and irradiated for 4 h at a neutron flux of 10 x 1011 n cm-2 s-1 and allowed decay periods of 4 days and 14 days respectively before counting.

For short and intermediate-lived radioisotopes gamma ray spectroscopy was performed using an Ortec High-Purity germanium coaxial gamma photon detector system with an efficiency of 71% and a resolution of 1.8 keV at the 60Co 1332 keV gamma line. Long lived radioisotopes were counted on an EG&G Ortec High-Purity Germanium detector with an efficiency of 20% and a resolution of 1.9 keV at the 60Co 1332 keV gamma line.

Cd and Pb were determined using GF-AAS; Cr, Cu, Fe, Mo, Ni, P, Sr and Zn were determined using ICP-OES; and Al, As, Ba, Br, Ca, Ce, Co, Cs, Eu, Hg, K, La, Mg, Mn, Na, Rb, Sb, Sc, Se, Sm, and Th were determined using INAA. In total thirty-one (31) elements were determined.

Quality Control Approximately 10% of the samples were analyzed in duplicate, with the differences between duplicates being less than 15%; at least one reagent blank and a certified reference material were also included in each analysis batch.

Reference materials used were NIST (National Institute of Standards and Technology, MD, USA) 1573a – Tomato Leaves, and IAEA (International Atomic Energy Agency, Vienna, Austria) 336 – Lichen. Reference material data are presented in Table 1.

There is no value for lead in NIST 1573a. All ICP-OES and GF-AAS analyses were also validated using certified multi-element solutions (PerkinElmer, MA, USA).

For most elements the difference between duplicate analyses was less than 10%, with a few having RSD between 10 and 20%; elements with RSDs greater than 20% were those which approached the detection limit of the technique. The agreement between techniques was excellent.

Recovery for all reference materials used was within acceptable limits of ± 20% of expected values.

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Table 1: Reference Material Data

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Table 2â&#x20AC;&#x201D;Elemental Concentrations in Instant/Soluble Coffees

All data expressed as mg kg-1 unless otherwise stated Numbers in bold represent detection limit values

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For the elements analyzed by INAA, validation was achieved by the irradiation and counting of single element standards (Certiprep) with identical geometry to that used for samples employing the improved relative standardization method10.

Table 3B: Intake estimates for potentially toxic elements from imported instant coffee

RESULTS AND DISCUSSION Element data for all samples is presented in Table 2. Samples were categorised as Jamaican, imported and decaffeinated to obtain mean values. The single brand of 100% Robusta variety coffee on the market was treated as its own category in the dataset. A serving size of 2g of instant coffee powder was used for the purpose of estimating dietary intake of essential elements and exposure to potentially toxic elements. Jamaican statistics on per capita coffee consumption also include beverages that are not 100% coffee; therefore use of a standard serving size is more useful. Daily intake estimates were calculated using the formula:

Table 3C: Intake estimates for potentially toxic elements from decaffeinated instant coffee

Intake (mg) per 2g serving = mean concentration (mg kg-1) x quantity (kg) (1) The concentrations and intake values are based on the dry weight of the coffee powder as it is assumed that the entire quantity is dissolved when prepared for consumption.

Potentially toxic elements: As, Al, Cd, Hg, Pb Intake estimates per 2g serving of instant coffee relative to Provisional Tolerable Weekly Intake (PTWI) for potentially toxic elements are presented in Tables 3A-3D. Mean values in bold type indicate detection limit values.

Table 3A: Intake estimates for potentially toxic elements from Jamaican instant coffee

Table 3D: Intake estimates for potentially toxic elements from Robusta instant coffee

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Aluminium is of concern due to growing evidence of its toxicity, resulting in the lowering of the PTWI from 7mg kg -1 of body weight (BW) to the current 1 mg kg-1 BW in 200611. The mean aluminium values for all categories of instant coffee were well above the limits of detection, and the concentration in the Robusta sample was lower than for other categories. The contribution of a single serving to the PTWI12 is less than 0.5% in all cases.

Table 4A: Intake estimates for essential nutrients from Jamaican instant coffee

Arsenic in foods occurs predominantly as organic species [13] and so is generally not a great concern (as opposed to inorganic arsenic species in polluted groundwater). The WHO has prescribed a PTWI of 0.015 mg kg-1 BW for inorganic arsenic12. The arsenic values for all categories of instant coffee were below the limit of detection; even if this absolute value is assumed to be all inorganic arsenic, the contribution to PTWI is a fraction of a percent in all instances.

Cadmium has not been shown to have any biological role in humans and is regarded as a carcinogen and nephrotoxin14.

All mean values for cadmium (and all samples) are below the limit of detection and would not contribute significantly to the PTWI of 0.007 mg kg-1 BW12.

Lead has been well established as a neurotoxin and has no known essentiality to humans. Only one sample of Jamaican instant coffee had measurable levels of lead, and even at this maximum value (0.094 mg kg-1) a serving of instant coffee would contribute a miniscule 0.04% of the PTWI of 0.05 mg kg-1 BW12 to dietary intake.

The toxicity of mercury and its compounds has been well studied and the neurotoxicity of mercury, from the fetus to adults has been well documented. However dietary concerns about mercury usually centre on fish and fishery products, and, to a lesser extent, rice. Most of the samples analysed had mercury levels below the limit of detection.

One sample of Jamaican instant coffee had measurable levels of mercury (0.22 mg kg-1) and one serving would contribute 1.10% of PTWI for men and 1.18% of PTWI for women. All other samples would contribute less than 0.5% of the PTWI for total mercury, 0.004 mg kg-1 of body weight12.

Intake estimates per 2g serving of instant coffee relative to Recommended Daily Intake (or Adequate Intake) levels for essential elements15 are presented in Tables 4A-4D.

Table 4B: Intake estimates for essential nutrients from imported instant coffee

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Table 4C: Intake estimates for essential nutrients from decaffeinated instant coffee

Macronutrients: Ca, K, Mg, Na, P Soluble coffee is known to be relatively enriched in several macronutrients including potassium, magnesium and phosphorus and moderately high in calcium and sodium16.

Table 4D: Intake estimates for essential nutrients from Robusta instant coffee

Potassium in the samples analyzed ranged from 3.56% in Folgers Instant Coffee to 4.36% in Coffee Roasters Jamaican Instant Coffee with an average content of 3.80%. Jamaican soluble coffee averages 3.95%, foreign soluble coffee averaged 3.63% and the two decaffeinated coffees averaged 3.76%. The lone soluble coffee sample made from the Robusta variety of coffee was 3.61%. Potassium was the highest inorganic constituent of the soluble coffee sampled by at least an order of magnitude. Percentage levels of potassium would typically make a food a significant source of this mineral. However, a serving size of 2 grams would on average, contribute less than 2% of the recommended daily intake (RDI) of potassium for both men and women (See Tables 4A-4D). The highest average of 1.68% of the RDI would be from the Jamaican soluble sample and the lowest average from the foreign coffees at 1.55%. The lone Robusta sample would contribute 1.54% to the RDI for potassium. Multiple servings would obviously make soluble coffee a more significant source of this macronutrient.

Magnesium in the samples ranged from 3207 mg kg-1 in Folgers Instant Coffee to 4600 mg kg-1 in Coffee Roasters Jamaican Instant Coffee with an average of 3919 mg kg-1.

Jamaican soluble coffee would on average contribute 2.0% of the RDI for magnesium for males and 2.6% for females.

The mean foreign soluble coffee would provide 1.6% and 2.1% of the RDI for magnesium respectively. Decaffeinated coffee averaged a similar RDI contribution as the Jamaican coffee at 2.0% and 2.6% for men and women respectively.

The Robusta sample contributed 1.9% and 2.5% (See Tables 4A-4D). Magnesium was second only to potassium in terms of its concentration levels; despite this the concentrations of magnesium in the samples would make soluble coffee a relatively greater source of magnesium than potassium although in the typical serving size this cannot be described as a significant.

Phosphorus ranged from 2518 mg kg-1 in Maxwell House Instant Coffee to 3291 mg kg-1 in Coffee Roasters Jamaican Instant Coffee with an average of 2865 mg kg-1. Jamaican coffee averaged the highest concentrations of coffee at

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3026 mg kg-1 and at these levels would contribute approximately 0.9% of the RDI for phosphorus for men and women per 2 g serving size (see Table 4A). Imported soluble coffee averaged 2614 mg kg-1 and would contribute about 0.7% of the RDI for phosphorus per 2g serving (see Table 4B). Two grams of decaffeinated coffee would contribute an average of 0.8% of the RDI for men and women at a mean concentration of 2820 mg kg-1 (see Table 4C). At 2899 mg kg-1 the Robusta sample would contribute roughly the same to the RDI as the decaffeinated samples (see Table 4D). Phosphorus is stored as phytic acid or phytate in plants. Because of its binding affinity for iron and its chelating effect on other elements such as zinc and calcium it is considered an anti-nutrient. Phytate levels in coffee can be relatively high. It is known that roasting does lower phytate levels so it is significantly lower in roasted and ground coffee but this is not the case for instant coffee17. Although the levels of phosphorus are relatively high in instant coffee the intake of between 5 and 6mg day-1 suggests it is similarly unlikely to be a significant source of phytic acid.

Calcium ranged from 1314 mg kg-1 in Nescafe Clasico Instant Coffee to 2059 mg kg-1 in Jablum Instant Coffee with an average of 1748 mg kg-1. A 2g serving size of Jamaican instant coffee would contribute 0.38% of the RDI of calcium for men and about 0.29% for women. Imported coffee which averaged 1336 mg kg-1 would contribute even less to the male and female RDI at 0.27% and 0.21% respectively.

Decaffeinated brands averaged 1869 mg kg-1 and would contribute 0.37% and 0.29% to the RDI for men and women respectively, per 2g serving size. The Robusta sample (1985 mg kg-1) would contribute approximately 0.39% of the RDI for men and 0.31% for women per 2g serving.

Despite the relatively high levels of calcium analyzed in the soluble coffee samples the contribution to RDI could be considered insignificant (see Tables 4A-4D).

Sodium, when compared to the other macronutrients discussed, showed large variation among the coffee analyzed.

In the samples it ranged from a low of 42 mg kg-1 in the Shirriff Instant Coffee, the lone Robusta sample, to a high of 2939 mg kg-1 in the True Brew Jablum Instant Coffee.

The second highest sample was Folgers Instant Coffee at 969 mg kg-1. In fact, the range of Jamaican coffees excluding the True Brew Jablum sample was 49mg kg-1 to 114 mg kg-1. With this high value included, Jamaican coffee (mean of 646.2mg kg-1) contributed only about 0.06% of the RDI

for sodium. Imported coffee, averaging 620.4 mg kg-1 contributed about 0.05% and decaffeinated coffees at an average of 335 mg kg-1 contributed about 0.03% to the RDI per 2g serving. Excess dietary sodium can lead to hypertension, and may have more serious implications in people with congestive heart failure, cirrhosis or renal disease. Sodium occurs naturally in most foods but most dietary sodium is added, particularly to processed foods. By some estimates, as much as 75% of dietary sodium comes from processed foods [18], and purchased meals may also be high in sodium. Considering that the typical westernized diet is not considered deficient in sodium but rather the converse it is likely a positive thing that a serving of soluble coffee cannot be considered a significant source of sodium.

Micronutrients: Cr, Cu, Fe, Mn, Mo, Se and Zn The process of coffee solubilisation, apart from resulting in instant coffee having high levels of potassium, magnesium and phosphorus and sodium also appears to result in low concentrations of elements such as copper and zinc19.

Chromium ranged from 122 µg kg-1 in Nescafe Clasico Instant coffee to 244 µg kg-1 in Coffee Roasters Jamaican Instant Coffee with a mean concentration of 192 µg kg-1 (see Table 2). In a 2g serving size the mean Jamaican soluble coffee concentration (208 µg kg-1) would contribute only 0.0012% and 0.0017% to the RDI for men and women respectively (see Table 4A). Imported coffee at a mean concentration of 168 µg kg-1 would contribute slightly less at 0.0010% and 0.0013% (see Table 4B). Decaffeinated coffee (mean of 184 µg kg-1) would contribute roughly 0.0010% and 0.0015% for men and women respectively.

The Shirriff branded Robusta coffee would contribute 0.0011% to the RDI for men and 0.0016% for women.

Copper was found to be below the limit of detection of 40 µg kg-1 for all but one sample, the Folgers Instant Coffee which had a concentration of 97 µg kg-1 (see Table 2). Jamaican coffee, decaffeinated coffee and the lone Robusta sample would all provide approximately 0.009% of the RDI for copper. Imported coffee would contribute 0.013% to the RDI. The Folgers coffee by itself would contribute 0.02% of the RDI per 2g serving.

Iron content ranged from a low of 19.2 mg kg-1 in Nescafe Clasico Instant Coffee to a high of 43.3 mg kg-1 in Folgers

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Instant Coffee, with a mean of 29.8 mg kg-1. Jamaican soluble coffee averaged 30.3 mg Fe kg-1 and would contribute 0.76% of the RDI required for men and 0.34% for women (see Table 4A). Imported coffee on average (34.7 mg kg-1) contributed more than Jamaican coffee but was still only 0.87% of the RDI for men and 0.39% for women in a 2g serving (see Table 4B). Decaffeinated samples averaged 24.1mg Fe kg-1 and the contribution to the RDI for men and women were 0.60% and 0.27% respectively (see Table 3C). The Robusta sample had 24.4 mg kg-1 and was similar to decaffeinated coffee in its contribution to RDI of 0.60% and 0.27% (See Tables 2 and 4D). According to the World Health Organization20 iron deficiency is worldâ&#x20AC;&#x2122;s most common and widespread nutritional deficiency. It is also one of the few nutritional deficiencies prevalent in the developed world. With such low concentrations per serving size even multiple cups of soluble coffee would be a negligible source of iron. Manganese concentrations ranged from 16.5 mg kg-1 in Nescafe Clasico Instant Coffee to 40.8 mg kg-1 in the Shirriff Instant Coffee. At this concentration the Robusta coffee would contribute a respectable 3.6% of the RDI for men and 4.5% for men in a 2g serving (see Table 4D).

Jamaican instant coffee averaged 29.4 mg kg-1 and would contribute 2.6% for the male RDI and 3.3% for females (see Table 4A). The imported coffee (mean content, 20.1 mg kg-1) would contribute roughly 1.7% and 2.2% to the RDI of manganese for men and women (see Table 4B). Decaffeinated coffee averaged 29.1 mg/kg and contributed 2.5% to the RDI of men and 3.2% to the RDI for women (See Table 4C).

Molybdenum ranged from below the limit of detection of 0.172 mg kg-1 in both the Nescafe Clasico Instant coffee and the Jamaica Mountain Peak decaffeinated coffee to 0.895 mg kg-1 in Maxwell House Instant coffee, with a mean concentration of 0.474 mg kg-1 (see Table 2). The lone Robusta brand, Shirriff Instant coffee, would contribute modestly to the RDI of both men and women at 2.3% per 2g serving size (see Table 4D). The average concentration of Jamaican instant coffee (0.460 mg kg-1) would contribute roughly 2.0% to the RDI of both men and women (see Table 4A).

Imported coffee would on average (mean 0.533 mg kg1) contribute 2.4% of the RDI for molybdenum (see Table 4B). Decaffeinated coffee averaged 0.401 mg kg

-1 and would provide only about 1.8% of the RDI (see Table 4C).

Selenium was detected in three samples, Folgers Instant (0.158 mg kg-1), Maxwell House Instant Coffee (0.201 mg kg-1) and Nescafe Clasico (0.290 mg kg-1). All other samples had variable limits of detection ranging from 0.100 mg kg-1 to 0.130 mg kg-1 (see Table 2). On average Jamaican soluble coffee, decaffeinated coffees and the Robusta sample would contribute approximately 0.4% to the RDI for both men and women in a 2g serving (see Tables 4A, 4C and 4D). The imported instant coffee averaged 0.217mg kg-1 and would provide 0.8% to the RDI for selenium in a 2g serving (see Table 4B).

Zinc, a trace metal of particular importance, ranged from 2.77 mg kg-1 in Nescafe Clasico Instant Coffee to 4.39 mg kg-1 in Coffee Roasters Jamaican Instant Coffee, with a mean of 3.54 mg kg-1.

Jablum Instant Coffee had zinc concentrations of 4.38 mg kg-1 (see Table 2). For a 2g serving of soluble coffee the highest provision to the RDI is 0.070% and 0.097% for men and women from Jamaican instant coffees (mean 3.86 mg kg-1). Imported coffees averaged 3.26 mg kg-1 and contributed 0.059% and 0.081% to the RDI for men and women respectively. Decaffeinated coffee averaged 3.12 mg Zn kg-1 and contributed 0.057% and 0.078% to the RDI for men and women respectively. The Robusta coffee brand Shirriff contributed 0.065% and 0.089% to the RDI for men and women respectively. The global prevalence of zinc deficiency is estimated at 31% 21. Zinc deficient soils or soil where zinc is not bioavailable may be the initial cause of zinc deficiency in diet. This may be exacerbated in the case of plants where absorption of zinc is hindered by high levels of fibre or antinutrients such as phytate. Instant coffee however is not a beverage that can be considered a source of dietary zinc.

Rubidium Rubidium ranged from 24.4 mg kg-1 in Jablum Instant Coffee to 68.7 mg kg-1 in Salada Mountain Peak Instant Coffee; the mean concentration was 53.2 mg/kg. The essentiality of rubidium has not been determined although evidence suggests a number of roles in various human systems. Rubidium is an alkali metal with the same oxidative state as potassium and thus may serve as an analogue for this metal in plant and other biological systems. Rubidium tends to be ubiquitous in vegetables and plant material and rubidium deficiency is not expected in the human diet. Coffee is known to have significant rubidium concentration and the bever-

JAMAICAN JOURNAL OF SCIENCE & TECHNOLOGY, Volume 26 ISSN: 1016-2054 | Dietary Intake of Trace and Heavy Metals from Consumption... age has as much as 4380 µg Rb L-1 litre. Estimates suggest a provisional daily requirement of 400 µg or less for rubidium22.

Based on Table 5 the contribution to the suggested daily intake would range from at least 12.2% to 34.4%. This would reaffirm that coffee is a fairly significant source of rubidium even in a 2g serving, should this ever be established as an ultra-trace nutrient.

Other Trace Elements Concentrations of Ba, Ce, Eu, La, Sm and Th were below the detection limits for almost all samples. While the levels of Sc were measurable, they are indeed quite low and do not warrant further discussion. While some of these elements are not known to have any defined physiological functions for humans, the prevailing wisdom may change with emerging research. It is notable that all samples had measurable concentrations of Cs, although at very low levels.

Table 5: Rubidium Content of Instant Coffee Samples and % RDI based on estimated RDI

CONCLUSIONS Moderate coffee consumption has been associated with benefits to human health, namely reduced risk of cardiovascular disease and Type 2 diabetes23. Instant coffees are not a significant dietary source of heavy metals and contribute a very low percentage of the Provisional Tolerable Weekly Intake for all potentially toxic elements examined. The dietary intake of most essential elements from instant coffees is also negligible, although moderate consumption would contribute some amount of potassium, magnesium, manganese and molybdenum to the Recommended Dietary Intake of those elements. Conversely, instant coffee is known to have polymers other than phytate which act as chelating agents for zinc, iron and copper24. This gives instant coffee additional anti-nutrient properties.

This may not be of significance except with high levels of consumption, however. Given that Jamaicans’ per capita coffee consumption is slightly less than 1 g day-13 even if all coffee consumption was soluble coffee its contribution to trace element nutrition or to potentially toxic element exposure would be for the most part insignificant to the Jamaican population.

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International Coffee Organization. World Coffee Trade. 2015. [cited March 17, 2015]. Available from: http:// ico.org/trade_e.asp?section=About_Coffee#sthash.dKu3SeWB.dpuf

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Jamaica Observer. Mavis Bank launches Tru Brew. 09 July 2014[cited March 17, 2015].Available from: http:// www.jamaicaobserver.com/business/Mavis-Bank-launches-True-Brew_17119866

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Coffee Industry Board of Jamaica. Coffee Industry Board of Jamaica Annual Report and Statement of Accounts 2012-2013. 2014.

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Global Coffee Report. Instant coffee consumption in emerging markets. March 2013 [cited March 17, 2015]. Available from: http://gcrmag.com/market-reports/view/instant-coffee-consumption-in-emerging-markets

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Ferdman RA. Almost half of the world actually prefers instant coffee. The Washington Post, 14 July 2014 [cited March 17, 2015]. Available from: http://www.washingtonpost.com/blogs/wonkblog/wp/2014/07/14/almost-halfof-the-world-actually-prefers-instant-coffee/

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Euromonitor International. Instant Coffee: Versatility and Convenience Drive Growth. 09 July 2014 [cited March 17, 2015]. Available from: http://blog.euromonitor.com/2014/07/instant-coffee-versatility-and-convenience-drivegrowth.html

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Lalor G. A Geochemical Atlas of Jamaica. Canoe Press, Kingston, Jamaica, 1996.

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Howe A, Hoo Fung L, Lalor G, Rattray R, Vutchkov M. Elemental composition of Jamaican foods. 1: A survey of five food crop categories. Environmental Geochemistry and Health. 2005; 27: 19–30.

9.

Lalor GC, Vutchkov MK, Grant C, Preston J, Figueiredo AMG, Favaro DIT. INAA of biological materials using the SLOWPOKE-2 Reactor in Jamaica. Journal of Radioanalytical and Nuclear Chemistry. 2000; 244 (2): 263–266.

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Kennedy G, St-Pierre J. NAA with the improved relative method and the interactive computer program EPAA. Journal of Radioanalytical and Nuclear Chemistry. 1993; 169 (2): 471–481.

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Joint FAO/WHO Expert Committee on Food Additives. Evaluation of Certain Food Additives and Contaminants: Sixty-seventh Report of the Joint FAO/WHO Expert Committee on Food Additives. WHO Technical Series No. 940. World Health Organization/Joint FAO/WHO Expert Committee on Food Additives, Geneva, Switzerland, 2007.

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Joint FAO/WHO Expert Committee on Food Additives. Evaluation of Certain Food Additives and Contaminants: Seventy-second Report of the Joint FAO/WHO Expert Committee on Food Additives. WHO Technical Series No. 959. World Health Organization/Joint FAO/WHO Expert Committee on Food Additives, Geneva, Switzerland, 2010.

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Ratnaike RN. Acute and chronic arsenic toxicity. Postgraduate Medical Journal. 2003; 79: 391–396.

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Hall AH, Shannon MW. Haddad and Winchester’s Clinical Management of Poisoning and Drug Overdose, 4th ed.

Saunders, PA, USA, 2007.

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United States Department of Agriculture/United States Department of Health and Human Services. Dietary Guidelines for Americans 2010. Washington, DC, USA, 2010.

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Oliveira M, Casal S, Morais S, Alves C, Dias F, Ramos S, Mendes E, Delerue-Matos C, Oliveira BPP. Intra- and in-

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terspecific mineral composition variability of commercial coffees and coffee substitutes: Contribution to mineral intake. Food Chem. 2012; 130: 702–709.

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McKenzie JM. Content of phytate and minerals in instant coffee, coffee beans and coffee beverages. Nutrition Reports International. 1984; 29: 387-395.

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United States Department of Agriculture/United States Department of Health and Human Services. Dietary Guidelines for Americans 2005. Washington, DC, USA, 2005.

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Fernandes AP, Santos MC, Lemos SG, Ferreira MMC, Nogueira AA, Nobrega JA. Pattern recognition applied to mineral characterization of Brazilian coffees and sugar-cane spirits. Spectrochimica Acta Part B. 2005; 60: 717724.

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World Health Organization. World Health Organization: Micronutrient deficiencies- Iron deficiency anaemia.

2015 [cited March 17, 2015]. Available from: http://www.who.int/nutrition/topics/ida/en/

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Caulfield LE, Black RE. Zinc deficiency. In: Comparative quantification of health risks: global and regional burden of diseases attributable to selected major risks. Edited by Ezzati M, Lopez AD, Murray CJ, Rodgers A. Geneva: World Health Organization. 2004; 257-279.

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Anke M, Angelow L, Muller R, Anke S. Recent progress in exploring the essentiality of the ultratrace element rubidium to the nutrition of animals and man. Biomedical Research on Trace Elements. 2005; 16 (3): 203–207.

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Ranheim T, Halvorsen B. Coffee consumption and human health – beneficial or detrimental? – Mechanisms for effects of coffee consumption on different risk factors for cardiovascular disease and type 2 diabetes mellitus.

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Wen X, Takenaka M, Murata M, Homma S. Antioxidative activity of zinc-chelating substance in coffee. Bioscience, Biotechnology and Biochemistry. 2004; 68: 2313-2318.

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5. Assessment of Cadmium, Mercury and Lead Concentrations in Jamaican Conch, Lobster, and Tilapia Leslie A. Hoo Fung1, Vaughn R. Rattray2 and Gerald C. Lalor1 1 International Centre for Environmental and Nuclear Sciences, University of the West Indies, Kingston, Jamaica 2 Department of Chemistry, University of the West Indies, Kingston, Jamaica ABSTRACT: The islands of the Caribbean basin depend on fisheries for local food and for earning foreign exchange through export. The concentrations of cadmium, mercury and lead in Jamaica are reported for samples of Queen Conch (Strombus gigas) and Caribbean spiny lobster (Panuliris argus/P. guttatus) as well as mercury in Snapper fish (Lutjanus campechanus) and compared with Tilapia fish (Oreochromis spp.) which are grown in artificial ponds with high water quality and fed high protein material which would be low in heavy metals. Cadmium, mercury and lead were below the limit of detection in all tilapia samples. Mean concentrations of cadmium, mercury and lead in conch were 0.74, 0.05 and 0.11 mg kg-1 fresh weight, respectively; means in lobster were 0.02, 0.04 and 0.09 mg kg-1 fresh weight, respectively.

Keywords: Jamaica, seafood, cadmium, lead, mercury, tilapia INTRODUCTION Jamaica has a population of some three million persons and because of geology and climate has a remarkable geochemistry1 including some soils with enhanced concentrations of trace elements, some of which are regulated by international agreements. Like numerous countries in or bordering the Caribbean, fisheries are highly important both for local consumption and for export. There are queen conch fisheries in the Bahamas, Turks and Caicos Islands, Cuba, Belize, the Dominican Republic, Mexico and Jamaica. Countries with spiny lobster fisheries include Belize, Jamaica, the Bahamas, and Cuba, The resources have been overexploited, in part because of poaching, and in many countries the rate of harvest is not sustainable2.

The spiny lobster (Panuliris argus / P. guttatus), queen conch (Strombus gigas) and tilapia (Oriochromis spp.) are important export products for Jamaica. Steps have been taken to protect these resources; for example fishing boats must be registered under the Fishing Industry Act (1975) and there are closed seasons for lobster (1 April to 30 June) and queen conch (usually 1 July to 31 October).

A National Total Allowable Catch (NTAC) is based on data on the stocks of conch in the Pedro Banks area (about 80km south of the island with a mean depth of 25m) from which most of Jamaicaâ&#x20AC;&#x2122;s conch harvest is tak-

en. Jamaica is allowed, under the Convention for International Trade in Endangered Species of Wild Fauna and Flora (CITES) to export conch based on good resource management3.

Freshwater aquaculture began in Jamaica in 1949, and several species of tilapia and carp have been introduced for pond cultivation. Jamaica had a tilapia production estimated at some 4200 tonnes in 1998, and 1300 tonnes of that being exported to the USA and UK4. Tilapia accounts for some 95% of Jamaicaâ&#x20AC;&#x2122;s aquaculture production5. In addition, some 5600 metric tones (mt) of tilapia were consumed locally in 20076.The tilapia are raised in artificial ponds, fed a high protein diet, typically pellets made of corn and fishmeal and the water quality is monitored. As the elemental intake is controlled the levels of heavy metals are low.

Fish, crustaceans and molluscs exported from Jamaica in 2014 were valued at US$12,587,0007, up from US$10,998,000 in 2013. With an increased international interest in the regulation of trace metal concentrations, in particular the potentially toxic cadmium, mercury and lead, it is therefore important to ensure that levels are in adherence to the regulations of the importing countries.

Fish and shellfish are an important part of human diet as they contain high quality protein, omega 3-fatty acids and

JAMAICAN JOURNAL OF SCIENCE & TECHNOLOGY, Volume 26 ISSN: 1016-2054 | Assessment of Cadium, Mercury and Lead Concentrations in Jamaican Conch, Lobster and Tilapiaâ&#x20AC;Ś...

other nutrients that contribute to cardiac health and childrenâ&#x20AC;&#x2122;s growth and development. But fish, particularly large fish at the top of the food chain tend to accumulate trace metals at levels that represent risks for some persons, particularly pregnant women and young children. This study surveys the present situation with regard to heavy metals and sets the stage for further work.

Table 1: Results of analysis of Certified Reference Material ERM CE-287R (mg kg-1)

METHODOLOGY Frozen tilapia fillets, lobster meat, and conch samples were received as subsamples from export shipments. Snapper were purchased from fish markets on the south side of the island, transported in ice and frozen in the laboratory. The frozen samples were thawed and homogenized with a blender before analysis. Approximately 1g fresh weight was weighed into a 70ml polyethylene vial with a hinged cap (Capitol Vial, Alabama) and 20ml of reverse aqua regia (1:3 v/v Trace Metal Grade hydrochloric: nitric acids, Fisher Scientific) was added to each vial and then covered with a polyethylene watch glass and left overnight. The following day, samples were digested in a graphite heating block (ModBlock, CPI International) at 1100C for 2 hours, cooled and made up to 50ml using 2% HNO3.

The range, mean and number of samples for all sample types are shown in Table 2 Table 2: Concentrations (mg kg fresh weight) for cd, Hg Pb in Conch, Lobster and Tillapia

Sample solutions were analyzed for Cd and Pb by Graphite Furnace Atomic Absorption Spectrophotometry (GFAAS), and for Hg by Cold Vapour Atomic Absorption Spectrophotometry (CV-AAS) using a PerkinElmer 5100PC Atomic Absorption Spectrophotometer. Quality control included duplicate analysis of 10% of samples. The reference material ERM-CE278r (Trace Elements in Mussel Tissue; Institute for Reference Materials and Measurements, Belgium) was included in each sample batch and was treated identically to the samples.

RESULTS AND DISCUSSION The agreement between measured and certified values of the reference material and the standard deviations, shown in Table 1, illustrate the accuracy and precision of the analytical methods used.

The fresh weight sample detection limits are also shown in Table 1. The detection limits reported are well below the elemental concentrations required by the regulations.

Cadmium Cadmium is generally considered to be extremely toxic, contributing to many conditions such as hypertension and cardiac and renal diseases which are of increasing prevalence in Jamaica and in many other countries. Cadmium accumulates in the tissues of crustaceans and mollusks, and consumption in large amounts can ultimately lead to accumulation in the body. Cadmium uptake may be enhanced by the dietary absence of certain nutrients such as iron and zinc, and retention may also depend on the age of the subject. High concentrations of cadmium are found in Jamai-

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can soils1 and it is possible that this is somehow reflected in foods obtained in Jamaican waters. Health effects, however, have not to date been observed in the Jamaican population [8] nor in New Zealand, where the dredge oyster (Tiostrea lutaria) from Bluff, Southland accumulates cadmium so well that consumption of a single one can more than double a normal daily dietary intake of cadmium for an adult. Yet a survey of 75 adults there, though they had high cadmium intakes, with the highest intake equivalent to more than 10 times the Provisional Tolerable Weekly Intake (PTWI) of 7 µg Cd/kg body weight set by Joint FAO/WHO Expert Committee on Food Additives9 showed no obvious health effects10.

The Codex Alimentarius prescribes a maximum level (ML) of 2 mg kg-1 Cd in mollusks and cephalopods11. The conch and lobster samples analysed are within this limit, with most samples well below it. While a few conch samples had elevated cadmium levels, this may be a function of the particular area in which these outliers were harvested. There is no prescribed limit for fish, and particularly pond-raised fish; all the tilapia samples had Cd concentrations below the limit of detection 0.005 mg kg-1.

Lead The toxicity of lead has been known from antiquity. Its role as neurotoxin and its effect on young children are of special concern. Although there have been numerous cases of lead poisoning due to improper disposal of used batteries and recycling of lead, it is not very soluble and no cases of unintentional poisoning due to food seem to have been reported.

The maximum limit (ML) for lead in fish set by the Codex Alimentarius is 0.3 mg kg-111, and as with mercury, MLs for other seafood or aquaculture products are not directly addressed. It is noteworthy, however, that the lead levels in the conch, lobster and tilapia caught in Jamaican waters is well below that level, with the unexplained exception of a single conch sample. These fisheries products would not be expected to make a significant contribution to the PTWI of 25 µg Pb/kg body weight9.

Mercury The toxicity of mercury has been known for centuries and the Minamata poisoning brought it forcibly to attention in recent times12. The element is now carefully regulated in developed countries and is being phased out from use in applications such as electrochemical processes, laboratory instruments, dental amalgam and gold refining. Elemental mercury, no doubt because of its extremely low vapour pressure is hardly toxic, but in the form of its methyl compounds it can

be highly damaging to humans. It is perhaps the best known of the trace element toxins in fish as it has been the topic of various advisories to restrict the consumption of some varieties of fish, especially by pregnant women and children. The range of mercury concentrations varies widely in fishes where it occurs almost entirely in the methylated form although it is usually reported as total mercury. Because they are among the top predators in the ocean, large pelagic fish like king mackerel, shark, swordfish, can show mean levels that exceed 0.9 mg/kg.

Mercury from fish and other seafood may account for as much as 40% of dietary mercury intake13. The ML for methyl mercury in fish and fish products is 0.5 mg kg-111. The levels of mercury found in these sample types were not significant, were well below the ML, and would not be expected to contribute significantly to the PTWI of 4 µg Hg/kg body weight9. Monitoring is recommended, however, particularly for large fish.

A small number (4) of snapper samples collected from the south coast of Jamaica contained on average 0.27 mg kg-1 fresh weight. While this is well below the ML, it may be significant given that the snapper analysed were relatively small fish. Further work is indicated.

CONCLUSIONS Of the sample types examined, tilapia is most consumed on the island. Conch and lobster, being high value products, are largely exported. This is also the case in other Caribbean territories which are producers of these fisheries products.

Based on these findings, and with the relatively low levels of consumption, it is unlikely that any of these would contribute significantly to dietary intake of cadmium, lead and mercury.

Periodic monitoring, especially of larger ocean fish, is nonetheless recommended.

We know of no cases in which trace metal concentrations have led to human illness in Jamaica [8] but this is a new field in the Caribbean and any problems may well be location specific so much more data are being gathered on the elemental composition of foods.

ACKNOWLEDGEMENTS The samples for analysis were provided by the Veterinary Services Division of the Ministry of Agriculture and Fisheries, Jamaica.

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REFERENCES 1.

Lalor G. A Geochemical Atlas of Jamaica, Canoe Press, Kingston, Jamaica, 1996.

2.

National Oceanic and Atmospheric Administration (NOAA), Fisheries, Office of Protected Resources. Queen Conch[cited October 15, 2015]. Available from: http://www.nmfs.noaa.gov/pr/species/invertebrates/queenconch.html

3.

Food and Agricultural Organization. Information on Fisheries Management in Jamaica [Internet]. October 2005. [cited October 15, 2015]. Available from: http://www.fao.org/fi/oldsite/FCP/en/JAM/body.htm

4.

Fitzsimmons K, Costa-Pierce BA, Rakocy JE, eds. Tilapia Aquaculture in the Americas, Vol. 2. The World Aquaculture Society, Baton Rouge, Louisiana, United States. Chapter: Future trends of tilapia aquaculture in the Americas. 2000; p. 252â&#x20AC;&#x201C;264.

5.

Ministry of Agriculture and Lands, Jamaica (MOA), Fisheries Division. Draft Fisheries Policy. 2008.

6.

Ministry of Agriculture and Fisheries, Fisheries Division, Jamaica and Statistical Institute of Jamaica (MOA/ STATIN).Source and Consumption of Fish in Jamaica 2001-2007.

7.

Statistical Institute of Jamaica (STATIN). Traditional and Non-Traditional Domestic Exports [cited October 15, 2015]. Available from: http://statinja.gov.jm/Trade-Econ%20Statistics/InternationalMerchandiseTrade/ newtrade.aspx

8.

Lalor GC. Review of cadmium transfers from soil to humans and its health effects in the Jamaican Environment. Science of the Total Environment. 2008; 400: 162-172.

9.

Joint FAO/WHO Expert Committee on Food Additives. Evaluation of Certain Food Additives and Contaminants: Sixty-seventh Report of the Joint FAO/WHO Expert Committee on Food Additives. WHO Technical Series No. 940. World Health Organization/Joint FAO/WHO Expert Committee on Food Additives, Geneva, Switzerland, 2007.

10.

McKenzie-Parnell JM, Kjellstrom TE, Sharma RP, Robinson MF. Unusually high intake and fecal output of cadmium, and fecal output of other trace elements in New Zealand adults consuming dredge oysters. Environmental Research. 1988; 46 (1): 1-14

11.

Codex Alimentarius Commission. Codex General Standard for Contaminants and Toxins in Food and Feed.

Codex Stan 193-1995. 2010.

12.

Ekino S, Susa M, Ninomiya T, Imamura K, Kitamura T. 2007. Minamata disease revisited: an update on the acute and chronic manifestations of methyl mercury poisoning. Journal of the Neurological Sciences. 2007; 262 (1-2): 131-44.

13.

World Health Organisation. Trace Elements and Human Nutrition. Geneva, Switzerland, 1996.

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Research in Progress Our Young Scientistsâ&#x20AC;&#x2122; Abstracts PRODUCTION OF SOLANUM tuberosum L MICROTUBER USING TEMPORARY IMMERSION SYSTEM Shischa Higgins, Donnett Jackson, Sandia McAnuff, Ryan Francis and Ann Marie Smith Scientific Research Council, Jamaica

BACKGROUND

METHODS

Jamaica aims at being self-sufficient in producing table Irish potato. Tissue culture is a means of propagating Solanum tuberosum L. plantlets for farmers; however, the disadvantage with this technique is the low yield of tubers and the small tuber size. Tissue cultured plants cannot be stored or transplanted directly into the field without an acclimatization stage; by comparison, liquid cultures are advantageous in mass producing microtubers which are easier to handle and transport by farmers who find it difficult to grow tissue culture irish potato plantlets - hence the need to produce microtubers which farmers are accustomed to growing. In the tuber induction stage, microtubers can be induced at all plant nodes indicating that tuberization is not restricted to specific regions (JimĂŠnez et al., 1995). The aim of this study is to investigate the difference between microtubers produced from 25 nodal segments and 50 nodal segments using 1000 mL of nutrient media.

Microtuber formation was developed using a temporary immersion system for potato microtuber production using Spunta 58 70 77 cultivar. The first step involved a multiplication stage on which tissue culture plantlets were grown on Murashige & Skoog (MS) multiplication media for three weeks. After three weeks the medium was replaced with a tuber MS induction medium for six weeks and plants were subjected to dark conditions. Each system contained 1000 mL of medium and one vessel was inoculated with 25 nodal segments and the other with 50 nodal segments.

25 nodal segments represent the least number of nodes used to produce microtubers. The quantity of tubers per vessel was determined as well as the fresh weight (g), length (cm) and diameter (cm) of microtubers per vessel.

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RESULTS AND DISCUSSION

CONCLUSION

The design of this system included 25 and 50 nodal segments in separate glass vessels, each containing 5L of MS media. After 2 months the average diameter (0.58 ± 0.2 cm) of the microtubers in the vessel containing 50 nodal segments was significantly higher (p<0.05) than the diameter (0.39 ± 0.03 cm) of the microtubers produced in the vessel containing 25 nodes. Similar, the fresh weight (0.42 ± 0.03 g) and length (1.17 ± 0.07 cm) of microtuber in the vessel containing 50 was significantly higher than the fresh weight (0.28 ± 0.069 g) and length (0.90 ± 0.139 cm) obtained in the vessel containing 25 nodal segments. The results obtained were comparable to other international studies.

The study revealed that when producing Solanum tuberosum microtubers via a temporary immersion system, 50 nodal segments may be used rather than 25 nodal segments to produce mircotubers that may be stored or handed over to farmers for propagation.

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GUIDE FOR AUTHORS Authorship of the paper submitted should be clearly indicated based on each author’s contribution to the paper. A covering letter must be attached to the paper confirming the following;     

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JAMAICAN JOURNAL OF SCIENCE & TECHNOLOGY, Volume 26 ISSN: 1016-2054 |

Jamaican Journal of Science & Technology Volume 26 | December 2015| Pages 7-56 JJST ISSN: 1016-2054 Published by: The Scientific Research Council, Information Services Division, Hope Gardens Complex, Kingston 6, Jamaica W.I.

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Jamaican Journal of Science & Technology Volume 26 | December 2015| Pages 7-56 JJST ISSN: 1016-2054 Published by: The Scientific Research Council, Information Services Division, Hope Gardens Complex, Kingston 6, Jamaica W.I.