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Revue Africaine de Santé et de Productions Animales © 2005 E.I.S.M.V. de Dakar
A RTICLE ORIGINAL The effects of force-feeding on sulphbromophtalein and indocyanine green pharmacokinetics in mullard ducks T. BENGONE-NDONG1*, S. BAKOU2, G. BENARD3, P. BENARD3,D. PREHN3 1 Institut de Recherches Agronomiques et Forestières (IRAF), BP 2246 Libreville Gabon. 2 Ecole Inter-Etats des Sciences et Médecine Vétérinaires (EISMV), Service d’Anatomie – Histologie, BP 5077 Dakar Fann Sénégal. 3 Ecole Nationale Vétérinaire de Toulouse, 23, chemin des Capelles 31076 Toulouse cedex France.
*
Correspondance et tirés à part, e-mail : tbengone@refer.ga
Abstract Disposition kinetics of Sulphobromophthalein (BSP) and Indocyanine Green (ICG) were used to evaluate hepatic function in mullard ducks with progressive fatty liver produced by force feeding. Two groups of 10 birds with fatty livers (after 3, 7 and 14 days of cramming period) and 1 control group were studied. Blood samples were collected for 180 minutes at predetermined times after the administration of an intravenous bolus (14 mg of BSP ; 0.74 mg of ICG per kilogram of body weight). Plasma dyes were analyzed spectrophotometrically. Pharmacokinetic variables were calculated using a two compartment open model. Volumes of distribution (p<0.001), area under the curve (p<0.05), half life of b phase (p<0.001) and mean residence time (p<0.001) were significantly decreased for both dyes. ICG clearance was significantly decreased. This study proved that the measurement of BSP and ICG disposition is suitable for assessing hepatic function and dysfunction in ducks with progressive fatty liver. These results also demonstrate that lipid accumulation within the hepatocytes modifies the excretion ratio of xenobiotics in the blood. (RASPA 3 (3-4) : 170-173).
Key – Words: Duck - Liver function - Force-feeding - Sulphobromophthalein - Indocyanine green Résumé Les effets du gavage sur la pharmacocinétique de la Bromosulfonephthaléine (BSP) et du vert d’Indocyanine chez les canards mulards L’exploration de la fonction hépatique a été envisagée au moyen de la cinétique de la Bromesulfonephthaléine (BSP) et du Vert d’Indocyanine (ICG) chez des canards mulards pendant le gavage. L’étude a été conduite sur deux lots de 10 animaux : un lot de canards maigres et un lot de canards en cours de gavage (aux jours 3, 7 et 14). Les doses adminitrées par voie intraveineuse étaient de 14 mg/kg pour la BSP et 0,74 mg/kg pour l’ICG. Dans les deux cas, le sang a été prélevé par ponction de la veine alaire pendant 180 minutes. Les plasmas ont été analysés par des méthodes spectrophotométriques. Les courbes exprimant les concentrations plasmatiques de BSP et d’ICG en fonction du temps ont été ajustées à un modèle pharmacocinétique ouvert à deux compartiments. Dans les deux cas, on observe une augmentation significative des volumes de distribution (p<0,001), des aires sous la courbe (p<0,05), des demi-vies d’élimination (p<0,001) et des temps moyens de séjour. La clairance diminue de façon significative dans le cas du vert d’indocyanine. Cette étude montre que la BSP et l’ICG peuvent être utilisés dans l’exploration de la fonction d’épuration hépatique chez le canard. Ces résultats démontrent également que l’accumulation des lipides dans les hépatocytes provoque une modification de la pharmacocinétique des xénobiotiques.
Mots-clés : Canard - fonction hépatique - Gavage - Bromesulfonephthaléine - Vert d’indocyanine
Introduction According to several authors, the fatty liver obtained by force feeding of ducks is probably a result of nutritional fat accumulation in the liver cells [16], [17], [18]. Cramming causes a large increase in the total proteins content, RNA and phospholipids in the liver [18]. The relative proportions and fatty acid compositions of the phospholipids are not altered. These phenomena seem to sow that the size of the liver cells increases during cramming ; these cells are the sites of intense and increasing metabolic activity at least until the middle of the cramming period. The purpose of this study was to see whether BSP and ICG disposition kinetics could be used to detect changes in hepatic function in ducks at various stages of liver fat accumulation. It is clear from studies by CAMPBELL [7], CLARKSON [10], CLARKSON and RICHARDS [11], [12], RAMACHANDRAN et al. [21] and BENGONE-NDONG et al. [3], [4] that the BSP test is suitable for the evaluation of liver function in birds. The disposition of ICG was also described [3], [4], [12]. RASPA Vol.3 N 03-4, 2005
Materials and Methods 1. ANIMALS
Three groups of 10 mullard ducks (Cairina moschata x Anas platyrhynchos) 14 weeks old, weighing about 4.2 kg were used for the experiment. The birds of the first group (controls) were housed in a single large room (25 m2). They were individually fed 200 g per day of a commercial duck starter diet. This group were given BSP and ICG with a two-week washout period between treatments. The birds in groups 2 and 3 were housed in individual cages and provided with individual waterers. The cages were located in an insulated building. After a prefattening programme, birds were force-fed twice daily with steamed maize for 14 days. Feed consumption was 340 g at the beginning and 530 g at the end of fattening.
2. EXPERIMENTATION 2.1. Compounds
Sulphobromophthalein was stored as a sterile 30‰ aqueous solution in glass ampoules. Indocyanine Green is unstable in aqueous solution and was distributed as a freeze-dried powder (50 mg) in a sterile flask. The solution therefore prepared just before administration. Both molecules were distributed by SERB, pharmaceutical laboratory (75020, Paris FRANCE).
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2.2. Administration and blood samples The birds were treated intravenously (right wing vein) with BSP or ICG at a dosage of 14 mg or 0.74 mg per kilogram of body weight. It was previously demonstrated that doses between 7 and 15 mg.kg-1 for BSP and 0.5 and 1 mg.kg-1 for ICG are suitable for assessing hepatic uptake-elimination function in ducks since relationship between administrated doses and pharmacokinetic parameters are linear [3]. Blood samples (2 ml) were collected into heparinized tubes from the left wing vein 2.5, 5, 7.5, 10, 12.5, 15, 17.5, 20, 25, 30, 40, 60, 90, 120 and 180 minutes after dye administration. The samples were centrifuged and the plasma immediately analysed for BSP or ICG concentration.
K 12
Centra l Compartment BLOOD
Peripheral Compartment LIVER
K 21
(1)
(2)
K 20
2.3. Assay procedure
The curves of drug concentrations in plasma plotted against time were fitted to a two-compartment open model for kinetic analysis. Such curves were described by the following exponential expression : C = Ae-at + Be-bt in which C is the plasma concentration at time t and A, B, a and b are constants. The pharmacokinetic models for BSP and ICG included an apparent first order elimination rate constant (k20) from the peripheral compartment (liver). All pharmacokinetic parameters were calculated using the SIPHAR program (version 4.0.b) : - Areas under the concentration-time curves : AUC = A/a + B/b - Total plasma clearance : Cl = dose/AUC - Mean residence time : MRT = (A/a2 + B/b2)/AUC - Apparent volume of distribution : Vb = dose/AUCxb - The b half-life : T 1/2 b = 0.693/b
Results The relationship between time and mean plasma concentrations of BSP and ICG in ducks at each stage of cramming were log-linear (figures 2 and 3). Plasma dye concentrations versus time curves were best represented by a biexponential model in the control group and after 7 and 14 days of cramming (figure 1). The values of the parameters describing the disposition kinetics of BSP and ICG are given in tables 1 and 2. The clearance of both BSP and ICG decreased as liver fattening increased, but the decline was not significant in the case of BSP. The volumes of distribution increased (p<0.001). Area under the curve differed between the different cramming stages (p<0.05 for BSP ; p<0.01 for ICG). The changes in T1/2b were reciprocal to volume of distribution.
Discussion In the present study, the hepatic uptake of BSP and ICG was examined to evaluate hepatic function in ducks with progressive fatty livers. BSP and ICG disposition were compared with those of control animals. No significant difference was observed between ducks that had been force-fed for 3 days and control birds. The modifications occurring in the liver of force-fed ducks are due mainly to an excess of triglycerides deposited in the
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Figure 1: The two-compartment open model associated with BSP and ICG parmacokinetics
1000 Concentration (mg.l-1 )
2.4. Data analysis
Elimina tion (BILE)
100 10 1 0,1 0,01
0
40
80
120
160
Time (minutes)
200
Figure 2 : Plasma concentrations of BSP after intravenous administration of 14 mg.kg -1. Data are expressed as mean Âą SD for ten ducks at day 0 ( ) and after 3 ( ), 7 ( ) and 15 days ( ) of cramming 100
concentration (mg.l-1 )
Plasma BSP concentrations were determined by reading the optimal density of 50 to 500 ml of plasma treated with 0.5 N sodium hydroxide (500 ml) at 584 nm against a control blank on a double beam Uvikon 860 spectrophotometer [14]. A set of standard samples prepared in plasma were assayed daily along with the other samples. ICG plasma levels were measured by means of a microassay adapted from a previously described method [22]. Plasma (200 ml) was mixed gently with 800 ml of cold acetonitrile in a 1.5 ml propylene tube. The samples were centrifuged at 3000 g for 5 minutes immediately after acetonitrile addition. The absorbance of each supernatant was measured at 784 nm using a double beam spectrophotometer (Uvikon 860). The optical densities were read against a blank plasma sample prepared as described above. A set of standard samples prepared in plasma were assayed daily along with the other samples.
10
1
0,1
0
40
80 120 Time (minutes)
160
200
Figure 3 : Plasma concentrations of ICG after intravenous administration of 0.74 mg.kg -1. Data are expressed as mean Âą SD for ten ducks at day 0 ( ) and after 3 ( ), 7 ( ) and 15 days ( ) of cramming
hepatic cell and partly to an increase in the total protein content [2], [19], [20]. The liver enlargement is a result of both hypertrophy and hyperplasia, as shown by the increase in the total amount of nucleic acids in the liver and the slight increase in the RNA/DNA ratio during the cramming period [18], [19]. The excess energy and the lower protein content in the diet of force-fed birds induced an increase in cell activity during the first week of force-feeding. This is why plasma BSP and ICG concentrations versus time curves were sometimes best represented by a monoexponential model after 3 days of cramming. The elimination of BSP and ICG from the plasma after 7 and 14 days of cramming was however significantly slower than in normal ducks as is shown by the high values of AUC, MRT and T1/2b. The ICG clearance at 14 days decreased by approximately 67%. The underlying reasons could be multiple and complex. Likely causes include reduced hepatic extraction and transport. RASPA Vol.3 N03-4, 2005
Effects of force-feeding on sulphbromophthalein and indocyanine green pharmacokinetics in mullard ducks
Table 1. BSP pharmacokinetic parameters in ten mullard ducks after a single intravenous administration of 14 mg/kg of body weight after 3, 7, and 14 days of cramming. Significant difference from control : * p<0.05 ; ** p<0.01 ; **** p<0.005 Parameters A (mg/l) α (l/min) B (mg/l) β (l/min) K 12 (l/min) K 20 (l/min) K 21 (l/min) T 1/2 α (min) T 1/2 β (min) Vβ (I) Vss (I) AUC (min*mg/l) MRT (min) Cl (ml/min/kg)
control 277 ± 43 0.22 ± 0.02 9.2 ± 4.3 0.04 ± 0.01 0.22 ± 0.02 0.04 ± 0.01 0.004 ± 0.002 3.1 ± 0.4 21 ± 8 0.31 ± 0.13 0.09 ± 0.02 1485 ± 245 8.6 ± 2.9 10.0 ± 0.2
3 days
7 days
216 ± 109** 0.21 ± 0.06* 11.3 ± 9.5 0.06 ± 0.04 0.19 ± 0.05 ** 0.07 ± 0.04 0.012 ± 0.020 3.5 ± 1.1 22 ± 18 0.36 ± 0.35 0.11 ± 0.07 1289 ± 363 7.4 ± 4.1 11.0 ± 0.3
191 ± 79 * 0.21± 0.04 7.5 ± 9.6 0.03 ± 0.02 * 0.20 ± 0.04 0.04 ± 0.03 0.004 ± 0.004 * 3.4 ± 0.7 35 ± 23 0.57 ± 0.36 * 0.15 ± 0.06 1077± 185 * 8.9 ± 4.4 11.0 ± 0.2
14 days 132 ± 44 *** 0.14 ± 0.03 ** 2.6 ± 1.2 * 0.01 ± 0.01** 0.13 ± 0.03 ** 0.01 ± 0.01 0.002 ± 0.001* 5.3 ± 1.2*** 122 ± 99 *** 1.80 ± 1.40** 0.65 ± 0.57*** 1434 ± 400 54.1 ± 48.8*** 9.0 ± 0.1
Table 2. ICG pharmacokinetic parameters in ten mullard ducks after a single intravenous administration of 0.74 mg/kg of body weight after 3, 7 and 14 days of cramming. Significant difference from control : * p<0.05 ; ** p<0.01 ; *** p<0.005. Parameters A (mg/l) α (l/min) B (mg/l) β (l/min) K 12 (l/min) K 20 (l/min) K 21 (l/min) T 1/2 α (min) T 1/2 β (min) Vβ (I) Vss (I) AUC (min*mg/l) MRT (min) Cl (ml/min/kg)
control 56.8 ± 11.5 0.24 ± 0.04 2.67 ± 0.76 0.028 ± 0.006 0.23 ± 0.03 0.030 ± 0.006 0.009 ± 0.003 2.9 ± 0.5 26 ± 7 0.08 ± 0.02 0.03 ± 0.01 334 ± 70 13 ± 3 2.2 ± 0.4
Several investigations have reviewed the relationship between hepatic disease and drug disposition in man [13], [15] and mammals [1], [5], [6], [8], [9]. Such reviews have been based on studies of the effects of hepatic disease on the elimination of dugs dependent on a single major determinant of hepatic clearance i.e. hepatic blood flow or the intrinsic metabolism. Effects of hepatic disease on hepatic blood flow have been studied by use of BSP and ICG, because the elimination of these dyes is flow limited. The results of this study indicate that force-feeding causes drastic changes in BSP and ICG dispositions. This is evidenced by observation of an increase in the volumes of distribution of both BSP and ICG. Increased hepatic mass probably contributes to the increased distribution of BSP and ICG. Differences in hepatic mass and volumes of distribution explain the differences in dye elimination at cramming stages and the resulting differences in AUC, T1/2β and MRT.
Bibliography 1. BACHMANN K.A., YANG C., JAHN D., SCHWARTZ J. 1988. The use of single sample clearance estimates to probe hepatic drug metabolism in rats I. Xenobiotica, 18, 151-159.
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3 days 23.7 ± 7.1*** 0.25 ± 0.04 2.60 ± 0.65 0.030 ± 0.009 0.23 ± 0.04 0.040 ± 0.010 0.019 ± .003*** 2.7 ± 0.5 23 ± 5 0.13 ± 0.02 0.07 ± 0.01 186 ± 27 16 ± 4 4.0 ± 0.6***
7 days 21.0 ± 9.4 *** 0.19 ± 0.06 ** 0.85 ± 0.17 *** 0.008 ± 0.001** 0.18 ± 0.06 ** 0.008± .002*** 0.007± 0.002 3.9 ± 1.3 89 ± 19* 0.40 ± 0.10 *** 0.23 ± 0.08 *** 232 ± 42 8.9 ± 4.4 3.0 ± 0.4 *
14 days 19.1 ± 8.4*** 0.16 ± 0.20 *** 1.83 ± 1.13 ** 0.004 ± .001*** 0.14 ± 0.10 *** 0.005± .002*** 0.020± 0.040 7.8 ± 1.6*** 173 ± 76 *** 0.31± 0.10** 0.20 ± 0.06*** 657 ± 319 54.1 ± 48.8*** 1.3 ± 0.5**
2. BENARD G., LABIE C. 1992. Composition chimique des foies de canard mulard au cours du gavage. Rev. Med. Vet., 143, 681-684. 3. BENGONE-NDONG T., BENARD G., PREHN D., COLAS E., GRIMM F., BENARD P. 1996. Etude de la cinétique de la bromesulfonephthaléine et du vert d’indocyanine chez le canard : détermination de la linéarité. Rev. Med. Vet., 147, 75-80. 4. BENGONE-NDONG T., BIAOU C.F., BENARD G., PREHN D., SAWADOGO G.J., BENARD P. 2003. Evaluation of sulphbromophthalein and indocyanine green liver function tests in ducks. Animalis, 2, 34-37. 5. BONASCH H., CORNELIUS C.E. 1964. Indocyanine green clearance. A liver function test for the dog. Am. J. Vet. Re., 25, 254-259. 6. BOOTHE D.M., BROWN S.A., JENKINS W.L., GREEN R.A., CULLEN J.M., CORRIER D.E. 1992. Indocyanine green disposition in healthy dogs and dogs with mild, moderate or severe dimethylnitrosamine-induced hepatic disease. Am. J. Vet. Res., 53, 382-388. 7. CAMPBELL J.G. 1957. Studies on the influence of sex hormones on the avian disease. I. Sexual differences in avian liver clearance curves. J. Endocrinol., 15, 339-345. 8. CENTER S.A., BUNCH S.E., BALDWIN B.H., HORNBUCKLE W.E. and TENNANT B.C. 1983a. Comparison of sulphobromophthalein, indocyanine green clearances in the dog. Am. J. Vet. Res., 44, 722-726. 9. CENTER S.A., BUNCH S.E., BALDWIN B.H., HORNBUCKLE W.E. and TENNANT B.C. 1983b. Comparison of sulphobromophthalein, indocyanine green clearances in the dog. Am. J. Vet. Res., 44, 727-730. 10.. CLARKSON M.J., 1961. Bromsulphthalein liver clearance in the bird. I : the relationship between plasma levels and bile and liver content in the turkey. Res. Vet. Sci., 2, 143-148.
172
T. BENGONE-NDONG et al. 11. CLARKSON M.J., RICHARDS T.G. 1967a. Bromsulphthalein liver clearance in the bird. II: transfer rates in normal turkeys. Res. Vet. Sci., 8, 48-52. 12. CLARKSON M.J., RICHARDS T.G. 1967. Steady-state plasma clearance of bromsulphthalein and indocyanine green measured by single injection. Res. Vet. Sci., 8, 454-462. 13. GILMORE I.T. 1986. Modern methods of diagnosis in liver disease, Journal of the Royal College of Physicians of London, 20, 201-205. 14. HARTMANN L. 1971. Clearance de la BSP, in: l’Expansion (Ed), Techniques modernes de laboratoire et explorations fonctionnelles, Paris, France,1971, pp 176-183. 15. KAWASAKI S., SUGIYAMA Y., IGA T., HANANO M., BEPPU T., SUGIURA M., SANJO K., IDEZUKI Y. 1988. Hepatic clearances of antipyrine, indocyanine green and galactose in normal subjects and in patients with chronic liver diseases. Clin. Pharmacol. Ther. 44, 217-224. 16. LABIE C., TOURNUT J. 1970. Recherches sur les modifications histologiques et biochimiques chez les oies soumises au gavage. Cah. Med. Vet., 39, 247-261. 17. LECLERCQ B., BLUM J.C. 1975. Etude de la suralimentation forcée : effets sur le métabolisme hépatique
et les formes de transport hépatiques chez le canard. Ann. Biol. Anim. Biochim. Biophys., 15, 559-568. 18. LECLERCQ B., DURAND G., DELPECH P., BLUM J.C. 1968. Note préliminaire sur l’évolution des constituants biochimiques du foie au cours du gavage. Ann. Biol. Anim. Biochim. Biophys., 8, 549-556. 19. NITSAN Z., NIR I., DROR Y., BRUCKENTAL I. 1973. The effects of forced feeding and of dietary protein level on enzymes associated with digestion, protein and carbohydrate metabolism in geese. Poult. Sci., 52, 474-481. 20. NIR I., PEREK M., KATZ Z. 1972. The influence of soybean meal supplemented to the maize diet of forced-fed geese upon their liver, organ and blood plasma components. Ann. Biol. Anim. Biochim. Biophys. 12, 77-89. 21. RAMACHANDRAN P.V., DAMODARAN S., CHANDRASEKARAN K.P. 1978. Liver function tests in fowls. Cheiron, 7, 93-98. 22. TUFENKJI A.E., ALVINERIE M., HOUIN G., GALTIER P. 1987. Pharmacokinetics of sulphobromophtalein, lidocaine and indocyanine green in the course of subclinical fascioliosis in sheep. Res. Vet. Sci., 43, 327-330.
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