Pharmacokinetics and tissue distribution of oxytetracycline in carp, Cyprinus carpio L., following different routes of administration

Abstract. The objective of this pharmacokinetic study was to investigate absorption, distribution, elimination and bioavailability of oxytetracycline (OTC) in carp, Cyprinus carpio L ., after different routes of administration, OTC was administered intravenously (i.v.), intramuscularly (i.m.) and orally at 60 mg/kg body weight. OTC levels were determined in plasma and several tissues. Analysis of the plasma drug concentration-time curves following i.v. OTC injection revealed three distinct phases. A three-compartment open model was used to derive pharmacokinetic parameters. Compared to mammals, a very extended final elimination half-life was observed (139.8±38.1 h). Following i.m. OTC administration, C_max was 56.8±10.9μg OTC/ml at 14 h post-injection. The Vd area was 2.1 ± 0.66 1/kg. Extreme differences were observed with respect to bioavailability following i.m. and oral administration; approximately 80 and 0.6%, respectively. Following i.m. injection tissue OTC determinations revealed that the drug was accumulating in pronephros, bone tissue and scales. After 21 days the OTC concentrations were 2.9±0.8, 5.2±0.3 and 4.7±3.1 μg/ ml, respectively. In tissue samples from the dorsal region (muscle), including the injection site, OTC could not be demonstrated at that time. The pharmacokinetic data are discussed in relation to the susceptibility of the immune system of fish for modulation.     

A Mammalian Growth Hormone-Releasing Hormone Increases Serum Growth Hormone Levels and Somatic Growth at Suboptimal Temperatures in Tilapia

The effects of a mammalian growth hormone-releasing hormone (GHRH), PrebGRF(1–78)OH (bGHRH), on growth and serum growth hormone (GH) levels were investigated in tilapias Oreochromis mossambicus and O. niloticus × O. aureus . Fish were injected intramuscularly or implanted intramuscularly (Silastic or cholesterol implants) with distilled water, 0.1 μg/kg bGHRH, 1.0 μg/kg bGHRH, 10.0 μg/kg bGHRH, or 100.0 μg/kg bGHRH and compared to untreated controls, fish implanted with 60 mg/kg 17α-methyltestosterone (MT), or a combination of bGHRH concentrations plus either MT, 0.01 μg/kg of a thyroid hormone (T₃), or 0.01 μg/kg of a glucocorticoid (DEX). The bGHRH increased serum GH levels in tilapia maintained at suboptimal temperatures (18 C). Serum GH levels were highest (5.3–0.45 ng/mL) for fish injected with 10.0 μg/kg bGHRH. Fish implanted with a Silastic implant containing 10.0 μg/kg bGHRH had significantly higher (4.35–0.35 ng/mL) serum GH levels than those with an equivalent dosage in a cholesterol implant. The addition of MT, thyroid hormones, and glucocorticoids did not increase serum GH levels above those obtained for fish receiving bGHRH alone. Tilapia reared at suboptimal temperatures and implanted with 10.0 μg/kg and 100.0 μg/kg bGHRH had significantly greater increases in weight and length than control fish. Fish implanted with bGHRH, MT or bGHRH plus MT had significantly higher moisture and protein content, while fat and ash contents were significantly lower than controls or sham-implanted fish. Fish implanted with bGHRH or bGHRH plus MT had significantly higher gonadosomatic indices than fish implanted with MT alone, shams or non-treated controls. This study demonstrates that a mammalian GHRH stimulates release of GH, promotes somatic and gonadal growth and may affect reproductive performance in tilapia.     

Multiple-dose pharmacokinetic and depletion studies of sarafloxacin in Atlantic salmon, Salmo salar L.

Abstract. Two multiple-dose pharmacokinetic and depletion studies with sarafloxacin hydrochloride in feed pellets were conducted with Atlantic salmon at 12.1 ± 1.1 †C and 9.3 ± 1.5†C, respectively. The dose regimens used were 10mg sarafloxacin kg^-1 biomass for 10 days, and 20mg sarafloxacin kg^-1 biomass for 5 days. In the 10-day study, the highest average concentrations of sarafloxacin in plasma, muscle and liver were 0.14, 0.39 and 0.88μgg(ml)^-1, respectively. In the 5-day study, the highest average concentrations in plasma, muscle, liver and skin were 0.40, 0.61, 1.56 and 0.19μg(ml)^-1, respectively. A comparison between the individual plasma concentrations and the corresponding tissue concentrations revealed significantly higher concentrations in tissue than in plasma during the depletion phase. A similar comparison made in the therapeutic phase from 3 days after first medication to one day after the last medication revealed significantly higher concentrations in liver and muscle than in plasma, and significantly lower concentrations in skin than in plasma. On withdrawal of the drug, sarafloxacin concentrations in plasma and the different tissues declined rapidly, Sarafloxacin was not detected in any plasma sample taken 6 days or more after the end of medication. The corresponding figures for muscle, skin and liver tissues were 14, 20 and 22 days, respectively. The half-lives of sarafloxacin varied in the different tissues included in the studies. The half-life was shortest in plasma, and increased in ascending order in muscle, liver and skin.     

Effects of Temperature on the Uptake and Depuration of 2-Methylisoborneol (MIB) in Channel Catfish Ictalurus punctatus

The microbial metabolite 2-methylisoborneol (MIB) imparts a muddy off-flavor to channel catfish Ictalurus punctatus . Uptake and depuration of MIB from fish are important considerations in the design and implementation of systems to remove off-flavors from fish prior to processing. The kinetics of MIB uptake by channel catfish were determined by placing fish in 6.5, 14.0, 25.0, and 34.0 C water containing approximately 1.0 μg/L chemically-synthesized MIB. Fish were sacrificed following 0, 2, 4, 8, and 24 h exposure to MIB. Fillet tissue samples were subjected to gas chromatographic and fat content analysis. The model for MIB uptake was:
MIB in fillet tissue (μg/kg) =−0.61 ± 4.2 [log( h ± 1)] ± 0.0076( T ) ( h ) ± 0.089( T ),
where h is the duration of exposure to MIB in hours and T is the water temperature in degrees C. The model accounted for 74% of the total variation observed in the tissue MIB concentrations and indicated that the fillet fat content was not strongly correlated with MIB uptake. To investigate the depuration of off-flavors, exposure to MIB was halted. Tissue samples were obtained 4, 8, 24, 48, and 72 h from fish held at 6.5, 14.0, 25.0, and 34.0 C. These data yielded the model:
MIB in fillet tissue (μg/kg) = 3.6 ± 0.176( T ) – 2.06 [log( h ± 1)] – 0.00296( T ) ( h ) ± 0.197 (% fat),
where h is the duration of exposure to MIB in hours, T is the water temperature in degrees C, and % fat is the % fat in the fillet tissue. The model accounted for 67% of the total observed variation in tissue MIB concentrations. Reducing the fat content of fish and optimizing water temperatures may augment MIB removal from fish tissues prior to processing.     

Measurement of Resting and Stress-elevated Serum Cortisol in Rainbow Trout Oncorhynchus mykiss in Experimental Net-pens

A commercially available heterogeneous, solid-phase tube enzyme-linked immunoassay (ELISA) was modified and validated for the measurement of serum cortisol in rainbow trout Oncorhynchus mykiss . The assay is accurate and precise. Resting and stress-elevated serum cortisol concentrations were measured in rainbow trout with a sensitivity of 1.5 ng/ml. Fish held in net-pens at a density of 0.4 kg/m³/cm had a resting cortisol level of 16.5 ± 3.8 ng/ml (mean ± SE). At 3 h postdisturbance, serum cortisol levels were not affected by the removal of fish from adjacent net-pens with dip nets or by the use of 200 mg/L tricaine methanesulfonate (MS-222) as an anesthetic for obtaining samples. However, an acute stress (60 s removal from water) elevated serum cortisol levels to 73.7 ± 9.4 ng/ml.     

Influence of ammonia on growth of Penaeus monodon Fabricius post-larvae

Abstract. Tiger shrimp, Penaeus monodon Fabricius, post-larvae (32·0±3mg, 1·43±0·03 cm) were exposed to control, 0·12, 0·60, 1·20 and 2·40mg/l ammonia-N (un-ionized plus ionized ammonia as nitrogen) which is equivalent to control, 6,32,63 and 126μg/1 NH₃-N (un-ionized ammonia as nitrogen) for 8 weeks in 25 ppt, as pH of 7·85–8·18 and 26–28°C by static renewal method. Growth in weight and length of the shrimps exposed to 1·20 and 2·40 mg/l ammonia-N were significantly lower (P < 0·05) than those exposed to control. The EC₅₀ (concentration that reduced growth by 50% of that of the controls) was 1·33 mg/l ammonia-N, 70 μg/l NH₃-N for weight gain, and 2·35 mg/1 ammonia-N, 123μg/l NH₃-N for length gain of P. monodon post-larvae. The'maximum acceptable toxicant concentration'(MATC) of ammonia-N and NH₃-N for P. monodon post-larvae was 0·60mg/l and 32μg/l, respectively after 6 weeks of exposure.     

The kidney of rainbow trout, Salmo gairdneri Richardson, in the acute phase of viral haemorrhagic septicaemia: in vivo experiments on the renal excretion of fluid, electrolytes and protein

Abstract. Kidney function was studied in unanaesthetized rainbow trout, Salmo gairdneri Richardson, with acute infection by VHS virus strain F-l causing viral haemorrhagic septicaemia. The experiments were performed in diseased and control fish with catheters chronically implanted in the dorsal aorta and the urinary bladder. Glomerular clearance of polyfructosan, tubular handling of fluid and electrolytes and urinary excretion of protein were studied. In diseased fish, plasma concentrations of inorganic solutes were reduced, but plasma osmolarity and systemic blood pressure remained unchanged. Glomerular filtration rate and urine flow, both linearly correlated, were reduced from 5.20±0.40 to 3.08±0.66 ml/h/kg and from 2.51±0.15 to 1.63±0.28 ml/h/kg, respectively (±SEM). The urine osmolarity was augmented due to increased Na and Mg concentrations in the urine. Net tubular secretion of Mg was observed in diseased fish whereas net reabsorption of Mg prevailed in the controls. Decreased renal protein excretion rate prevented a renal loss of protein and, therefore, diminished plasma protein concentrations in VHS-infected fish may be attributed to extrarenal pathophysiological mechanisms.     

Artificial Propagation and Induction of Triploidy in Largemouth Bass Micropterus salmoides and Ploidy Discrimination using Erythrocyte Length

We describe an artificial propagation procedure and simple ploidy discrimination techniques using erythrocyte major axis length for largemouth bass Micropterus salmoides. Hormonal treatments of 5 mg/kg of carp pituitary and 50 μg/kg of leutinizing hormone-releasing hormone (LHRH) produced viable gametes in 21-24 h, and triploidy was induced using a pressure treatment of 563 kg/cm² on embryos for a 1-min duration exactly 5 min following fertilization. We produced about 500 fingerling triploids and about 500 diploid controls, and verified genetic status of a subset of each group using flow cytometry. Erythrocyte length was measured for 10 known diploid and 10 known triploid individuals. Remaining fish were internally microtagged with group-specific tags and mixed to test the model. We developed ploidy discrimination intervals based on the 99% confidence limits of mean erythrocyte length (MEL, N = 25 erythrocytes) for individual fish, which were 14.43-16.66 μ.m for triploids, and 10.23-13.62 μm for diploids. Logistic regression provided the discrimination model: Ploidy status (±) = -196.16 + 13.97 x MEL, with positive (+) outcomes considered triploid. Both discrimination techniques were 100% effective at differentiating ploidy of the 22 microtagged largemouth bass recollected from the mixed population. We did not observe a significant change in erythrocyte length as fish size increased, indicating that erythrocyte length is an accurate predictor of ploidy for all sizes of largemouth bass.     

Influence of starvation, body size and temperature on ammonia excretion in the marine bivalve Tapes decussatus (L.)

Abstract. Ammonia excretion rate of the marine bivalve Tapes decussatus (L.) varied with body weight, temperature and starvation. There was a steady state in the excretion rates, in which these rates neither increased nor decreased during the first 6 days of starvation. The highest rates of ammonia excreted during the steady state (before decline to lower level) depended on the temperature (715 ± 86 and 395 ± 55 μg NH₄.N/clam/h × 10⁻²) at 28°C and 20°C, respectively. At 16°C, the steady state extended from 6h to 18d starvation. Ammonia excretion rates were higher for starved clams than for fed clams at all sizes, e.g. clams of 0·07 g dry flesh weight (28 ± 9 and 13 ± 5 μg NH₄.N/clam/h × 10⁻² respectively) at 16°C but not at 20°C and 28°C. At each temperature, weight-specific ammonia excretion rates were related to dry flesh weight of starved clams but were not related to fed ones.     

Effect of Luteinising Hormone-Releasing Hormone Analogue and Human Chorionic Gonadotropin on Ovulation, Plasma and Ovarian Levels of Gonadal Steroids in Greenback Flounder Rhombosolea tapirina

Female greenback flounder Rhombosolea tapirina were treated with either des Gly¹⁰ [D-Ala⁶] LHRH ethylamide (LHRH-a) at 50 or 100 pg/kg intraperitoneal injection (ipi), LHRH-a cholesterol pellet (LHRH-a pellet) 100 μg/kg implanted intraperitoneally, or human chorionic gonadotropin (hCG) 1,000 IU/kg ipi. Treatment with hCG, LHRH-a (50 μg/kg) ipi and LHRH-a pellet increased the total number of ovulations above control levels. LHRH-a (100 μg/kg) ipi was not consistently more effective than the control and both LHRH-a (50 μg/kg) ipi and LHRH-a pellet induced more ovulations than LHRH-a (100 μg/kg) ipi. Of those fish that ovulated, most ovulated more than twice, and most ovulations occurred at daily intervals. Oocyte diameters increased and oocyte stage advanced significantly in response to all exogenous hormone treatments. This was accompanied by increases in plasma and ovarian levels of 17β-estradiol (E₂, and in most cases, plasma and ovarian levels of testosterone (T). Plasma and ovarian levels of 17,20β-dihydroxy-4-pregnen-3-one (17,20βP) were not consistently elevated in association with reproductive events.     

Anoxia survival in common carp and crucian carp is related to high zinc concentration in tissues

ABSTRACT:   Ten lots of common carp and six lots of crucian carp (each lot of 100 fish) were treated under anoxia for five days at water temperatures of 25–31°C. The average per group mortality of common carp was 17%, but none of the 600 crucian carp died. The digestive tract tissues of the common carp that died had significantly lower zinc concentrations than those that survived (59 ± 41 vs 142 ± 60 μg/[g fresh tissue], P  < 0.001). The digestive tract tissues of the crucian carp had mean zinc concentrations of 652 ± 458 μg/(g fresh tissue). One lot of common carp that had low tolerance for anoxia was fed a high zinc diet (2000 mg zinc/kg diet) for 1, 2 or 6 months and then subjected to 5 days anoxia. The survival rates of those fed the high zinc diet 1 and 2 months increased from 0 to 50%, respectively; all of fish that had fed a high zinc diet for 6 months survived. Thus, anoxia survival in common carp and crucian carp is closely related to the high concentrations of zinc in their tissues.     

The influence of antibiotics on the immune system: immuno-pharmokinetic investigations on the primary anti-SRBC response in carp, Cyprinus carpio L., after oxytetracycline injection

Abstract. The present study was performed to obtain further details on the immuno-modulating effects of oxytetracycline (OTC) in carp, Cyprinus carpio L. The immuno-logical study was extended by investigating the pharmacokinetic behaviour of OTC. Oxytetracycline, at a dosage of 60 mg/kg, was administered five times intra-peritoneally at 3-day intervals; the first injection was given 1 day before immunization. The kinetics of the primary anti-sheep red blood cell (SRBC) response, during a short-term OTC treatment revealed that the kinetics of the plaque-forming cell (PFC) response was not significantly affected by OTC nor its solvent. In contrast, the number of plaque-forming cells was significantly decreased. The anti-SRBC antibody production was delayed by 2–4 days in both experimental groups (OTC and solvent). However, within 12–14 days post-immunization, the same antibody levels were detected as in the control group. High OTC plasma levels (>50 μg/ml) were detected about 10 h after a single i. p. injection. A mean plasma elimination half-life (T¹/₂) of 34·5 h was calculated.     

Anaesthetic efficacy and physiological responses to clove oil-anaesthetized kelp grouper Epinephelus bruneus

The efficacy of clove oil as an anaesthetic and at producing a physiological response (plasma cortisol and glucose) was evaluated in the kelp grouper, Epinephelus bruneus . To acquire complete anaesthesia in less than 3 min and recovery in <10 min, three doses of clove oil were tested at 18, 22 and 26 °C. Although higher anaesthetic doses resulted in shorter induction times and longer recovery times, and a lower temperature resulted in longer anaesthesia induction and slower recovery, we found the optimal dose and administering temperature of clove oil to be 250–300 mg L⁻¹ at water temperature of 18 °C, 150–200 mg L⁻¹ at water temperature of 22 °C and 50–100 mg L⁻¹ at water temperature of 26 °C respectively. Following the administration of 150 mg L⁻¹ of clove oil at 22 °C, the plasma cortisol level was highest (4.24 ± 1.571 μg dL⁻¹) after 12 h and the plasma glucose was highest (92.7 ± 9.61 mg dL⁻¹) after 2 h. These results should be useful to the aquaculture industry, where anaesthesia is necessary for a range of activities.     

The effects of temperature on the uptake and metabolism of benzo[a]pyrene in isolated gill cells of the gulf toadfish,Opsanus beta

The effects of acclimation temperature and acute temperature change on the uptake and metabolism of the procarcinogen benzo[a]pyrene (BaP) by gill cells of the gulf toadfish, Opsanus beta, were examined. BaP was rapidly accumulated by isolated gill cells and uptake rates were directly proportional to BaP concentration in the medium (1 to 100 μg/ml). Uptake rates were higher in cells isolated from fish acclimated to 18°C when compared to cells from 28°C acclimated fish at all incubation temperatures. When cells were exposed to BaP at the respective acclimation temperatures of the fish, uptake rates were similar (0.14 ± 0.01 at 18°C and 0.12 ± 0.01 μg BaP/s/10 mg cells at 28°C). This finding is discussed in view of results which showed a partial compensation of membrane fluidity in plasma membranes isolated from fish from the two acclimation temperatures. At higher incubation temperatures, cells from fish acclimated to 18°C metabolized BaP at a greater rate than those at 28°C (49.6 ± 1.92 and 43.0 ± 2.24 μg/g/8h, respectively, at 23°C). Low but detectable activities of common biotransformation enzymes (aryl hydrocarbon hydroxylase, glutathione-S-transferase) and cytochrome P-450 content were found, however, no significant differences were evident between cells from fish acclimated to different temperatures. To whom to address correspondence     

Pharmacokinetics and physio‐metabolic response of single and multiple dose of fenbendazole in Labeo rohita (Hamilton, 1822) fingerlings

An experiment was carried to determine the plasma fenbendazole (FBZ) concentration and physio‐metabolic responses in juveniles of Labeo rohita (90 ± 4 g) after oral administration of single doses at 10, 20 and 50 mg, 20 mg FBZ/kg b.wt. in multiple times on 1st, 3rd and 7th day. The blood samples were collected at 0.5, 1, 2, 4, 8, 12, 24, 30, 48, 72, 96 and 120 hr, after single‐dose administration, and regularly (upto 15 day) in multiple dose. Plasma FBZ concentration was determined up to the limit of detection (LoD) of 0.09 µg/ml by HPLC. There was no parent drug detected in plasma for administration of 10 mg FBZ/kg b.wt. The drug attained the peak concentration (C_max) 1.85 and 3.09 µg/ml in plasma at 4 hr (T_max) after administration of 20 and 50mg FBZ/kg b.wt. respectively. Plasma FBZ was detectable up to 96 and 120 hr with concentration 0.09 ± 0.007 and 0.098 ± 0.006 µg/ml, respectively, after single‐dose administration of 20 and 50mg/kg b.wt. In case of multiple‐dose administration, the maximum concentration of FBZ was 1.01 ± 0.03 µg/ml on 7th day that was less than to the single dose at 50 mg/kg b.wt. However, FBZ was detected up to 11 day after multiple doses. The study revealed that the hepatic antioxidant enzymes activities like superoxide dismutase, catalase and glutathione‐S‐transferase were significantly affected by increasing FBZ in single and multiple doses. The results of the present study could reveal that single‐ or multiple‐oral administration of FBZ at 20 mg/kg b.wt. in feed as antihelminthic drug in L. rohita could be considered as the safe dose.     

Pharmacokinetics and bioavailability of oxytetracycline in vannamei shrimp (Penaeus vannamei) and the effect of processing on the residues in muscle and shell

The present study examined the pharmacokinetics and bioavailability of oxytetracycline (OTC) in vannamei shrimp (Penaeus vannamei) after intra-sinus (10 mg/kg) and oral (10 and 50 mg/kg) administration and also investigated the net changes of OTC residues in the shrimp after the thermal, acid and alkaline processing methods. The hemolymph concentrations of OTC after intra-sinus dosing were best described by a two-compartment open model. The oral bioavailability was found to be 48.2 and 43.6% at doses of 10 and 50 mg OTC/kg, respectively. The peak hemolymph concentrations after 10 and 50 mg OTC/kg doses were 3.37 and 17.4 μg/ml; the times to peak hemolymph concentrations were 7 and 10 h. The elimination half-lives were found to be 15.0 and 11.5 h for the low and high dose, respectively. The residual OTC was rapidly eliminated from muscle with the elimination half-life value of 19.4 and 15.4 h, respectively, for the groups treated with doses of 10 and 50 mg/kg. The residual OTC levels in the muscle fell below the MRL (0.2 μg/g) at 72 and 96-h post-dosing at dose levels of 10 or 50 mg/kg, respectively. Residual OTC levels in muscle and shell were approximately 20–50% lower in the thermal treatment such as boiling, baking and frying. By the acid treatment, OTC residues were reduced to >80%, while those were reduced to around 30% by alkaline treatment.     

Bioencapsulation of erythromycin using adult brine shrimp, Artemia franciscana (Latreille)

Live adult brine shrimp, Artemia franciscana (Latreille), were enriched with erythromycin to determine if Artemia could accumulate therapeutic levels for subsequent feeding to young fish. Three trials were conducted to determine the erythromycin incorporation and survival rates of enriched Artemia when fed either liposomes containing erythromycin or various erythromycin suspensions. Erythromycin concentration in Artemia fed a liposome suspension was low (∼ 5 μg mL⁻¹) relative to Artemia fed the direct suspension (> 100 μg mL⁻¹) over the same time period. When enriched with suspensions up to 1 g erythromycin L⁻¹ sea water for 14 h, Artemia survival was not significantly affected ( P > 0.05) relative to controls. Using a suspension of 1 g L⁻¹, tissue erythromycin concentrations of 109 ± 16 μg erythromycin mL⁻¹  Artemia homogenate (mean ± SEM) were achieved after 12 h. Concentrations above 170 μg mL⁻¹ were obtained using suspensions of 2–5 g L⁻¹, but Artemia survival significantly ( P < 0.05) decreased.     

Possible Effects of Sodium Chloride Treatment on Quality of Effluents from Alabama Channel Catfish Ponds

Channel catfish ponds are treated with salt (sodium chloride) to increase chloride concentration and prevent nitrite toxicity in fish. A survey indicated that most farmers try to maintain chloride concentration of 50 to 100 mg/L in ponds by annual salt applications. Averages and standard deviations for selected water quality variables in salt-treated ponds were as follows: chloride. 87.2 ± 37.5 mg/L; total dissolved solids (TDS), 336 ± 96 mg/L; specific conductance, 512 ± 164 μmhos/cm. Maximum values were 189 mg/L for chloride, 481 mg/L for TDS, and 825 μmhos/cm for specific conductance. Good correlations between specific conductance values and both chloride and TDS concentrations suggest that specific conductance can be a rapid method for estimating concentrations of these two variables in surface water. The maximum limit for chloride concentration in Alabama streams allowed by the Alabama Department of Environmental Management is 230 mg/L. The usual recommended upper limit of TDS for protection of aquatic life in freshwater streams is 1,000 mg/L. Based on the observed relationship between TDS concentration and specific conductance in Alabama catfish ponds, 1,000 mg/L TDS corresponds to 1,733 μmhos/cm specific conductance. It is unlikely that effluents from salttreated catfish ponds would violate the in-stream chloride standard of 230 mg/L or harm aquatic life in streanis. Nevertheless, chloride concentrations in ponds should be measured before salt application as a safe guard against excessive salt application and chloride concentrations above the in-stream chloride standard.     

Residues of [14C]-malachite green in eggs and fry of rainbow trout, Oncorhynchus mykiss (Walbaum), after treatment of eggs

Abstract. Rainbow trout, Oncorhynchus mykiss (Walbaum), eggs were exposed to [methane-¹⁴C] malachite green chloride on day 0 and on every third day thereafter through day 24, with a final treatment administered to fry on day 31. Eggs or fry were sampled before each treatment, and at selected times from day 31 to day 59. Malachite green equivalence in eggs and fry was determined by sample oxidation and liquid scintillation counting. Total malachite green residues increased throughout the exposure period to 0.271 ± 0.042 μg g⁻¹ (± SD) on day 31. Residues declined to 0.055 ± 0.011 μg g⁻¹ on day 59. The depuration phase declined monoexponentially with a half-life of 13.3 days for the absolute amount (μg sample⁻¹) and a half-life of 9.7 days for the concentration of malachite green residues (μg g⁻¹). Growth dilution accounted for the 25% increase in the elimination (9.7 days) of malachite green residues. Extracts from treated eggs and fry were analysed by reversed-phase liquid chromatography. Three peaks were resolved in treated eggs: chromatic malachite green, leuco malachite green and an unknown polar metabolite. Only two peaks were resolved in the fry: leuco malachite green and the unknown polar metabolite. The most prominent residue in all samples was leuco malachite green.     

Stimulation of gonadotropin secretion and spermiation in carp by pimozide-LRH-A treatment: Effects of dose and time of day

To define the optimal combination time and LRH-A (luteinizing releasing hormone analogue) administration route for inducing spermiation in carp, two experiments were carried out. In the first experiment, carp kept in raceways in running recycled water in August (at a water temperature of 15–20°C) were injected intraperitoneally with pimozide (PIM) at 1 or 10 mg/kg body weight as well as with Des-Gly¹⁰ [D-Tryp⁶] LH-RH-ethylamide (LRH-A) at 0.1, 1 or 10 μg/kg; these products were given 6 h apart. The response was evaluated by the volume of milt and the number of spermatozoa emitted and by the gonadotropin (GTH) concentration in the plasma. At doses of 0.1, 1 and 10 μg/kg LRH-A, the total milt production was increased 2, 4 and 6-fold respectively when these treatments were combined with 1 mg PIM. In the second experiment, adult male carp, held in cages in a pond in a natural environment (at a water temperature of 11–14°C) at the beginning of May, received an intraperitoneal injection of PIM (1 mg/kg body weight) and a silicone rubber implant of LRH-A (Des-Gly¹⁰ [D-Ala⁶] - LH-RH-ethylamide) at dose of 10 μg/kg body weight simultaneously. Other groups of fish received PIM plus a blank implant. All of the treatments were given at either 6 a.m. or 8 p.m. When given at 6 a.m., the treatment induced a similar rise in plasma GTH (12–15 ng/ml) 24 h later in the groups treated with PIM and LRH-A implant alone or in combination. When the same treatment was given at 8 p.m., no response was observed 24 or 48 h later. Spermiation was not stimulated in any of the groups.