A pharmacokinetic study of flumequine in sea bass, Dicentrarchus labrax (L.), after a single intravascular injection

The pharmacokinetic properties of flumequine following a single intravascular injection (10 mg kg^–1 fish) were studied in sea bass, Dicentrarchus labrax (L.), 120 g held at 18 °C. The absorption half life ( t _1/2α) and the elimination half life ( t _1/2β) of the drug were calculated to be 1.05 and 10.71 h, respectively. Tissue penetration of flumequine seemed to be moderate because both the apparent volume of distribution of the drug at steady-state ( V _d(ss)) and the extensive apparent volume of the central compartment ( V _c) were found to be small (1.51 and 0.626 L kg^–1). The mean residence time ( MRT ) was short (09.73 h) and the total clearance (CL_T) of the drug was rapid (0.156 L kg^–1 h^–1).     

Characterization of grouper nervous necrosis virus (GNNV)

Grouper nervous necrosis virus (GNNV) was isolated from moribund grouper larvae, Epinephelus sp., using a fish cell line GF-1. The present study describes the biochemical and biophysical properties of GNNV and the expression of GNNV in diseased grouper larvae. Viral protein was detectable in most of the GNNV-infected GF-1 cells by the fluorescent antibody technique (FAT) after 12 h post-infection (p.i.), although no cytopathic effect (CPE) appeared at that time. Clear CPE developed on the third day, and complete disintegration of the monolayer occurred over the subsequent two days. The infectivity of GNNV can be blocked following treatment at 60 °C for 1 h. GNNV was sensitive to pH 3 and pH 10–12 with a 4 log₁₀ drop in infectivity. Purified GNNV was analysed by SDS–PAGE, and then stained with periodic acid silver. The positive staining indicated that its two capsid proteins were glycoproteins. Genomic RNAs of GNNV were extracted from purified virions and analysed. The molecular weights of genomic RNAs were 1.02 × 10⁶ and 0.50 × 10⁶ Da. The T2 region of the coat protein gene of GNNV was amplified by polymerase chain reaction (PCR), and the multiple alignment of the T2 sequence of two GNNV isolates with four genotypes of fish nodaviruses revealed that these two isolates (GNNV9410 and GNNV9508) belong to the red-spotted grouper nervous necrosis virus (RGNNV) genotype. The tissue distribution of GNNV in naturally infected grouper larvae was investigated by in situ hybridization using a dig-labelled probe, which showed that GNNV was not only detected in the brain and retina, but also in the gill, skeletal muscle, liver, pyloric gland, intestine and blood cells in the heart.     

Effects of a peroxide-based commercial product on bacterial load of larval rearing water and on larval survival of two species of Sparidae under intensive culture: preliminary study

Larvae of two Mediterranean Sparidae species, Sparus aurata and Dentex dentex , were used to test the efficacy of a peroxide-based product (Ox-Aquaculture^©) on the reduction in bacterial load in larval rearing water and its effects on larval survival. Eleven-day-old S. aurata larvae and 15-day-old D. dentex larvae were exposed to different concentrations of Ox-Aquaculture^© (50, 100 and 200 mg L⁻¹, and 20 and 50 mg L⁻¹ respectively) for 1 h. Results indicated that 50 and 20 mg L⁻¹ were the most effective concentrations for the reduction in bacterial load (at least one order of magnitude) after 1 h treatment, without affecting larval survival and/or vitality in 11 dph S. aurata and 15 dph D. dentex larvae respectively. Ox-Aquaculture^© concentrations of 200 and 50 mg L⁻¹ during 1 h affected negatively final survival rate of the larvae of S. aurata and D. dentex respectively.     

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.     

Use of neuroactive catecholamines to chemically induce metamorphosis of hatchery-reared flat oyster, Ostrea angasi, larvae

Low numbers and unreliable wild catch of the native flat oyster, Ostrea angasi , spat has resulted in the NSW flat oyster industry being reliant on hatchery-produced spat. The need to produce culchless spat in the hatchery stimulated investigation of several catecholamines to induce metamorphosis in O. angasi larvae. Larvae were treated with one of four neuroactive catecholamines (epinephrine, epinephrine bitartrate, l -Dopa and GABA) at one of four concentrations (10⁻³, 10⁻⁴, 10⁻⁵ or 10⁻⁶  m ) for one of three treatment durations (0.5, 1–2 h) to determine morphogenic action for culchless spat production. Epinephrine bitartrate at 10⁻³ and 10⁻⁴  m and epinephrine at 10⁻³, 10⁻⁴ and 10⁻⁵  m , for a treatment duration of 1–2 h, produced significantly greater numbers of spat and culchless spat, compared with any other treatment combination. The other catecholamines tested did not induce a significant increase in the total number of spat or culchless spat, over untreated controls. Separate trials found that long-term treatment (24 h) with epinephrine bitartrate and epinephrine at morphogenic concentrations inhibited metamorphosis. Consecutive daily use of epinephrine bitartrate increased the numbers of spat and culchless spat produced, but did not affect larval or short-term post-larval survival. Treatment with 10⁻³  m epinephrine bitartrate or 10⁻⁴  m epinephrine for 1 h is recommended for routine commercial production of culchless flat oyster spat.     

Long-term effect of photoperiod manipulation on growth, maturation and flesh quality in Atlantic halibut

The aim of this study was to investigate the effect of continuous light at different stages during the production cycle of Atlantic halibut Hippoglossus hippoglossus L. on growth, age at first maturity, endocrine parameters and flesh quality. A group of juvenile halibut [mean (SD), initial weight 191.3 g (44.7)] was reared in indoor tanks under ambient temperature conditions for 38 months until harvesting (mean final weight, 4.6 kg). The entire photoperiod experiment was divided into four phases, where the fish in each phase were exposed to either natural photoperiod (62°33'N) or continuous light (L). Thus, the following five photoperiod combinations were tested: (a) Control group (NNNN), (b) Group 2A (NLNN), (c) Group 2B (NNLN), (d) Group 2C (NNNL) and (e) Production group (LNNN). Exposure to continuous light stimulated growth, and the final mean weights of Groups 2A and 2B were 23% and 11% higher than those of the Control group (NNNN). The final plasma 11-ketotestosteron levels were lower in Groups 2A (2.94 ng mL⁻¹) and 2B (2.46 ng mL⁻¹) compared with the Control (5.29 ng mL⁻¹), Group 2C (5.09 ng mL⁻¹) and the Production group (4.78 ng mL⁻¹) during spring 2007 (age 4 years), indicating higher age at first maturity in Groups 2A and 2B. Photoperiod regime had only a minor, and transient, effect on flesh-quality traits of the fish, whereas a significant seasonal effect was seen with a tendency towards increased gaping, lower pH, lower hardness and lower shear force in July compared with December and March.     

Continuous exposure to infectious pancreatic necrosis virus during early life stages of rainbow trout, Oncorhynchus mykiss (Walbaum)

Rainbow trout, Oncorhynchus mykiss (Walbaum), were exposed continuously to infectious pancreatic necrosis virus (IPNV) at 0, 10¹, 10³ or 10⁵ plaque forming units (pfu) L⁻¹ of water to estimate the effects of chronic IPNV exposure on early life stages. Fish density averaged 35 fish L⁻¹ (low density) or 140 fish L⁻¹ (high density), and the tank flow rate was 250 mL⁻¹ min. Virus exposure began at 6 days before hatch and continued until fish were 44 days old. Cumulative per cent mortality, analysis of survival and hazard functions, and discrete-time event analysis were used to explore the patterns of survival and mortality. In eggs and fish exposed to IPNV, mortality significantly greater than in the 0 pfu L⁻¹ exposure did not occur until IPNV concentration was 10⁵ pfu L⁻¹ at low fish density and 10³ pfu IPNV L⁻¹ at high fish density. These results suggest that in the natural aquatic environment, where rainbow trout densities are likely to be considerably lower than in this study, mortality resulting from infection with IPNV will very likely not occur when ambient concentrations of virus are ≤10³ pfu IPNV L⁻¹. In aquaculture rearing units, trout density is likely to be as high or higher than the densities used in this study. Therefore, continuous inputs of virus at concentrations greater than 10¹ pfu L⁻¹ may result in IPN epidemics in aquaculture facilities.     

Perception of light intensity by Haliotis discus discus based on locomotor activity patterns

ABSTRACT:   An apparatus to measure the locomotor activity of aquatic benthic organisms at variable low light levels was developed and the diurnal behavioral pattern of the abalone Haliotis discus discus was measured at various low light intensities. During the experiment, abalone were exposed to 12 h light–dark cycles of complete darkness, 0 µmol/m²/s throughout the 12 h dark cycle and, during periods I (days 1–8) and III (days 19–26), the 12 h light cycles were set at 10 µmol/m²/s. During period II (days 10–17), abalone were exposed to a light level during the 12 h light cycles of 1 × 10⁻⁵, 1 × 10⁻⁶, 1 × 10⁻⁷ or 1 × 10⁻⁸ µmol/m²/s and the changes in locomotor activity assessed. At daytime levels of 1 × 10⁻⁵ µmol/m²/s, typical behavioral patterns were observed of high locomotory activity during the night-time cycle. However, at lower light intensities, the distinction between day and night activity patterns became less clear and, at intensities lower than 1 × 10⁻⁷ µmol/m²/s, the difference between activity during the light and dark cycles became negligible. Based on this, we conclude that the threshold of light level perception in relation to locomotor activity is approximately 1 × 10⁻⁷ µmol/m²/s. The significance of these results in relation to the entrainment of behavior in abalone is discussed.     

Effects of dietary phosphorus level on non-faecal phosphorus excretion from yellowtail (Seriola quinqueradiata Temminck & Schlegel) fed purified and practical diets

Non-faecal phosphorus (P) was determined for large yellowtail to estimate a minimum available P requirement (Experiment  1) and to justify inorganic P supplementation in a fish meal-based diet (Experiment 2). In Experiment 1, purified diets with incremental P concentrations were fed to yellowtail (mean weight 917 g) at a feeding rate of 1.5% of body weight. The peaks of non-faecal P excretion appeared 5–6 h after feeding in fish fed more than 4.5 g available P kg⁻¹ dry diet. Broken-line analysis indicated that the minimum available P requirement was 4.4 g kg⁻¹ dry diet. In Experiment 2, a purified diet (PR) containing 6.5 g available P kg⁻¹ and a fish meal-based diet with (F1) and without (F0) additional phosphorus were fed to yellowtail (mean weight 1.1 kg) at 1.5% (PR) and 2% (F0 and F1) feeding rates respectively. There was no significant difference in P excretion between fish fed the F0 (5.5 g soluble P kg⁻¹ dry diet) and the PR diet. However, significantly higher (34.5%) amounts of non-faecal P excretions (7.4 g soluble P kg⁻¹ dry diet) were found in fish fed F1 compared with the F0 diet. This suggested that there was an excess of dietary P in the F1 diet and that supplementation is not needed in fish meal-based diets for large yellowtail.     

Propagation of yellow grouper nervous necrosis virus (YGNNV) in a new nodavirus-susceptible cell line from yellow grouper, Epinephelus awoara (Temminck & Schlegel), brain tissue

A nodavirus was isolated from diseased yellow grouper, Epinephelus awoara , larvae cultured in southern Taiwan. The histopathology and RT–PCR results confirmed that it was a fish nodavirus; its coat protein gene sequence was similar to that of red spotted grouper nervous necrosis virus (RGNNV) and it is named yellow grouper nervous necrosis virus (YGNNV). A new nodavirus-susceptible cell line, grouper brain (GB) was established and characterized from the brain tissue of yellow grouper. The GB cells multiplied well in Leibovitz's L-15 medium supplemented with 10% foetal bovine serum at temperatures between 24 and 32 °C, and have been subcultured more than 80 times, becoming a continuous cell line. The GB cell line consists of fibroblast-like cells and some epithelioid cells. The cell line yielded titres of YGNNV up to 10^8.5 TCID₅₀ mL^–1. The GB cells effectively replicated the virus at 28 °C, which could be purified to homogeneity by caesium chloride gradient centrifugation. Electron microscopy studies showed that purified virus particles were 25–30 nm in diameter. The cytoplasm of infected cells was filled with aggregates of virus particles. These results indicate that the GB cell line is a significant tool for the study of fish nodaviruses.     

The assessment of the chemical composition of fishmeal by near infrared reflectance spectroscopy

The use of near infrared reflectance spectroscopy (NIRS) was investigated as an alternative method for predicting moisture (M), oil, crude protein (CP), ash, salt as NaCl, total volatile nitrogen (TVN) and buffer capacity in fishmeal. The NIRS calibration models were developed using the modified partial least squares (MPLS) regression technique. One thousand and ten ( n =1010) fishmeal samples were used to predict chemical composition for quality control in the fishmeal industry. Equations were selected based on the lowest cross validation errors (SECV). The coefficient of determination in calibration ( R ²) and SECV were 0.93 and 3.9 g kg^–1 dry matter (DM); 0.85 and 5.7 g kg^–1 DM; 0.92 and 3.7 g kg^–1 DM; 0.91 and 4.7 g kg^–1 DM; 0.88 and 6.7 g kg^–1 DM; 0.94 and 1.8 g kg^–1 DM; for M, CP, oil, ash, TVN and NaCl, respectively. It was concluded that NIRS can be used as a method to monitor the quality of fishmeal under industrial conditions.     

Fishery resources and economic gain in three mangrove areas on the south-east coast of India

Abstract The fishery resources and fishing income in three different mangrove areas were evaluated. The mangrove-rich area provided high catches of shellfish and finfish and yielded high fishery income, compared with the mangrove-poor areas. Luxuriant mangrove areas supported a catch of 11 kg shellfish ha⁻¹ day⁻¹ and 4.5 kg finfish ha⁻¹ day⁻¹, corresponding with an economic gain of US$ 14 (Rs 603.7) day⁻¹ for shellfish and US$ 3 (Rs 130.5) day⁻¹ for finfish. The value of maintaining mangroves to ensure better fishery resources and to support coastal economies was identified. Management practices for conserving and enhancing mangroves and associated fishery resources are suggested.     

Quantitative real time PCR for the measurement of white spot syndrome virus in shrimp

Quantitative real time PCR, recently developed in molecular biology, is applied in this paper to quantify the white spot syndrome virus (WSSV) in infected shrimp tissue. The WSSV content in moribund shrimp of all species tested ( Penaeus stylirostris, P. monodon, P. vannamei ) ranged from 2.0 × 10⁴ to 9.0 × 10¹⁰ WSSV copies μg^–1 of total DNA ( n =26). In whole moribund post-larvae, 4.3 × 10⁹ WSSV copies μg^–1 of DNA were detected which is equivalent to 5.7 × 10¹⁰ WSSV copies g^–1 of post-larvae. The comparison of WSSV content between different tissues showed that muscle and hepatopancreas tissues contained 10 times less virus than gills, pleopods and haemolymph. With inocula of known virus content, bioassays by immersion challenge showed that a minimum of five logs of WSSV copies was necessary to establish disease in the challenged shrimp. In contrast, five logs of WSSV copies injected into shrimp muscle produced a LT-50 of 52 h. This real time polymerase chain reaction (PCR) technique is sensitive (four copies), specific (negative with DNA from shrimp baculoviruses and parvoviruses), dynamic (seven logs) and easy to perform (96 tests in <4 h).     

Immune Responses and Resistance against Vibrio parahaemolyticus Induced by Probiotic Bacterium Arthrobacter XE-7 in Pacific White Shrimp, Litopenaeus vannamei

A 63-d feeding experiment was conducted to determine the effects of dietary supplementation of probiotic bacterium Arthrobacter XE-7 on immune responses and resistance against Vibrio parahaemolyticus in the Pacific white shrimp, Litopenaeus vannamei . The probiotic bacteria were administered orally at four different doses of 0, 10⁶, 10⁸, and 10¹⁰ colony-forming unit (CFU)/g feed for shrimp. On Day 50, the shrimp were challenged with Vibrio parahaemolyticus by bath. On Days 7, 21, 49, and 63, six shrimp per tank were sampled to take intestine and hemolymph. With increasing dietary supplementation of probiotic bacteria, shrimp mortality decreased from 63.16% (the control) to 55.9% (10⁶ CFU/g feed), 51.75% (10⁸ CFU/g feed), and 51.78% (10¹⁰ CFU/g feed), respectively. Vibrio counts in intestine of shrimp fed probiotic bacterium Arthrobacter XE-7 was generally lower than that in the control shrimp ( P  < 0.05). The probiotic bacteria generally increased the immune parameters in shrimp, that is, total hemocyte counts, percentage phagocytosis, respiratory burst activity, and serum phenoloxidase activity. The results showed that probiotic bacterium Arthrobacter XE-7 can be used as probiotic in shrimp feed.     

Use of full-fat winged bean Psophocarpus tetragonolobus seed meal as a protein feedstuff in fish-meal free diets for African catfish Clarias gariepinus

Mature winged bean Psophocarpus tetragonolobus seeds were quick-cooked and the full-fat meal derived was used to completely replace menhaden fish meal as a dietary protein source for the African catfish Clarias gariepinus . Five dry practical diets (400 g crude protein kg⁻¹ and 17.5 kJ gross energy g⁻¹ dry diet) containing menhaden fish meal (diet 1) or winged bean meal with or without graded levels of supplemental L -methionine (diets 2, 3, 4 and 5; 0, 5, 10 and 15 g kg⁻¹, respectively) were fed to catfish fingerlings (5.8  +  1.2 g) for 70 days. Weight gain, growth rate, feed conversion and protein utilization by catfish fed a winged bean meal diet without L -methionine supplementation (diet 2) was inferior ( P  > 0.05) to that in catfish fed the other diets, where performance differed nonsignificantly. Carcass protein of catfish was lower ( P  < 0.05) while liver protein was higher ( P  < 0.05) in catfish fed the winged bean meal diet without methionine supplementation. Results suggest that winged bean meal cannot replace fish meal as a protein source in catfish diets except with a minimum supplementation with 5 g L -methionine kg⁻¹ diet.     

Effects of dietary phosphorus and lipid levels on utilization and excretion of phosphorus and nitrogen by rainbow trout (Oncorhynchus mykiss). 2. Production-scale study

In a 8-week production-scale experiment at a commercial trout farm, the effects of dietary lipid level and phosphorus level on phosphorus (P) and nitrogen (N) utilization of rainbow trout (initial mean weight 99 g) were assessed. A low-phosphorus, high-lipid experimental diet (457 g protein, 315 g lipid, 9.1 g P  kg^–1 dry diet) was compared with a commonly used commercial diet (484 g protein, 173 g lipid, 13.6 g P  kg^–1 dry diet). P and N budgets were constructed using data from the production-scale experiment and digestibility data for the two diets. In addition, orthophosphate and ammonia-N waste were measured in effluent over one 24-h period. Relative to the commercial diet, the experimental diet resulted in significantly increased feed efficiency ratio, N retention and P retention, and substantially reduced dissolved, solid and total P waste (g kg^–1 dry feed). Although N retention resulting from the experimental diet was higher, this was attributable to higher N (protein) digestibility of the experimental diet. Solid N waste (g kg^–1 dry feed) resulting from the experimental diet was substantially lower, but dissolved N waste (g kg^–1 dry feed) was not significantly different relative to the commercial diet. Mean effluent orthophosphate production (mg day^–1 kg^–1 fish) of fish fed the experimental diet was substantially lower than that of fish fed the commercial diet ( P  < 0.05), but effluent ammonia-N production (mg day^–1 kg^–1 fish) was not significantly affected by dietary treatment.     

Evaluation of indigenous marine periphytic Amphora, Navicula and Cymbella grown on substrate as feed supplement in Penaeus monodon postlarval hatchery system

Three isolated marine diatoms ( Amphora , Navicula and Cymbella ) grown on substrate were evaluated as feed supplement for Penaeus monodon postlarvae (PL) in hatchery system for a period of 19 days without changing water. Specific growth rate (day⁻¹) (0.27 ± 0.0) and survival (%) (56.3 ± 1.8) of PLs were significantly higher ( P  < 0.05) in treatment tanks when compared with the control (0.20 ± 0.0; 36.0 ± 1.5, respectively). Shrimp PLs reared in substrate-based tanks had significantly higher ( P  < 0.05) levels of protein, lipid (521.0 ± 7.0; 304.0 ± 2 g kg⁻¹ dry weight, respectively), ecosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) (189.0 ± 2.0; 176.0 ± 2 g kg⁻¹ of total fatty acid, respectively) than the control (435.0 ± 22.0; 258.0 ± 22 g kg⁻¹ dry weight; 172.0 ± 5.0; 152 ± 2 g kg⁻¹ total fatty acid, respectively). The periphytic diatoms contained protein and lipid (430–490; 230–260 g kg⁻¹ dry weight, respectively), EPA (30–150 g kg⁻¹ of total fatty acids), DHA (20–30 g kg⁻¹ of total fatty acids) and nine essential amino acids. The results showed that isolated marine periphytic diatoms grown on substrate could be used as feed supplement in enhancing the growth and survival of P. monodon postlarvae.     

Variations in spawning ground area and egg density of the Japanese sardine in Pacific coastal and oceanic waters

The spawning ground of the Japanese sardine, Sardinops melanostictus (Schlegel), was distributed over the oceanic waters as well as the coastal waters along the Pacific coasts of western and eastern Japan during 1978–1992. The area of the spawning ground in the coastal waters on the continental shelf has ranged from 95 000 km² in 1992 to 143 000 km² in 1988, constituting 44–77% of the total area of the spawning ground. The area of the coastal spawning ground was relatively constant in spite of the large fluctuations in egg abundance, i.e. size of the spawning population, from 88 trillion (1987) to 668 trillion (1989) in the waters. Spawning adults seemed to extend over the coastal waters irrespective of the size of the spawning population. In contrast to the coastal waters, the spawning area in the oceanic waters offshore of the continental shelf increased from 31 000 km² in 1978 to 183 000 km² in 1988 and then shrank to 83 000 km² in 1992, as a function of the spawning population size. The egg distribution density in the coastal waters stayed less than 6000 m⁻² mo⁻¹, but it reached as high as 27 400 m⁻² mo⁻¹ in the expanded spawning ground in the oceanic waters. The oceanic waters seemed to function as a reserve spawning ground for the sardine in years of extremely high spawning population.     

The effect of dietary phosphatidylcholine and its constituent fatty acids on microdiet ingestion and fatty acid absorption rate in gilthead sea bream, Sparus auratus, larvae

This study tested the effect of the level of dietary phosphatidylcholine (PC) and its constituent medium-chain fatty acids on microdiet ingestion (μg diet larva⁻¹ h⁻¹) and the absorption rate of the free fatty acid [¹⁴C]16:0 (pmole larva⁻¹ h⁻¹) in 15, 20, 21, 25, 26, 30 and 31-day-old gilthead sea bream, Sparus auratus L., larvae. Fish were fed four microdiets (A, B, C and D): microdiet A contained no phospholipid (PL), while microdiet B included 10 g kg⁻¹ Artemia nauplii PL (3.7 g kg⁻¹ PC). Microdiets C and D contained 10 g kg⁻¹ purified saturated PC dimyristoyl (C_14:0) and polyunsaturated PC dilinoleoyl (C_{18:2[cis]−9,12}), respectively.
Larvae from one or both of the PC microdiets demonstrated significantly higher ( P < 0.05) ingestion rates (μg diet larva⁻¹ h⁻¹) than the non-PL microdiet control in 15, 21, 22, 25 and 26-day-old larvae and the Artemia PL microdiet in 15, 22 and 26-day-old larvae. However, microdiet ingestion and fatty acid absorption rate appeared to be independent of the associated medium carbon chain saturated or polyunsaturated fatty acid moiety of the PC diets. Apparent absorption, as measured by the retention of radio-labelled [¹⁴C]16:0 following 8 h of non-labelled microdiet feeding, was possibly related to feeding.