RESUMEN
The effect of ß-glucans 1,3/1,6 from Saccharomyces cerevisiae yeast at different inclusion percentages (0.0, 0.2, 0.4, 0.6, and 0.8%) in the diet for tropical gar (Atractosteus tropicus) larvae was evaluated on growth, digestive enzyme activity and, relative expression of the immune system genes. The bioassay started on the third day after hatching (DAH) and lasted 21 days, using a total of 1500 larvae of 0.055 ± 0.008 g and, a total length of 2.46 ± 0.26 cm. Larviculture was carried out in a recirculation system with 15 tanks of 70 L using a density of 100 organisms per experimental unit. No significant differences in larval growth were observed by the inclusion of ß-glucans (p > 0.05). Digestive enzymes showed changes in lipase and trypsin activities, presenting higher values in fish fed 0.6% and 0.8% ß-glucans diets compared to the other treatments (p < 0.05). Leucine-aminopeptidase, chymotrypsin, acid phosphatase, and alkaline phosphatase activity showed higher activities in larvae fed with a 0.4% ß-glucan diet compared to the control group. The relative expression of intestinal membrane integrity (mucin 2) muc-2, (occludins) occ, (nucleotide-binding oligomerization domain) nod-2, and immune system lys (lysosome) genes showed over-expression in larvae fed the 0.4% ß-glucan diet to the rest of the treatments (p < 0.05). The inclusion of ß-glucans at 0.4-0.6% in diets for A. tropicus larvae could improve larviculture, as effects on the increase in the activity of several digestive enzymes and the expression of genes of the immune system.
Asunto(s)
Peces , beta-Glucanos , Animales , Larva , Peces/metabolismo , Intestinos , Dieta/veterinaria , Expresión Génica , beta-Glucanos/metabolismoRESUMEN
Determination of the main nutritional requirements at different developmental stages is a prerequisite in the formulation of feeds for newly cultured fish species. In the present study, the lipid and protein requirements of larvae of a native Mexican fish, the three-spot cichlid Cichlasoma trimaculatum, were assessed using a two-factor experimental design that considered four protein (35, 40, 45, and 50%) and two lipid levels (16 and 22%) on growth, survival, and digestive enzyme activities. The best growth and feed efficiency results were obtained when larvae were fed diets including 45% protein and 22% lipids. Comprehensive evaluation of the profile of digestive enzymes using multivariate analysis also demonstrated significant differences in nutritional condition generated by varying inclusion of nutrients. Thus, an increase in protein led to an increase in alkaline protease activity and a reduction in leucine aminopeptidase activity, and the reduction of dietary lipid content led to a significant increase in lipase and trypsin enzymatic activities. Based on our results, C. trimaculatum larvae have a high capacity to hydrolyze both nutrients (protein and lipids) for the high digestive enzyme activities and increase their growth, particularly with a diet containing 45% protein and 22% lipids.
Asunto(s)
Cíclidos/fisiología , Dieta , Grasas de la Dieta , Proteínas en la Dieta , Animales , Cíclidos/metabolismo , Lipasa , Lípidos , Proteínas , TripsinaRESUMEN
Functional characteristics of digestive proteases and in vitro digestibility of several protein sources were studied in hatchlings of Central American river turtles, Dermatemys mawii. Acidic and alkaline proteases from the stomach and intestines were used, and optimums for acidic proteases were registered at 55°C and pH 2, while alkaline proteases were found at 55°C and pH 9. Ten protein ingredients, both vegetable and animal sources, were explored by the pH-STAT method, only for alkaline proteases. The degree of hydrolysis was at its highest for squid meal and lowest for blood meal, while the total free amino acids level was at its highest for squid meal, and lowest for wheat gluten meal. Our results indicate that D. mawii has a broad capacity to digest both animal and vegetable sources, and suggests some ingredients more suitable to design artificial diets for this species.
Asunto(s)
Dieta/veterinaria , Digestión/fisiología , Péptido Hidrolasas/metabolismo , Tortugas/metabolismo , Aminoácidos , Alimentación Animal , Fenómenos Fisiológicos Nutricionales de los Animales , Animales , Estados UnidosRESUMEN
Aquaculture in Mexico has been developed by the cultivation of commercial species. In Tabasco, the cultivation of native species is mainly limited by the lack of nutrition studies to support its crop profitability. Among these species is the tropical gar (Atractosteus tropicus), which has great potential for cultivation. However, the nutritional value of carbohydrates in diets for this species which contribute to improved growth and survival, have not been evalulated,. Thus, in the present investigation, isoprotein and isolipid diets have been designed based on the substitution of cellulose by corn starch (D1: 0% starch-15% cellulose, D2: 7.5% starch-7.5% cellulose and D3: 15% starch-0% cellulose) and compared with a commercial trout diet (45% protein and 16% lipids). A total of 1800 larvae (0.008 ± 0.002 g and 10.5 ± LT 0.126 mm) were used, distributed in a recirculation system in order to evaluate growth and survival for 30 days. The results show higher growth and survival of 97% of larvae fed the D3 diet, while cannibalism in the species was mitigated. Major digestive enzyme activities occurred (acid protease, alkaline protease, trypsin, chymotrypsin, leucine aminopeptidase, carboxypeptidase A, lipase, α-glucosidase and amylase) for larvae fed D3. It is concluded that the contribution of corn starch (15%) replacing cellulose in the diet improves growth and survival of this species.
Asunto(s)
Acuicultura , Dieta/métodos , Peces/fisiología , Almidón , Amilasas/metabolismo , Animales , Leucil Aminopeptidasa/metabolismo , México , Péptido Hidrolasas/metabolismo , Tripsina/metabolismoRESUMEN
Changes in digestive enzyme activity and histology were studied in Atractosteus tropicus embryos, larvae and juvenile periods. Alkaline protease, chymotrypsin, carboxypeptidase A, lipase and α-amylase were detected in all periods and gradually increased until reaching the maximum peak in juveniles; meanwhile, acid protease was first detected at 5 days after hatching (dah) when first feeding started and trypsin and leucine aminopeptidase activities were detected from 19 dah, their values being increased gradually until reaching a maximum value at 31 dah. Acid and alkaline phosphatase activities increased from yolk-sac absorption (3 dah) until day 31 after hatching. Zymogram for acid protease showed two bands in active forms (0.4 and 0.5 Rfs) from day 5 after hatching and a third protease form (0.3 Rf) that appears at 31 dah. Two active forms (26.3 and 24.9 kDa) were detected using SDS-PAGE alkaline proteases zymogram at 5 dah, and an additional active form (44.1 kDa) was detected at 7 dah. Regarding the histological development of the digestive system, the exocrine pancreas containing zymogen granules was already visible at 3 dah, whereas at 5 dah first gastric glands were already detected in the stomach. Between 7 and 9 dah, the digestive tract of A. tropicus resembled that of a juvenile specimen with a well-developed and short oesophagus, stomach divided into a glandular and non-glandular (pyloric) stomach, folded intestine with pyloric caeca and a well-developed spiral valve (posterior intestine). Considering this, larvae of A. tropicus are capable of digesting several foods from yolk absorption (3 dah), maximizing its activities at 15 dah, age at which the organisms maximize its capability to absorb nutrients from diets provided.
Asunto(s)
Peces/embriología , Tracto Gastrointestinal/embriología , Tracto Gastrointestinal/metabolismo , Animales , Regulación del Desarrollo de la Expresión Génica/fisiología , Regulación Enzimológica de la Expresión Génica/fisiología , Larva/metabolismoRESUMEN
A study was performed in order to understand the development of digestive enzymes during initial ontogeny of Cichlasoma trimaculatum, for which the activity of acidic and alkaline proteases, lipases, amylases and phosphatases was determined by means of biochemical and electrophoretic analysis. Our results showed that the activity of alkaline proteases, trypsin and chymotrypsin is present from day 6 after hatching (dah) during exogenous feeding with Artemia nauplii. The activities of carboxypeptidase A and leucine aminopeptidase are present from the first days, increasing at 6 dah and reaching their maximum activity at 9 dah while acid protease activity started at 9 dah. Furthermore, the lipase activity is detected on 6 dah and keeps increasing and decreasing on 17 dah. Amylase activity is detected on 3 dah, presenting fluctuations until 45 dah, where it reaches its maximum activity. Acid and alkaline phosphatases are detected from 3 dah and reach a maximum activity between 13 and 19 dah. The SDS-PAGE electrophoresis revealed six types of bands in the alkaline proteases, with molecular weight between 113.4 and 20.4 kDa. First three bands appear on 6 dah, but it is until 11 dah when all isoforms appear. Based on these results, it is considered that this species completes its digestive enzymatic machinery from day 9 after hatching, therefore is recommended to perform the transition from live feed to inert feed at 15 dah.
Asunto(s)
Acuicultura/métodos , Cíclidos/crecimiento & desarrollo , Sistema Digestivo/enzimología , Regulación del Desarrollo de la Expresión Génica/fisiología , Regulación Enzimológica de la Expresión Génica/fisiología , Factores de Edad , Amilasas/metabolismo , Animales , Carboxipeptidasas A/metabolismo , Quimotripsina/metabolismo , Cíclidos/metabolismo , Sistema Digestivo/crecimiento & desarrollo , Electroforesis en Gel de Poliacrilamida , Leucil Aminopeptidasa/metabolismo , Lipasa/metabolismo , Péptido Hidrolasas/metabolismo , Tripsina/metabolismoRESUMEN
Tropical gar (Atractosteus tropicus) is an economically and socially important freshwater species from Southeastern Mexico, with a high aquaculture potential. With this in mind, the purpose of this study was to characterize the digestive proteases of tropical gar juveniles through biochemical and electrophoretic analyses. Twenty specimens with an average weight of 73.6 ± 12.7 g were used to obtain stomach and intestinal tissue from which multienzymatic extracts were prepared. The general activities of the acid and alkaline proteases were evaluated, as well as the specific activities of trypsin, chymotrypsin, leucine aminopeptidase and carboxypeptidase A. The effect of the pH and temperature on the proteases was also analyzed, together with the composition of the multienzymatic extracts using protease inhibitors and electrophoretic tests. Results showed that A. tropicus have a functional stomach in which protein hydrolysis starts with pepsin and which contains endo- and exopeptidases (trypsin, chymotrypsin, leucine aminopeptidase and carboxypeptidase A) and proteases that are resistant to high temperatures (45 and 55 °C for alkaline and acid proteases, respectively) and pH values. Using zymogram technique, we found two acid protease isoforms (0.35 and 0.71 rf) and five alkaline protease isoforms (83.7, 43.7, 27.5, 24.0 and 19.4 kDa), which decrease or disappear with the different inhibitors. Thus, this species is considered to be a carnivore capable of adapting to its environment by consuming different types of proteins from preys and also could adapt rapidly to consume a compound diet with different animal protein sources.
Asunto(s)
Digestión/fisiología , Peces/metabolismo , Tracto Gastrointestinal/enzimología , Péptido Hidrolasas/metabolismo , Animales , Acuicultura/métodos , Carboxipeptidasas A/metabolismo , Quimotripsina/metabolismo , Electroforesis/veterinaria , Peces/fisiología , Concentración de Iones de Hidrógeno , Isoenzimas/metabolismo , Leucil Aminopeptidasa/metabolismo , México , Estadísticas no Paramétricas , Temperatura , Tripsina/metabolismoRESUMEN
The characterisation of digestive proteases in native freshwater fish such as the Mayan cichlid Cichlasoma urophthalmus provides scientific elements that may be used to design balanced feed that matches with the digestive capacity of the fish. The purpose of this study was to characterise the digestive proteases, including the effect of the pH and the temperature on enzyme activity and stability, as well as the effect of inhibitors using multienzymatic extracts of the stomach and intestine of C. urophthalmus juveniles. Results showed that the optimum activities of the acid and alkaline proteases occurred at pH values of 3 and 9, respectively, whereas their optimum temperatures were 55 and 65 °C, respectively. The acid proteases were most stable at pH values of 23 and at temperatures of 3545 °C, whereas the alkaline proteases were most stable at pH values of 69 and at 2555 °C. The inhibition assays recorded a residual activity of 4% with pepstatin A for the acid proteases. The inhibition of the alkaline proteases was greater than 80% with TPCK, TLCK, EDTA and ovalbumin, and of 60 and 43.8% with PMSF and SBT1, respectively. The results obtained in this study make it possible to state that C. urophthalmus has a sufficiently complete digestive enzyme machinery to degrade food items characteristic of an omnivorous fish species, although specimens showed a tendency to carnivory.
Asunto(s)
Cíclidos/metabolismo , Digestión , Intestinos/enzimología , Péptido Hidrolasas/metabolismo , Estómago/enzimología , Animales , Proteínas de Peces/metabolismo , Inhibidores de ProteasasAsunto(s)
Cíclidos/fisiología , Digestión , Tracto Gastrointestinal/enzimología , Tracto Gastrointestinal/crecimiento & desarrollo , Fenómenos Fisiológicos Nutricionales de los Animales , Animales , Cíclidos/crecimiento & desarrollo , Larva/enzimología , Larva/crecimiento & desarrollo , Larva/fisiologíaRESUMEN
Common snook (Centropomus undecimalis) is one of the most important marine species under commercial exploitation in the Gulf of Mexico; for this reason, interest in developing its culture is a priority. However, larviculture remains as the main bottleneck for massive production. In this sense, our objective was to determine the changes of digestive enzymes activities using biochemical and electrophoretic techniques during 36 days of Common snook larviculture fed with live preys (microalgae, rotifers, and Artemia). During larviculture, all digestive enzymatic activities were detected with low values since yolk absorption, 2 days after hatching (dah) onwards. However, the maximum values for alkaline protease (6,500 U mg protein(-1)), trypsin (0.053 mU × 10(-3) mg protein(-1)), and Leucine aminopeptidase (1.4 × 10(-3) mU mg protein(-1)) were detected at 12 dah; for chymotrypsin at 25 dah (3.8 × 10(-3) mU mg protein(-1)), for carboxypeptidase A (280 mU mg protein(-1)) and lipase at 36 dah (480 U mg protein(-1)), for α-amylase at 7 dah (1.5 U mg protein(-1)), for acid phosphatases at 34 dah (5.5 U mg protein(-1)), and finally for alkaline phosphatase at 25 dah (70 U mg protein(-1)). The alkaline protease zymogram showed two active bands, the first (26.3 kDa) at 25 dah onwards, and the second (51.6 kDa) at 36 dah. The acid protease zymogram showed two bands (RF = 0.32 and 0.51, respectively) at 34 dah. The digestive enzymatic ontogeny of C. undecimalis is very similar to other strictly marine carnivorous fish, and we suggest that weaning process should be started at 34 dah.
Asunto(s)
Digestión , Hidrolasas/metabolismo , Perciformes/metabolismo , Animales , Larva/enzimología , Larva/crecimiento & desarrollo , Perciformes/crecimiento & desarrolloRESUMEN
The effects of prolonged feed deprivation (40 days at 18° C) and re-feeding (30 days) on body mass, growth and the activity of selected pancreatic and intestinal enzymes were evaluated in migrating European glass eels Anguilla anguilla by comparison with a control group fed to satiation with hake Merluccius merluccius roe for the duration of the experiment. Feed deprivation resulted in mass loss and a reduction in digestive function, as revealed by a decrease in the total and specific activities of pancreatic (trypsin and α-amylase) and intestinal brush border (alkaline phosphatase and leucine aminopeptidase) enzymes. The total activity of intestinal brush border enzymes diminished after 5 days of feed deprivation, whereas that of pancreatic enzymes did not decrease until 10 days, indicating that the intestine is more sensitive to feed deprivation than the pancreas. Re-feeding A. anguilla that were starved for 40 days resulted in compensatory growth, with specific growth rates that were 2·6 times higher than the control group. This compensatory growth was associated with the recovery of trypsin and intestinal brush border enzyme activities, which were restored to control levels within 5 days of re-feeding. The ability to maintain pancreatic enzyme activity during 40 days of feed deprivation, and rapidly recover capacity for protein digestion upon re-feeding, would enable A. anguilla at this glass eel stage to withstand periods without food but rapidly provide amino acids for protein synthesis and growth when suitable food was available.
Asunto(s)
Anguilla/fisiología , Migración Animal , Digestión/fisiología , Inanición/metabolismo , Fosfatasa Alcalina/fisiología , Anguilla/crecimiento & desarrollo , Animales , Intestinos/enzimología , Leucil Aminopeptidasa/fisiología , Páncreas/enzimología , Tripsina/fisiología , alfa-Amilasas/fisiologíaRESUMEN
Several samples of P. splendida larvae were obtained from eggs until day 60 after hatching (dah) to determine acid and alkaline proteases, trypsin, chymotrypsin, leucine aminopeptidase, α-amylase, lipase, and acid and alkaline phosphatase activities using biochemical techniques. Additionally, SDS-PAGE alkaline protease zymogram and PAGE acid protease zymogram were carried out to identify active isoforms during larviculture. Alkaline protease and chymotrypsin were present at the moment of hatching, increased gradually reaching the maximum values at 35 dah. Trypsin and leucine aminopeptidase activities were low from hatching, increasing gradually as larvae grew. Alkaline protease zymogram showed four zymogens, which appears at different days, remaining present until the end of the larviculture (95.2 kDa at 11 dah, 26.4 kDa at 9 dah, 21.4 kDa at 3 dah, and 23.3 kDa at hatching). Pepsin activity was present at day 7 after hatching and increased progressively until the end of the larviculture. Acid protease zymogram only showed one zymogen (0.65 rf), which appear at 6 dah. Lipase was high at the time of hatching and increased until 15 dah, after which decreased gradually. Amylase was high from the beginning and until 15 dah and then decreased rapidly to almost nothing onward. Alkaline and acid phosphatases presented a high activity at the egg stage, fell slightly during the first feeding and increased again from 20 to 30 dah. Results obtained in this study show that larvae can be fed artificial diets starting on day 10 after hatching.
Asunto(s)
Cíclidos/embriología , Cíclidos/metabolismo , Enzimas/metabolismo , Tracto Gastrointestinal/embriología , Tracto Gastrointestinal/metabolismo , Envejecimiento , Animales , Regulación del Desarrollo de la Expresión Génica/fisiología , Regulación Enzimológica de la Expresión Génica/fisiología , Larva/crecimiento & desarrolloRESUMEN
The development of digestive enzymes during the early ontogeny of the Mayan cichlid (Cichlasoma urophthalmus) was studied using biochemical and electrophoretic techniques. From yolk absorption (6 days after hatching: dah), larvae were fed Artemia nauplii until 15 dah, afterward they were fed with commercial microparticulated trout food (45% protein and 16% lipids) from 16 to 60 dah. Several samples were collected including yolk-sac larvae (considered as day 1 after hatching) and specimens up to 60 dah. Most digestive enzymes were present from yolk absorption (5-6 dah), except for the specific acid proteases activity (pepsin-like), which increase rapidly from 8 dah up to 20 dah. Three alkaline proteases isoforms (24.0, 24.8, 84.5 kDa) were detected at 8 dah using SDS-PAGE zymogram, corresponding to trypsin, chymotrypsin and probably leucine aminopeptidase enzymes, and only one isoform was detected (relative electromobility, Rf = 0.54) for acid proteases (pepsin-like) from 3 dah onwards using PAGE zymogram. We concluded that C. urophthamus is a precocious fish with a great capacity to digest all kinds of food items, including artificial diets provided from 13 dah.
Asunto(s)
Cíclidos/crecimiento & desarrollo , Enzimas/metabolismo , Animales , Larva/enzimología , Larva/crecimiento & desarrollo , Factores de TiempoRESUMEN
The activities of several digestive enzymes during larval development of the spotted sand bass (Paralabrax maculatofasciatus) were evaluated using electrophoretic techniques. The results show the presence of three isoforms of alkaline protease from day 2 after hatching (ah) and the early appearance of one pepsin-like band from day 12 ah onwards. In addition, two lipase bands first appeared on day 2 ah, and there was a change in the molecular weight of one band from day 15 ah onwards. Several alpha-amylase isoforms were observed from hatching up to day 5 ah. These results indicate that the important digestive enzymes develop rapidly in these larvae, supporting the possibility of early weaning at day 12 ah using artificial diets.
Asunto(s)
Lubina/crecimiento & desarrollo , Digestión/fisiología , Tracto Gastrointestinal/enzimología , Lipasa/metabolismo , Péptido Hidrolasas/metabolismo , alfa-Amilasas/metabolismo , Animales , Electroforesis en Gel de Poliacrilamida , Larva/enzimología , Fenómenos Fisiológicos de la NutriciónRESUMEN
Spotted sand bass Paralabrax maculatofasciatus is a potential aquaculture species in Northwest Mexico. In the last few years it has been possible to close its life cycle and to develop larviculture technology at on pilot scale using live food, however survival values are low (11%) and improvements in growth and survival requires the study of the morpho-physiological development during the initial ontogeny. In this research digestive activity of several enzymes were evaluated in larvae, from hatching to 30 days after hatching (dah), and in live prey (rotifers and Artemia), by use of biochemical and electrophoretic techniques. This paper, is the first of two parts, and covers only the biochemical analysis. All digestive enzyme activities were detected from mouth opening; however the, maximum activities varied among different digestive enzymes. For alkaline protease and trypsin the maximum activities were detected from 12 to 18 dah. Acid protease activity was observed from day 12 onwards. The other digestive enzymes appear between days 4 and 18 after hatching, with marked fluctuations. These activities indicate the beginning of the juvenile stage and the maturation of the digestive system, in agreement with changes that occur during morpho-physiological development and food changes from rotifers to Artemia. All enzymatic activities were detected in rotifers and Artemia, and their contribution to enhancement the digestion capacity of the larvae appears to be low, but cannot be minimised. We concluded that the enzymatic equipment of P. maculatofasciatus larvae is similar to that of other marine fish species, that it becomes complete between days 12 and 18 after hatching, and that it is totally efficient up to 25 dah.