The importance of pepsin‐like acid protease quantification in aquaculture studies: a revision of available procedures and presentation of a new protocol for its assessment

Assessing the activity of acid pepsin‐like protease is a common procedure in many biological, physiological and nutritional studies in gastric fish species. After reviewing the available literature on fish digestive acid proteases (stomach acid pepsin‐like proteases), especially those species of relevance for aquaculture purposes, published during a time span of ten years (2008–2018), authors detected some possible methodological and/or interpretative inconsistencies in this kind of studies when quantifying pepsin‐like enzymes. Thus, we concluded that special attention should be paid on the following issues when assessing digestive acid protease activity: (i) the proper preparation of stomach extracts, (ii) the use of the proper reaction conditions at the normal range of physiological pH and temperature values, and (iii) the proper quantification of the number of peptide bonds hydrolyzed for acid protease unit calculation. The implementation of these recommendations, and the proposal of more suitable (previously tested) methodological steps, will promote the standardization of acid pepsin‐like protease analytical procedures, as well as improve the reliability of comparative studies dealing with this proteolytic enzyme between different fish species of aquacultural importance.info:eu-repo/semantics/acceptedVersio

Composición y Método Cinético de Medición de la Actividad Lipasa

Número de publicación: ES2358029 A1 (05.05.2011) También publicado como: ES2358029 B2 (23.03.2012 Número de Solicitud: Consulta de Expedientes OEPM (C.E.O.) P200902117(20.10.2009)La presente invención pertenece al campo de las composiciones químicas utilizadas en métodos de determinación de actividad enzimática. En particular, la presente invención se encuadra en los métodos cinéticos de determinación de la actividad lipasa que utilizan sustratos sintéticos.Universidad de Almerí

Celdilla de digestión-Lixiviación con membrana semipermeable, sistema de cerrado hermético y de control de temperatura

Número de publicación: ES2524521 A1 (09.12.2014) También publicado como: ES2524521 B1 (13.10.2015)ES2524521 B8 (13.05.2016) Número de Solicitud: Consulta de Expedientes OEPM (C.E.O.) P201201027 (08.10.2012)La invención es una celdilla de digestión-lixiviación con membrana semipermeable, sistema de cerrado hermético y de control de temperatura que es un dispositivo para pruebas de digestibilidad y lixiviación de moléculas o iones a nivel laboratorio. La invención es un diseño novedoso que permite mantener aislada la cámara interna (donde se realiza la digestión-lixiviación de materiales) de la cámara externa (donde se reciben los productos de la digestión-lixiviación) conectadas exclusivamente a través de la membrana semipermeable, de tamaño de poro intercambiable, para el paso de las moléculas o iones de tamaño deseado. La celdilla contiene un dispositivo que permite mantener un control de la temperatura en el interior de la cámara interna. Las aplicaciones de la celdilla están en laboratorios académicos o de investigación (empresas, centros de investigación, institutos, Universidades) para la realización de pruebas de digestibilidad in vitro de macromoléculas o de lixiviación de moléculas o iones.Universidad de Almerí

Digestibilidad in vitro del dsRNA específico por enzimas del tracto digestivo del camarón Litopenaeus vannamei

Objective. The digestibility of specific dsRNA by action of the enzymes of digestive tract of the whiteleg shrimp Litopenaeus vannamei was determined in vitro. Materials and methods. Digestive enzyme activity (amylase, lipase, protease, DNase and RNase) was measured in the stomach, digestive gland, and anterior, middle, and posterior intestine of juvenile shrimp, and the digestibility of DNA, RNA and the dsRNA-ORF89, specific to WSSV, was determined by in vitro assays, as well as electrophoretic and densitometric analyses. Results. The highest enzymatic activity was found in the digestive gland: amylase (81.41%), lipase (92.60%), protease (78.20%), DNase (90.85%), and RNase (93.14%). The highest digestive capacity against DNA, RNA, and dsRNA was found in the digestive gland (5.11 ng of DNA per minute, 8.55 ng of RNA per minute, and 1.48 ng dsRNA per minute). Conclusions. The highest digestibility of dsRNA-ORF89, specific to WSSV, was found in the digestive gland, whereas the lowest digestibility was observed in the posterior intestine. This is the first report regarding the digestibility of dsRNA-ORF89 by whiteleg shrimp digestive tract enzymes, with potential therapeutic importance in shrimp culture to prevent WSSV disease through balanced feed.Objetivo. La digestibilidad del dsRNA específico para el virus de la mancha blanca (WSSV) por acción de las enzimas del tracto digestivo del camarón Litopenaeus vannamei fue analizada in vitro. Material y métodos. Se midió la actividad de enzimas digestivas (proteasa, amilasa, lipasa, ADNasa y ARNasa) en el estómago, la glándula digestiva, el intestino anterior, medio y posterior en juveniles de camarón patiblanco y se evaluó la digestibilidad de ácidos nucleicos ADN, ARN y dsRNA-ORF89 especifico contra el virus WSSV, por análisis electroforéticos y densitometría. Resultados. La actividad enzimática más alta se encontró en la glándula digestiva del camarón: amilasa (81.41%), lipasa (92.60%), proteasa (78.20%), ADNasa (90.85%) y ARNasa (93.14%). Se evidenció la capacidad digestiva del camarón patiblanco contra el ADN, ARN y dsRNA-ORF89 encontrando en la glándula digestiva la mayor digestión (5.11 ng de ADN por minuto, 8.55 ng de ARN por minuto y 1.48 ng de dsRNA por minuto). Conclusiones. La mayor digestibilidad del dsRNA-ORF89, específico contra el virus WSSV, se encontró en la glándula digestiva y la menor en el intestino posterior. Este es el primer informe relacionado con la digestibilidad del dsRNA-ORF89 por las enzimas del camarón patiblanco con potencial importancia terapéutica en el cultivo de camarón para prevenir la enfermedad del WSSV a través del alimento balanceado

Caracterización parcial de proteasas digestivas del chucumite (Centropomus parallelus)

Diversos estudios han abordado la caracterización de las diversas enzimas digestivas, lo cual es esencial para entender los tipos, modos de acción y nivel de actividad de dichas enzimas. Objetivos. La caracterización parcial de las proteasas digestivas se estudió en juveniles del chucumite (Centropomus parallelus, Poey 1869). Métodos. Se utilizaron técnicas bioquímicas y electroforéticas para la caracterización de proteasas. Resultados. La actividad de proteasas ácidas (estomacales) fue mayor a las proteasas alcalinas (intestinales). Asimismo, la actividad de la quimotripsina fue superior a la actividad de la tripsina. El pH óptimo para las proteasas estomacales fue de 2 con una alta estabilidad en el rango de pH 2-8, mientras que las proteasas intestinales mostraron dos picos con actividad a pH 8 y 10, siendo altamente estables a los pH 8 y 12. Las temperaturas óptimas se obtuvieron a 45 y 55 °C para proteasa estomacales e intestinales, respectivamente. Mediante el uso de inhibidores de proteasas se comprobó la presencia de pepsina en los extractos estomacales y de diversas serina proteasas en los extractos intestinales. Los zimogramas mostraron cinco bandas con actividad proteolítica alcalina (79.5, 48.9, 37.0, 29.0 y 22.5 kDa), las cuales fueron fuertemente inhibidas con PMSF, SBT1 y ovoalbúmina. Conclusiones. Por lo anterior, se considera que C. parallelus tiene una alta capacidad digestiva similar a otros peces marinos carnívoros estrictos

Palatability of animal oils included in the diet of the Mexican axolotl and its effect on growth and survival

Objective: The growth, survival and condition factor of Ambystoma mexicanum juveniles fed with three commercial feed-based diets coated with different animal oils as potential palatability enhancers were evaluated. Methodology: Three diets were prepared using commercial feed coated with fish, chicken and krill oil. The experiment lasted 81 days, the food was provided to the axolotls (6 per experimental unit, in triplicate) every 48 h, at 4% of the total biomass. The total weight of each experimental unit was recorded every 15 days, at the end of the experiment individual weight and height and survival were recorded. The following variables were calculated: Fulton's K, specific growth rate, coefficient of variation of final weight and size heterogeneity. Results: All three treatments showed an increase in total biomass, with a trend towards better performance in chicken and fish oil treatments compared to krill oil. When performing the statistical analysis, it was found that there were no significant differences among the treatments for any of the variables recorded. Implications: The three oils used in the present study were good palatable agents in the food intake of A. mexicanum. Conclusions: It is recommended to use chicken oil as an attractant additive in the formulation of a specific diet due to its low cost.Objective: The growth, survival, and condition factor of Ambystoma mexicanum juveniles fed with three commercial feed-based diets coated with different animal oils as potential palatability enhancers were evaluated. Methodology: Three diets were prepared using commercial feed coated with fish, chicken, and krill oil. The experiment lasted 81 days, the food was provided to the axolotls (6 per experimental unit, in triplicate) every 48 h, at 4% of the total biomass. The total weight of each experimental unit was recorded every 15 days, at the end of the experiment individual weight and height and survival were recorded. The following variables were calculated: Fulton's K, specific growth rate, coefficient of variation of final weight and size heterogeneity. Results: All three treatments showed an increase in total biomass, with a trend towards better performance in chicken and fish oil treatments compared to krill oil. When performing the statistical analysis, it was found that there were no significant differences among the treatments for any of the variables recorded. Implications: The three oils used in the present study were good palatable agents in the food intake of A. mexicanum. Conclusions: It is recommended to use chicken oil as an attractant additive in the formulation of a specific diet due to its low cost

Somatic indices and nutritional composition of the roe of the native fish Dormitator latifrons: no aplica

Objective: To evaluate some reproductive aspects of D. latifrons and the nutritional quality of its eggs. Design/methods/approach: Eighty-two fish were randomly collected (August 2021), and their sex, length, weight, somatic indices, Fulton condition index (K), proximate composition, and amino acid and lipid composition (fatty acids) of the gonads were determined. Results: Of the total specimens collected, 62% were females; length and weight values were higher in males, but their gonadosomatic index (GSI) was lower than in females. Somatic indices did not show differences between different weight ranges. In the roe of D. latifrons, the average proximate composition was 24.3% protein and 8.5% lipids. The most abundant essential amino acids were leucine and lysine. Linoleic acid (C18:2n6) was the fatty acid with the highest concentration. Limitations/implications: It is necessary to complement the analysis of the amino acid and fatty acid profile of the roe in wild organisms to relate the changes caused by balanced feed. Findings/conclusions: This study shows that the D. latifrons roe is a good source of amino acids and PUFA. Keywords: proximate composition, fatty acids, essential amino acidsObjective: To evaluate some reproductive aspects of D. latifrons and the nutritional quality of its eggs. Design/methods/approach: Eighty-two fish were randomly collected (August 2021), and their sex, length, weight, somatic indices, Fulton condition index (K), proximate composition, and amino acid and lipid composition (fatty acids) of the gonads were determined. Results: Of the total specimens collected, 62% were females; length and weight values were higher in males, but their gonadosomatic index (GSI) was lower than in females. Somatic indices did not show differences between different weight ranges. In the roe of D. latifrons, the average proximate composition was 24.3% protein and 8.5% lipids. The most abundant essential amino acids were leucine and lysine. Linoleic acid (C18:2n6) was the fatty acid with the highest concentration. Limitations/implications: It is necessary to complement the analysis of the amino acid and fatty acid profile of the roe in wild organisms to relate the changes caused by balanced feed. Findings/conclusions: This study shows that the D. latifrons roe is a good source of amino acids and PUFA