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Recibido el 25-08-2020 . Aceptado el 25-07-2020 *Autor de correspondencia. Correo electrónico: pinto_ruiz@yahoo.com.mx Nota técnica / Technical note Efecto del método de extracción de líquido ruminal en la técnica de gas in vitro Effect of the ruminal liquid extraction method in the in vitro gas technique Efeito do método de extração de líquido ruminal na técnica de gases in vitro Roselia Ramírez Díaz 1 , René Pinto Ruiz 2 *, Francisco Guevara Hernández 2 , José Apolonio Venegas Venegas 2 , Mariela Beatriz Reyes Sosa 1,2 y Luis Alberto Miranda Romero 2 1 Estudiante de Doctorado en Ciencias Agropecuarias y Sustentabilidad, Universidad iVBORw0KGgoAAAANSUhEUgAAABAAAAAQCAIAAACQkWg2AAABL0lEQVR4nGP5//8/Awzce7f1 /tvtr76cg4uI8RgpCnsqCXnDRVgg1Ndfzw/fq3j//TYDKgBqBqJbr1bZKnVws0lCNQBV77gR /+vvl0jDY1dezN1/s1eETwBZG9AgoAIPjYVAPSANQLOBqhnwAqACoDKgHhagu+EumXxQC00d GzMP3CygMqBiFqAv4dK59tcgTgK6DS745OOhkw9bINqAilmQwwQZAG0D+gQYShby1UYyBSce tkDCgAW/04EqTjxsdVadAtHAAA9W/HqQuYQ1AF2FogHIx+UNiGqgH+6/2wbnsgBjHqsGYIhB GMBQOvdkAoQNVMwCTCfAmIdExfLzVkASGDgQBhoQ5FQFKgb5AZhOIEkDj0+AMQhUxgDxNDCF AOMca+KDm42S+OB6iEneAKkQp/ZDwx7EAAAAAElFTkSuQmCC Autónoma de Chiapas. Correo electrónico: (RR) ramirez.rrd@gmail.com, . 2 Universidad Autónoma de Chiapas, Facultad de Ciencias Agronómicas, carretera Ocozocoautla-Villaflores km 84.5, Villaflores, Chiapas, México, C. P. 30470. Correo electrónico: (RP) pinto_ruiz@yahoo. iVBORw0KGgoAAAANSUhEUgAAABAAAAAQCAIAAACQkWg2AAABL0lEQVR4nGP5//8/Awzce7f1 /tvtr76cg4uI8RgpCnsqCXnDRVgg1Ndfzw/fq3j//TYDKgBqBqJbr1bZKnVws0lCNQBV77gR /+vvl0jDY1dezN1/s1eETwBZG9AgoAIPjYVAPSANQLOBqhnwAqACoDKgHhagu+EumXxQC00d GzMP3CygMqBiFqAv4dK59tcgTgK6DS745OOhkw9bINqAilmQwwQZAG0D+gQYShby1UYyBSce tkDCgAW/04EqTjxsdVadAtHAAA9W/HqQuYQ1AF2FogHIx+UNiGqgH+6/2wbnsgBjHqsGYIhB GMBQOvdkAoQNVMwCTCfAmIdExfLzVkASGDgQBhoQ5FQFKgb5AZhOIEkDj0+AMQhUxgDxNDCF AOMca+KDm42S+OB6iEneAKkQp/ZDwx7EAAAAAElFTkSuQmCC com.mx, iVBORw0KGgoAAAANSUhEUgAAABAAAAAQCAIAAACQkWg2AAABL0lEQVR4nGP5//8/Awzce7f1 /tvtr76cg4uI8RgpCnsqCXnDRVgg1Ndfzw/fq3j//TYDKgBqBqJbr1bZKnVws0lCNQBV77gR /+vvl0jDY1dezN1/s1eETwBZG9AgoAIPjYVAPSANQLOBqhnwAqACoDKgHhagu+EumXxQC00d GzMP3CygMqBiFqAv4dK59tcgTgK6DS745OOhkw9bINqAilmQwwQZAG0D+gQYShby1UYyBSce tkDCgAW/04EqTjxsdVadAtHAAA9W/HqQuYQ1AF2FogHIx+UNiGqgH+6/2wbnsgBjHqsGYIhB GMBQOvdkAoQNVMwCTCfAmIdExfLzVkASGDgQBhoQ5FQFKgb5AZhOIEkDj0+AMQhUxgDxNDCF AOMca+KDm42S+OB6iEneAKkQp/ZDwx7EAAAAAElFTkSuQmCC ; (FG) francisco.guevara@unach.mx, iVBORw0KGgoAAAANSUhEUgAAABAAAAAQCAIAAACQkWg2AAABL0lEQVR4nGP5//8/Awzce7f1 /tvtr76cg4uI8RgpCnsqCXnDRVgg1Ndfzw/fq3j//TYDKgBqBqJbr1bZKnVws0lCNQBV77gR /+vvl0jDY1dezN1/s1eETwBZG9AgoAIPjYVAPSANQLOBqhnwAqACoDKgHhagu+EumXxQC00d GzMP3CygMqBiFqAv4dK59tcgTgK6DS745OOhkw9bINqAilmQwwQZAG0D+gQYShby1UYyBSce tkDCgAW/04EqTjxsdVadAtHAAA9W/HqQuYQ1AF2FogHIx+UNiGqgH+6/2wbnsgBjHqsGYIhB GMBQOvdkAoQNVMwCTCfAmIdExfLzVkASGDgQBhoQ5FQFKgb5AZhOIEkDj0+AMQhUxgDxNDCF AOMca+KDm42S+OB6iEneAKkQp/ZDwx7EAAAAAElFTkSuQmCC ; (JV) polo_tex1@hotmail.com, ; (MR) iVBORw0KGgoAAAANSUhEUgAAABAAAAAQCAIAAACQkWg2AAABL0lEQVR4nGP5//8/Awzce7f1 /tvtr76cg4uI8RgpCnsqCXnDRVgg1Ndfzw/fq3j//TYDKgBqBqJbr1bZKnVws0lCNQBV77gR /+vvl0jDY1dezN1/s1eETwBZG9AgoAIPjYVAPSANQLOBqhnwAqACoDKgHhagu+EumXxQC00d GzMP3CygMqBiFqAv4dK59tcgTgK6DS745OOhkw9bINqAilmQwwQZAG0D+gQYShby1UYyBSce tkDCgAW/04EqTjxsdVadAtHAAA9W/HqQuYQ1AF2FogHIx+UNiGqgH+6/2wbnsgBjHqsGYIhB GMBQOvdkAoQNVMwCTCfAmIdExfLzVkASGDgQBhoQ5FQFKgb5AZhOIEkDj0+AMQhUxgDxNDCF AOMca+KDm42S+OB6iEneAKkQp/ZDwx7EAAAAAElFTkSuQmCC mareso84@gmail.com, iVBORw0KGgoAAAANSUhEUgAAABAAAAAQCAIAAACQkWg2AAABL0lEQVR4nGP5//8/Awzce7f1 /tvtr76cg4uI8RgpCnsqCXnDRVgg1Ndfzw/fq3j//TYDKgBqBqJbr1bZKnVws0lCNQBV77gR /+vvl0jDY1dezN1/s1eETwBZG9AgoAIPjYVAPSANQLOBqhnwAqACoDKgHhagu+EumXxQC00d GzMP3CygMqBiFqAv4dK59tcgTgK6DS745OOhkw9bINqAilmQwwQZAG0D+gQYShby1UYyBSce tkDCgAW/04EqTjxsdVadAtHAAA9W/HqQuYQ1AF2FogHIx+UNiGqgH+6/2wbnsgBjHqsGYIhB GMBQOvdkAoQNVMwCTCfAmIdExfLzVkASGDgQBhoQ5FQFKgb5AZhOIEkDj0+AMQhUxgDxNDCF AOMca+KDm42S+OB6iEneAKkQp/ZDwx7EAAAAAElFTkSuQmCC ; (LM) debraj.aryal@hotmail.com, . . Resumen Los estudios de gas in vitro requieren del uso de animales canulados; sin embargo, el alto costo de la cirugía limita su aplicación. Por lo que, el objetivo del estudio fue evaluar el efecto de la técnica de extracción de líquido ruminal por sonda oro-ruminal sobre los resultados derivados de la técnica de producción de gas in vitro (TPG). La harina de Elaeis guineensis se utilizó como sustrato. El líquido ruminal se obtuvo de dos formas: de animales canulados y sonda oro-ruminal. Se estimó la degradación de la materia seca (DGRMS), parámetros de la cinética de fermentación y volumen fraccional por la TPG. La comparación de medias se realizó utilizando el procedimiento de Tukey (p<0,05). Los resultados indicaron la falta de diferencias estadísticas significativas (p>0,05). Se concluyó que, la técnica de extracción de líquido ruminal, no modificó la DGRMS, los parámetros y fracciones de fermentación de E. guineensis. Palabras clave: in vitro, ovinos, degradación, fermentación. 682 Esta publicación científica en formato digital es continuación de la Revista Impresa: Depósito legal pp 196802ZU42, ISSN 0378-7818. Rev. Fac. Agron. (LUZ). 2021, 38(3): 681-691. Julio-Septiembre. Ramírez et al. ISSN 2477-9407 Abstract In vitro gas studies require the use of cannulated animals; however, the high cost of the surgery limits its application. Therefore, the objective of the study was to evaluate the effect of the ruminal fluid extraction technique by oro-ruminal probe affects the results derived from the in vitro gas production technique (GPT). Elaeis guineensis meal was used as a substrate. Ruminal fluid was obtained in two ways: cannulated animals and oro-ruminal probe. Dry matter degradation was estimated (DMD), parameters of fermentation kinetics and fractional volume by the GPT. The comparison of means was carried out using the Tukey procedure (p<0.05). The results indicated the lack of significant statistical differences (p>0.05). It was concluded that the ruminal fluid extraction technique did not modify the DGRMS, the parameters and fermentation fractions of E. guineensis. Key words: in vitro, sheep, degradation, fermentation. Resumo Os estudos de gases in vitro requerem o uso de animais canulados; entretanto, o alto custo da cirurgia limita sua aplicação. Portanto, o objetivo do estudo foi avaliar o efeito da técnica de extração de fluido ruminal por sonda ororruminal sobre os resultados derivados da técnica de produção de gás in vitro (TPG). Farinha de Elaeis guineensis foi usada como substrato. O líquido ruminal foi obtido de duas formas: de animais canulados e sonda ororruminal. A degradação da matéria seca (DGRMS), os parâmetros cinéticos da fermentação e o volume fracionário foram estimados por TPG. A comparação das médias foi realizada pelo procedimento de Tukey (p<0,05). Os resultados indicaram a ausência de diferenças estatísticas significativas (p>0,05). Concluiu-se que a técnica de extração do fluido ruminal não modificou o DGRMS, os parâmetros e as frações de fermentação de E. guineensis. Palavras-chave: in vitro, ovelha, degradação, fermentação. Introducción La técnica de producción de gas in vitro es ampliamente usada para determinar cinética de fermentación de alimentos a través del volumen de gas liberado, producto de la fermentación (Posada y Noguera, 2005), también permite conocer las fracciones de fermentación y estimar energía metabolizable, energía Introduction The in vitro gas production technique is widely used to determine the kinetics of food fermentation through the volume of gas released, a product of fermentation (Posada and Noguera, 2005), it also allows to know the fermentation fractions and estimate metabolizable energy, net energy, digestibility of organic matter, 683 Esta publicación científica en formato digital es continuación de la Revista Impresa: Depósito legal pp 196802ZU42, ISSN 0378-7818. Rev. Fac. Agron. (LUZ). 2021, 38(3): 681-691. Julio-Septiembre. Ramírez et al. ISSN 2477-9407 neta, digestibilidad de la materia orgánica, biomasa microbiana y gases efecto invernadero por fermentación entérica (Jiménez-Santiago et al., 2019, Ramírez et al., 2020). No obstante, para realizar este tipo de investigaciones, es necesario contar con animales canulados para obtener inóculo a partir del líquido ruminal (Vargas-Bayona et al., 2013), lo que limita su implementación en diversos laboratorios, debido a los altos costos de la cirugía, el manejo sanitario de los animales (Martín et al., 2005) y, en algunos casos, la mortalidad post cirugía, debido a la susceptibilidad de los animales a enfermedades. Por tal razón se han utilizado otras técnicas para obtener líquido ruminal (LR), como es el uso de la sonda oro-ruminal, la cual es económica, práctica y menos invasiva (Shen et al., 2012). Esta técnica se utilizó con éxito para hacer transfaunaciones de animales saludables a otros con desórdenes digestivos y enfermedades metabólicas (Pereira et al., 2018). Si bien, el uso de animales canulados para la extracción de LR es considerada una forma convincente con relación a los resultados de los estudios que arroja, también deben considerarse las ventajas del uso de sondas oro-ruminales. Sin embargo, no existe información que indique la eficiencia del método de extracción de líquido ruminal por sonda oro- ruminal en los resultados derivados de la técnica de gas in vitro, pero se demostró su eficacia al obtener muestras ruminales para conocer el desarrollo de poblaciones microbianas del rumen, determinación de ácidos microbial biomass and greenhouse gases due to enteric fermentation (Jiménez-Santiago et al., 2019; Ramírez et al., 2020). However, to carry out this type of research, it is necessary to have cannulated animals to obtain inoculum from ruminal fluid (Vargas-Bayona et al., 2013), which limits its implementation in various laboratories, due to the high costs of surgery, animal health management (Martín et al., 2005) and, in some cases, post-surgery mortality due to the animals’ susceptibility to diseases. For this reason, other techniques have been used to obtain ruminal fluid (RL), such as the use of the oro-ruminal probe, which is economical, practical and less invasive (Shen et al., 2012). This technique was used successfully to transfaun healthy animals to others with digestive disorders and metabolic diseases (Pereira et al., 2018). Although the use of cannulated animals for the extraction of LR is considered a convincing way in relation to the results of the studies it produces, the advantages of the use of oro-ruminal probes should also be considered. However, there is no information that indicates the efficiency of the ruminal fluid extraction method by oro-ruminal probe in the results derived from the in vitro gas technique, but its effectiveness was demonstrated when obtaining ruminal samples to know the development of microbial populations of the rumen, determination of volatile fatty acids, minerals and pH (Ramos-Morales et al., 2014, Lourenco et al., 2019). Therefore, the objective of the present study was to evaluate 684 Esta publicación científica en formato digital es continuación de la Revista Impresa: Depósito legal pp 196802ZU42, ISSN 0378-7818. Rev. Fac. Agron. (LUZ). 2021, 38(3): 681-691. Julio-Septiembre. Ramírez et al. ISSN 2477-9407 grasos volátiles, minerales y pH (Ramos-Morales et al., 2014, Lourenco et al., 2019). Por lo anterior, el objetivo del presente estudio fue evaluar y comparar los resultados obtenidos en la técnica de gas in vitro al utilizar líquido ruminal obtenido por cánula o sonda oro-ruminal. Materiales y métodos La investigación se realizó en el Laboratorio de Nutrición Animal de la Facultad de Ciencias Agronómicas de la Universidad Autónoma Chiapas, ubicado en el municipio de Villaflores, Chiapas. La región presenta una precipitación media anual de 1.100 mm y una temperatura media anual de 25 °C. Se utilizó como sustrato harina de palmiste (Elaeis guineensis) y fue analizada para determinar materia seca (MS), proteína cruda (PC) según AOAC (2000), fibra detergente neutro (FDN) y fibra detergente ácida (FDA) de acuerdo con la técnica descrita por Van Soest et al. (1991). El líquido ruminal (LR) fue extraído de tres ovinos machos canulados (CR) y de tres ovinos por sonda oro-ruminal (SR), todos de la raza Katahdin de 42 ± 1,5 kg y de aproximadamente un año de edad. Los animales se alimentaron con una dieta isoproteíca e isoenergética compuesta de 60 % de pasto Cynodon nlemfuensis, 26 % maíz y 14 % harina de palmiste, la cual fue formulada para satisfacer sus necesidades nutricionales de acuerdo a NRC (National Research Council) (NRC, 2007) y en todo momento tuvieron un trato de acuerdo con los protocolos de la Ley Federal and compare the results obtained in the in vitro gas technique when using ruminal fluid obtained by cannula or oro-ruminal probe. Materials and methods The research was carried out in the Laboratorio de Nutrición Animal de la Facultad de Ciencias Agronómicas of the Universidad Autónoma Chiapas, located in the municipality of Villaflores, Chiapas. The region has an average annual rainfall of 1,100 mm and an average annual temperature of 25 °C. Palm kernel cake (Elaeis guineensis) was used as substrate and it was analyzed to determine dry matter (DM), crude protein (PC) according to AOAC (2000), neutral detergent fiber (NDF) and acid detergent fiber (FDA) according to the technique described by Van Soest et al. (1991). The ruminal fluid (LR) was extracted from three cannulated male sheep (CR) and from three sheep by oro-ruminal probe (SR), all of the Katahdin breed, 42 ± 1.5 kg and approximately one year old. The animals were fed an isoprotein and isoenergetic diet composed of 60 % Cynodon nlemfuensis grass, 26 % corn and 14 % palm kernel cake, which was formulated to meet their nutritional needs according to NRC (National Research Council) (NRC, 2007) and at all times they were treated in accordance with the protocols of the Federal Animal Health Law in force, NOM-062-ZOO-1999 and under the animal welfare standards established in Facultad de Ciencias Agronómicas 685 Esta publicación científica en formato digital es continuación de la Revista Impresa: Depósito legal pp 196802ZU42, ISSN 0378-7818. Rev. Fac. Agron. (LUZ). 2021, 38(3): 681-691. Julio-Septiembre. Ramírez et al. ISSN 2477-9407 de Sanidad Animal vigente, NOM- 062-ZOO-1999 y bajo las normas de bienestar animal establecidas en la Facultad de Ciencias Agronómicas de la Universidad Autónoma de Chiapas. Antes de la extracción de LR, los ovinos tuvieron un periodo de ayuno de 12 h, ello con la finalidad de conseguir un fluido ruminal mas consistente en composición y actividad (Williams, 2000 y Posada y Noguera, 2005). Para la extracción de LR por SR se utilizó una manguera de 1,25 m de longitud con un diámetro externo de 1” y 3 mm de espesor de pared, con perforaciones en el extremo que fue introducido en el saco ruminal, el extremo opuesto se conectó a un matraz kitasato para colectar el LR y el cual a su vez fue conectado a una bomba de succión. Para introducir la sonda, se utilizó un abrebocas metálico cubierto con una banda de caucho para evitar lesiones. Se extrajo 100 mL de LR/ animal/muestreo, el exceso de saliva se retiró previo a usar el LR. La cinética de fermentación, fracciones y degradación de la materia seca in vitro se determinó mediante el procedimiento descrito por Menke y Steigass (1988), para lo cual se usaron frascos color ámbar de 125 mL de capacidad a los que se les colocó 0,5 g de MS de harina de palmiste. Se preparó una solución mineral reducida compuesta de K 2 HPO 4 (0,45 g.L -1 ), KH 2 PO 4 (0,45 g.L -1 ), NaCO 3 (0,6 g.L -1 ), (NH4) 2 SO 4 (0,45 g.L -1 ), NaCl (0,9 g.L -1 ), MnSO 4 (0,18 g.L -1 ), CaCl 2 (0,12 g.L -1 ), L-cisteína (0,25 g.L -1 ) y Na 2 S (0,25 g.L -1 ) y se usaron dos fuentes de líquido ruminal, correspondiente a cada método de de la Universidad Autónoma de Chiapas. Before the LR extraction, the sheep had a fasting period of 12 h, this in order to achieve a more consistent ruminal fluid in composition and activity (Williams, 2000 and Posada and Noguera, 2005). For the LR extraction by SR, a 1.25 m long hose with an external diameter of 1” and 3 mm wall thickness was used, with perforations at the end that was introduced into the ruminal sac, the opposite end was it was connected to a kitasato flask to collect the LR and which in turn was connected to a suction pump. To insert the probe, a metal gag covered with a rubber band was used to prevent injury. One hundred (100) mL of LR/ animal/sample was extracted, excess saliva was removed prior to using the LR. The in vitro kinetics of fermentation, fractions and degradation of dry matter were determined by the procedure described by Menke and Steigass (1988), for which amber bottles of 125 mL capacity were used, to which 0.5 g of DM of palm kernel cake was placed. A reduced mineral solution composed of K 2 HPO 4 (0.45 g.L -1 ), KH 2 PO 4 (0.45 g.L -1 ), NaCO 3 (0.6 g.L -1 ), (NH 4 ) 2 SO 4 (0.45 g.L -1 ) was prepared, NaCl (0.9 g.L -1 ), MnSO 4 (0.18 g.L -1 ), CaCl 2 (0.12 g.L -1 ), L-cysteine (0.25 g.L -1 ) and Na 2 S (0.25 g.L -1 ) and two sources of ruminal liquid were used, corresponding to each extraction method (cannula and oro-ruminal probe), subsequently and under a continuous flow of carbon dioxide (CO 2 ), 90 mL of inoculum were added to each flask diluted ruminal (1:10) according to treatment. 686 Esta publicación científica en formato digital es continuación de la Revista Impresa: Depósito legal pp 196802ZU42, ISSN 0378-7818. Rev. Fac. Agron. (LUZ). 2021, 38(3): 681-691. Julio-Septiembre. Ramírez et al. ISSN 2477-9407 extracción (cánula y sonda oro- ruminal), posteriormente y bajo un flujo continuo de bióxido de carbono (CO 2 ), a cada frasco se le agregaron 90 mL de inóculo ruminal diluido (1:10) de acuerdo al tratamiento. La presión del gas producido se midió con un manómetro marca Infra modelo 63100/1-4 (0 a 1 kg.cm -2 ) a 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 24, 28, 32, 36, 40, 44, 48, 54, 60, 66 y 72 horas de incubación. Los valores de presión (kg.cm -2 ) se transformaron a volumen de gas (mL.g -1 materia seca) con la ecuación de regresión (volumen= presión/0,019 con R 2 =0,98), y se estimaron los parámetros de la cinética de producción de gas: volumen máximo (Vmáx; mL.g -1 ), tasa (S; h) y fase de retardo (L; h), para el modelo logístico V = Vmáx/1+e (2-4*S (T-L)) (Schofield y Pell, 1995), utilizando el Sistema de Análisis Estadístico SAS (SAS, 2011). La degradación in vitro de la materia seca (DGRMS) se determinó a 72 h. Se calculó por diferencia entre el peso de la materia seca inicial, antes de ser incubada, y el peso de la materia seca residual después de 72 h de incubación. Al final del periodo de incubación, el contenido de cada frasco se filtró a través de papel de filtrado previamente pesado. Los papeles con residuo se secaron a 65 °C por 48 h, se pesaron y se restó el peso del papel filtro. Las fracciones de fermentación se obtuvieron mediante el volumen fraccional (Vf) de gas de fermentación producido a tres intervalos de tiempo: 0 h a 8 h (Vf 0-8 ), 8 h a 24 h (Vf 8-24 ) y 24 h a 72 h (Vf 24-72 ) de incubación, que corresponden a carbohidratos solubles, carbohidratos de reserva The pressure of the gas produced was measured with an Infra model 63100 / 1-4 (0 to 1 kg.cm -2 ) manometer at 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 24, 28, 32, 36, 40, 44, 48, 54, 60, 66 and 72 hours of incubation. The pressure values (kg.cm -2 ) were transformed into gas volume (mL.g -1 dry matter) with the regression equation (volume = pressure/0.019 with R 2 = 0.98), and the parameters of gas production kinetics were estimated: maximum volume (Vmax; mL.g -1 ), rate (S; h) and lag phase (L; h), for the logistic model V = Vmax/1 + e (2-4 * S (TL)) (Schofield and Pell, 1995), using the SAS Statistical Analysis System (SAS, 2011). In vitro degradation of dry matter (DGRMS) was determined at 72 h. It was calculated by the difference between the weight of the initial dry matter, before being incubated, and the weight of the residual dry matter after 72 h of incubation. At the end of the incubation period, the contents of each vial were filtered through pre-weighed filter paper. The papers with residue were dried at 65 °C for 48 h, weighed and the weight of the filter paper was substracted. The fermentation fractions were obtained by the fractional volume (Vf) of fermentation gas produced at three time intervals: 0 h to 8 h (Vf 0-8 ), 8 h to 24 h (Vf 8-24 ) and 24 h to 72 h (Vf 24-72 ) of incubation, which correspond to soluble carbohydrates, reserve carbohydrates and structural carbohydrates, respectively (Sandoval et al., 2016). These fractional volumes (mL.g -1 ) were transformed into fractions (g.kg -1 ) of fast (FR), medium (FM) and slow (FL) fermentation by means of the following regression 687 Esta publicación científica en formato digital es continuación de la Revista Impresa: Depósito legal pp 196802ZU42, ISSN 0378-7818. Rev. Fac. Agron. (LUZ). 2021, 38(3): 681-691. Julio-Septiembre. Ramírez et al. ISSN 2477-9407 y carbohidratos estructurales, respectivamente (Sandoval et al., 2016). Estos volúmenes fraccionales (mL.g -1 ) fueron transformados a fracciones (g.kg -1 ) de rápida (FR), media (FM) y lenta (FL) fermentación mediante las siguientes ecuaciones de regresión (Miranda et al., 2015): FR (g.kg -1 ) = Vf 0-8 /0,4266 (R 2 = 0,944 1), FM (g.kg -1 ) = Vf 8-24 / 0,6152 (R 2 = 0,998), FL (g.kg -1 ) = Vf 24-72 /0,3453 (R 2 = 0,965 3). Se evaluaron dos tratamientos con nueve repeticiones en cada uno, las cuales fueron obtenidas de los valores medios, productos de la repetición por tres veces consecutivas del experimento, según recomienda Udén et al. (2012). Se utilizó un diseño completamente al azar con dos tratamientos y nueve repeticiones. Los datos obtenidos de las variables estudiadas se analizaron mediante PROC GLM (SAS, 2011) y las medias de tratamientos se compararon con la prueba de Tukey (p<0,05). Resultados y discusión El contenido de MS de la harina de palmiste fue de 91,9 %, con 14,47 % de PC, 69,7 % de FDN y 45,56 % de FDA. Los parámetros de la cinética de fermentación y las fracciones (rápida, media y lenta) de harina de Elaeis guineensis inoculada con LR obtenido por CR y por SR no presentaron diferencias estadísticas (p>0,05; Cuadro 1). La tasa de fermentación promedio fue de 0,0275 h -1 , con una L de 6,575 h, mientras que el Vmáx promedio equations (Miranda et al., 2015): FR (g.kg -1 ) = Vf 0-8 /0.4266 (R 2 = 0.944 1), FM (g.kg -1 ) = Vf 8-24 /0.6152 (R 2 = 0.998), FL (g .kg -1 ) = Vf 24-72 /0.3453 (R 2 = 0.965 3). Two treatments were evaluated with nine repetitions in each one, which were obtained from the mean values, products of the repetition for three consecutive times of the experiment, as recommended by Udén et al. (2012). A completely randomized design with two treatments and nine repetitions was used. The data obtained from the variables studied were analyzed using PROC GLM (SAS, 2011) and the treatment means were compared with the Tukey test (p <0.05). Results and discussion The DM content of the palm kernel cake was 91.9 %, with 14.47 % of CP, 69.7 % of NDF and 45.56 % of FDA. The parameters of the fermentation kinetics and the fractions (fast, medium and slow) of Elaeis guineensis meal inoculated with LR obtained by CR and SR did not present statistical differences (p> 0.05; Table 1). The average fermentation rate was 0.0275 h -1 , with an L of 6.575 h, while the average Vmax was 316.81 mL.g -1 . For its part, the DGRMS of E. guineensis meal at 72 h of incubation showed a similar behavior in both extraction methods (p> 0.05). The lack of differences between the two LR extraction techniques on the variables evaluated is probably due to the fact that the LR obtained by SR 688 Esta publicación científica en formato digital es continuación de la Revista Impresa: Depósito legal pp 196802ZU42, ISSN 0378-7818. Rev. Fac. Agron. (LUZ). 2021, 38(3): 681-691. Julio-Septiembre. Ramírez et al. ISSN 2477-9407 fue de 316,81 mL.g -1 . Por su parte, la DGRMS de la harina de E. guineensis a las 72 h de incubación mostró un comportamiento similar en ambos métodos de extracción (p>0,05). La falta de diferencias entre ambas técnicas de extracción de LR sobre las variables evaluadas, probablemente se deba a que el LR obtenido por SR, no se contaminó con saliva y fue representativo. Lo que concuerda con Martín et al. (2005), quienes cuantificaron las poblaciones de bacterias y hongos ruminales en muestras de contenido ruminal obtenidos por CR y SR en bovinos y señalaron que, mientras el LR no se contamine por saliva los resultados serán similares ente sí. Por su parte, Shen et al. (2012) y Ramos-Morales et al. (2014), no encontraron diferencias en la estimación de ácidos grasos volátiles, amoniaco, lactato, potasio, cloruro, calcio y fósforo y pH, cuando usaron SR y animales canulados, por lo que concluyeron que el uso de SR podría ser una alternativa para tomar muestras ruminales. Por otro lado, Terré et al. (2013), no encontraron diferencias en la cuantificación de bacterias gramnegativas y grampositivas al extraer LR por CR y SR, sin embargo, si se presentaron diferencias en la cuantificación de pH y AGV, concluyendo que, a pesar de las diferencias encontradas los resultados obtenidos con las dos técnicas están altamente correlacionadas, por lo que, se puede utilizar un factor de corrección para aproximar ambos métodos y permitir comparaciones entre estudios. was not contaminated with saliva and was representative. What agrees with Martín et al. (2005), who quantified the populations of ruminal bacteria and fungi in samples of ruminal content obtained by CR and SR in bovines and pointed out that, as long as the LR is not contaminated by saliva, the results will be similar to each other. For their part, Shen et al. (2012) and Ramos-Morales et al. (2014), found no differences in the estimation of volatile fatty acids, ammonia, lactate, potassium, chloride, calcium and phosphorus and pH, when they used SR and cannulated animals, for which they concluded that the use of SR could be an alternative to take ruminal samples. On the other hand, Terré et al. (2013), did not find differences in the quantification of gram-negative and gram-positive bacteria when extracting LR by CR and SR, however, there were differences in the quantification of pH and AGV, concluding that, despite the differences found, the results obtained with the two techniques they are highly correlated, therefore, a correction factor can be used to approximate both methods and allow comparisons between studies. Conclusions The extraction of ruminal fluid by cannula or oro-ruminal probe does not affect the results obtained in the in vitro gas technique, so they could be used interchangeably. Acnowledgement The authors thank the Instituto de Ciencia, Tecnología e Innovación 689 Esta publicación científica en formato digital es continuación de la Revista Impresa: Depósito legal pp 196802ZU42, ISSN 0378-7818. Rev. Fac. Agron. (LUZ). 2021, 38(3): 681-691. Julio-Septiembre. Ramírez et al. ISSN 2477-9407 Conclusiones La extracción de líquido ruminal por cánula o sonda oro-ruminal no afecta los resultados obtenidos en la técnica de gas in vitro, por lo que podrían utilizarse indistintamente. Agradecimientos Los autores agradecen al Instituto de Ciencia, Tecnología e Innovación del estado de Chiapas, México, por el apoyo proporcionado en la convocatoria del Sistema Estatal de Investigadores 2019. Literatura citada Jiménez-Santiago, Á., G., Jiménez-Ferrer, A., Alayón-Gamboa, E. J., Pérez- Cuadro 1. Efecto de LR obtenido por cánula y sonda oro-ruminal sobre la degradación de la materia seca, parámetros y fracciones de fermentación de harina de Elaeis guineensis. Table 1. Effect of LR obtained by cannula and oro-ruminal probe on the degradation of dry matter, parameters and fermentation fractions of Elaeis guineensis meal. Variables Cánula Sonda oro ruminal Promedio DGRMS 72 h (%) 43,45 ± 2,70 45,59 ± 1,80 44,520 S (h -1 ) 0,028 ± 0,02 0,027 ± 0,03 0,0275 L (h) 6,52 ± 1,90 6,63 ± 0,25 6,575 Vmáx (mL.g -1 ) 323,26 ± 21,64 310,36 ± 4,44 316,810 FR (g.kg -1 ) 131,08 ± 26,08 150,99 ± 2,30 141,035 FM (g.kg -1 ) 167,70 ± 19,07 142,01 ± 4,92 154,855 FL (g.kg -1 ) 476,93 ± 18,14 482,83 ± 49,80 479,880 FT (g.kg -1 ) 775,71 ± 49,98 753,02 ± 16,20 764,365 S: Tasa de fermentación, L: Fase de retardo, Vmáx: Volumen máximo de gas, FR: Fracción rápida, FM: Fracción media, FL: Fracción lenta, FT: Fracción Total, DGRMS: Degradación de la materia seca, ±: desviación estándar. 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