本试验旨在探究一种猪胃肠道内容物与粪便微生物梯度分离方法。试验首先用分离缓冲液对猪胃肠道内容物及粪便样品进行稀释，寻找最佳稀释比例，再进行梯度离心得到胃肠道内容物及粪便样品中的微生物菌体。本方法得到胃、空肠和回肠内容物样品与分离缓冲液的最佳稀释比例范围为8.0%~12.0%，样品的称取量范围为3.2~4.8 g；盲肠、结肠内容物及粪便样品与分离缓冲液的最佳稀释比例范围为2.0%~4.0%，样品的称取量范围为0.8~1.6 g。对饲喂3种饲粮蛋白质水平（12.0%、15.0%和18.0%）30 kg的猪胃肠道内容物与粪便微生物进行分离，并对其回肠内容物和粪便中微生物氨基酸组成以及胃肠道内容物与粪便中微生物多样性进行检测。结果显示：回肠内容物和粪便中微生物菌体大部分氨基酸对饲粮不同蛋白质水平的营养模式响应效果显著（P<0.05），胃肠道内容物与粪便样品微生物的聚合酶链反应-变性梯度凝胶电泳（PCR-DGGE）条带丰富和清晰。试验结果表明，该方法分离程度高，获取菌体干重多，同时还具有成本低廉、操作简单、重复性好等特点，可为更深层次的研究肠道微生物的生理功能提供方法。

This experiment was conducted to investigate a gradient centrifugation separation method for microorganisms in gastrointestinal contents and faeces of swine. First, dilute the gastrointestinal contents and faeces with separation buffer to find the fittest dilution ratio. Then gradiently centrifuge and acquire microbial bacteria from gastrointestinal contents and faeces samples. By this method, the best dilution ratio of gastro, jejunum and ileum contents to separation buffer ranged from 8.0% to 12.0%, and the weight of samples ranged from 3.2 to 4.8 g. The best dilution ratio of cecum, colon contents and faeces ranged from 2.0% to 8.0%, and the weight ranged from 0.8 to 1.6 g. We detected the amino acids composition of microorganisms in ileum and faeces as well as the diversity of microorganisms in gastrointestinal content and faeces of weighed at 30 kg pigs fed with three kinds of protein level diets (12.0%, 15.0% and 18.0%). The results showed that the most amino acids composition of microbes in ileum contents and faeces were significantly responsible to different protein levels of nutritional model (P<0.05). The bands in polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) figure were rich and legible with microorganisms separated from gastrointestinal contents and faeces. The study demonstrates that this method has high separation degree and obtains substantial dry cell weight. Meanwhile it owns advantages of less cost, easy operation and good repeatability, which provides a method for studying the physiological function of gastrointestinal microbes deeply.