A multi-glycomic platform for the analysis of food carbohydrates - Monosaccharide, linkage and polysaccharide (FITDOG) composition analyses of different varieties of apple (figure 5)

Carbohydrates comprise the largest fraction of most diets and exert a profound impact on health. Components such as simple sugars and starch supply energy while indigestible components, deemed dietary fiber, reach the colon to provide food for the tens of trillions of microbes that make up the gut microbiota. The interactions between dietary carbohydrates, our gastrointestinal tracts, the gut microbiome, and host health are dictated by their structures. However, current methods for analysis of food glycans lack the sensitivity, specificity, and throughput needed to quantify and elucidate these myriad structures. This protocol describes a multi-glycomic approach to food carbohydrate analysis where the analyte might be any food item or digested product. The carbohydrates are extracted by… and the resulting samples are subjected to employing rapid-throughput liquid chromatography-tandem mass spectrometry (LC-MS/MS) methods. Quantitative analyses of monosaccharides, glycosidic linkages, polysaccharides, and alcohol soluble carbohydrates are performed in 96-well plate format to reduce the biomass of sample required and enhance throughput. Detailed stepwise processes for sample preparation, LC-MS/MS, and data analysis are provided. We illustrate the application of the protocol to a diverse set of foods as well as different apple cultivars and various fermented foods. Furthermore, we show the utility of these methods in elucidating glycan-microbe interactions in germ-free and colonized mice. These methods provide a framework for elucidating relationships between dietary fiber, the gut microbiome, and human physiology. These structures will further guide nutritional and clinical feeding studies that enhance our understanding of the role of diet in nutrition and health.

碳水化合物是多数膳食中占比最高的营养组分，对人体健康具有深远影响。单糖与淀粉等组分可为机体提供能量，而被归类为膳食纤维的难消化组分则可抵达结肠，为构成肠道菌群的数十万亿微生物提供营养基质。膳食碳水化合物、胃肠道、肠道微生物组与宿主健康之间的相互作用，由其分子结构所决定。然而，当前食品聚糖分析方法在量化与阐明这类海量结构所需的灵敏度、特异性及通量上均存在明显不足。本方案介绍一种面向食品碳水化合物分析的多聚糖组学方法，其分析物可覆盖任意食品或消化产物。碳水化合物经提取后，所得样品将采用高通量液相色谱-串联质谱（LC-MS/MS）方法进行分析。以96孔板形式开展单糖、糖苷键、多糖及醇溶性碳水化合物的定量分析，可减少所需样品生物量并提升分析通量。本方案提供了样品制备、液相色谱-串联质谱分析及数据分析的详细分步流程。我们以多种食品、不同苹果品种及各类发酵食品为例，展示了本方案的应用效果。此外，我们还验证了这些方法在阐明无菌小鼠与定植小鼠体内聚糖-微生物相互作用方面的实用性。这些方法为解析膳食纤维、肠道微生物组与人体生理之间的关联提供了标准化研究框架。相关结构信息将进一步指导营养学与临床喂养研究，加深我们对膳食在营养与健康中所扮演角色的理解。