Evaluation of freeze-drying for quantification of the microbiome and metabolome in neonatal faecal samples

Appropriate sampling and storage processes are crucial for the accurate determination of the gut microbiome and metabolome. The present study evaluated the effect of the freeze-drying process on meconium and regular faecal samples of newborn babies. Triplicate samples were freeze-dried for 6 or 12 ​h and compared to samples that were only frozen. Microbiome analysis using 16 ​S rRNA sequencing showed that the microbial composition was similar between samples with freeze-drying and those without freeze-drying in general, while the determination efficiency of Bifidobacterium was significantly increased after freeze-drying for either time. A total of 300 commonly observed gut metabolites were quantified and classified into 13 categories, including amino acids, bile acids, carbohydrates, carnitines, fatty acids and so on. The compositions of most metabolites were consistently preserved before or after freeze-drying for the two time protocols. Moreover, the sensitivity of detection was significantly improved after freeze-drying, even for volatile compounds such as short-chain fatty acids. Accordingly, freeze-drying for 6 ​h is recommended for the pretreatment of bulk meconium and regular faecal samples for the determination of gut microbiota.