Analysis Of Diversity Of Bacteria And Small Molecule Compounds In The 3 Kefir Samples

Kefir is a traditional fermented milk with various of health care functions,which was derived from the fermentation of Kefir grains with milk as the main raw material and contains abundant lactobacillus resources.In the process of kefir fermentation,a variety of small molecular compounds are produced.In order to explore the different source and main differential metabolites of bacteria in kefir from different sources,especially Lactic acid bacteria.In this study firstly collected 3 kefir from different regions(2 samples were from the folk family numbered K1 and K3;1 sample was from commercial,numbered K2).Then 16 S rDNA amplification sequencing technology was used to analyze the diversity of bacteria,especially Lactic acid bacteria in 3 kefir samples,and finally Liquid Chromatography-Mass Spectrometry(LC-MS)technology was used to analyze the differences of small molecular compounds in 3 kefir samples.The results were as follows:(1)The results of 16 S rDNA amplification technology showed that the bacteria and Lactic acid bacteria in the 3 kefir samples had a rich diversity,and the diversity of bacteria and Lactic acid bacteria in Kefir K1 were higher than those in K2 and K3.The bacteria belonged to 28 phyla and 374 classes.Firmicutes and Proteobacteria were the common dominant phyla in all 3 Kefir.All Lactic acid bacteria belong to 5 genera and 14 species,and the 5 genera included Lactobacillus,Lactococcus,Streptococcus,Leuconostoc and Gamella.The results also show that different kefir’s Lactic acid bacteria dominant bacteria groups were different,in which Lactobacillus was the dominant bacteria genus in Kefir K1 and K3;Lactococcus was the dominant bacteria genus in Kefir K2.Lactobacillus kefiranofaciens is the dominant Lactobacillus species in traditional Kefir K1 and K3;Lactococcus lactis is the dominant lactobacillus species in Kefir K2.(2)The metabolites in 3 Kefir samples were analyzed by LC-MS technique,and 419 and 209 metabolites were detected in the positive and negative ion modes.Subsequently,pair-to-pair comparison of metabolites in 3 Kefir samples were conducted,and major differential metabolites were screened according to VIP≥1,Folder change > 2and p<0.05.Key information was extracted from KEGG database and the functions of major differential metabolites were preliminarily analyzed.The pair-to-pair comparison results were as follows:(1)Kefir K1 compared with K2,14 different metabolites in Kefir K1 were higher than those in K2,and they belonged to 9 categories,including amino acids,vitamins,alkaloids,amines,aromatic compounds,nucleosides,lignans,fatty acids and fatty acylates.The KEGG pathway of major differential metabolites of Kefir K1 and K2 was analyzed,and it was speculated that Kefir K1 was superior to K2 in anti-inflammation,anti-tumor,hypoglycemic and other health functions as well as promoting the growth of bacterial communities.(2)Kefir K1 compared with K3,19 major differential metabolites in Kefir K1 were higher than that in K3,and they belonged to 7 categories,including amino acids,alkaloids,vitamins,sugars,organic heterocyclic compounds,fatty acylates and lignans.The KEGG pathway of major differential metabolites of Kefir K1 and K3 were analyzed,and it was speculated that Kefir K1 was superior to K3 in flavor formation,antibacterial,preservative and anti-cancer,which was consistent with the results that the diversity of bacteria and Lactic acid bacteria in Kefir K1 and the total number of Lactic acid bacteria were higher than that of K3.(3)Kefir K2 compared with K3,12 of the main differential metabolites in K2 were higher than that in K3,which belonged to 6 categories,including amino acids,alkaloids,sugars,organic acids,nucleosides and lignans.Through the KEGG pathway analysis of the major differential metabolites of Kefir K2 and K3,it was speculated that Kefir K2 was superior to K3 in maintaining intestinal homeostasis and systemic immunity,high microbial activity,and high yield of some compounds.The above results showed that different kefir had different bacterial composition and abundance(including Lactic acid bacteria),and small molecular compounds were also significantly different.Various indexes of kefir prepared with commercial kefir K2 grain were lower than traditional kefir K1,and better than traditional Kefir K3.At present,although the development of commercial Kefir starter had achieved certain results and scale,the composition of microbial flora was not as complex and diverse as that of traditional Kefir,which also leads to different composition and content of small and medium molecular substances in various Kefir products,and the quality of products was uneven.This study provides data support for the development of commercial kefir starter culture and the development and utilization of Lactic acid bacteria in Kefir.