Inhibition Of Portulaca Oleracea Fermentation Broth On Model Bacteria Of Acne And Preliminary Analysis Of Its Antibacterial Components

Acne is one of the common skin diseases.The main pathogenic bacteria of acne are Propionibacterium acnes(PA),Staphylococcus aureus(SA)and Staphylococcus epidermidis(SE).The inhibition model of acne was established by the inhibition of these three bacteria.Portulaca oleracea is a common medicinal and food homologous plant with many active ingredients.After probiotic fermentation,the active components and physiological functions of portulaca oleracea would change significantly,which has great development value.In this paper,the inhibitory effect of portulaca oleracea fermentation broth on model acne bacteria,optimization of fermentation conditions and analysis of antibacterial components of fermentation broth were studied.The main research contents are as follows:(1)Lactobacillus plantarum,Saccharomyces cerevisiae and Bacillus subtilis were cultured in Purslane extract for fermentation.Oxford cup method was used to conduct bacteriostatic experiments on the extract and three kinds of fermentation broth.The results showed that Lactobacillus plantarum fermented portulaca oleracea had the best inhibitory effect on PA,SA and SE,and the diameters of inhibition zones were 18.1±0.4 mm,20.4±1.2 mm and 12.5±0.6 mm,respectively.Bacillus subtilis fermentation broth only had strong antibacterial activity against SA,and the inhibition zone reached 24 mm.However,its inhibition ability to the other two acne model bacteria was slightly weaker,and the diameter of inhibition zone was significantly smaller than 4 mm.Saccharomyces cerevisiae fermentation broth showed no inhibition to the three acne model bacteria.Therefore,Lactobacillus plantarum was selected as the fermentation strain in the follow-up experiments.(2)Fermentation conditions such as solid-liquid ratio,inoculation amount,temperature and glucose addition were optimized by taking the diameter of inhibition zone of three acne model bacteria as the target.The results showed that 5% feed liquid had stronger inhibition on PA than fermentation broth,and the diameter of inhibition zone was 16.5±0.5 mm.The inhibition of SA and SE in 5% feed solution was slightly worse than that in fermentation solution.At 35℃,the inhibition of PA was the strongest,and the diameter of inhibition zone reached 19.8± 0.6mm.5% inoculation had the strongest inhibitory effect on PA,and the diameter of antibacterial circle was 13.9± 0.5mm.The inhibition effect of 3% glucose on PA was the strongest,and the diameter of inhibition zone was 13.2± 0.4mm.After optimization,the fermentation conditions were determined as 5% solid-liquid ratio,5% inoculation amount,3% glucose dosage and 35℃.Under the optimized conditions,the diameter of inhibition zone for PA and SA was 20 mm.The scavenging rate of hydroxyl radical and DPPH radical was increased from 17% to 33% and 44% to 72%.(3)Qualitative detection of antibacterial components in the extract,Lactobacillus plantarum seed and fermentation broth was carried out by high resolution mass spectrometry.Through database comparison,444 substances were detected.Most of them were organic acids,fatty acids,short peptides,terpenoids,phenols and so on.The contents of gluconic acid,phenylpropionic acid,fructozine,pantothenic acid,gullo-lactone,pentadecanoic acid and dodecanoic acid in fermentation broth were significantly increased compared with that in bacteria liquid(P-value <0.05).Compared with the extract,carnitine,AG957,N-(3,5-dimethylphenyl)-2-(4-hydroxyphenyl)acetamide,glucose acid,heptadecanoic acid,choline sulfate,alkaloids MQ-B2,pyridoxine,indole acrylic acid,betaine,colonic acid and Kalkitoxin were significantly increased(P-value <0.05).Fructozine,carnitine,AG957 and N-(3,5-dimethylphenyl)-2-(4-hydroxyphenyl)acetamide were selected from these substances which were significantly increased after fermentation to verify the inhibition of PA,SA and SE.The results showed that fructozine had certain inhibitory effect on three kinds of bacteria,while the other three substances showed no inhibitory effect.The results showed that fructozine was not detected in the extract and fermentation broth by liquid mass spectrometry.The content of fructozine in 48 h fermentation broth was 100 ug/L,and the diameter of the corresponding inhibition zone was 8 mm.(4)Shake flask fermentation conditions were applied to 5 L fermenter.The p H change in 5 L fermentor was the same as that in shaker.The p H of the fermentation broth decreased gradually and stabilize at 3.0-3.2.The antioxidant capacity of fermentation broth in 5 L fermentor was also improved.The scavenging rates of hydroxyl radical and DPPH radical in 5 L fermentor were increased from 17% to 26%and 47% to 64%.The fermentation broth of 5 L fermentor still had significant inhibitory effect on PA,SA and SE.The fermentation broth of 5 L fermentor had the strongest inhibition on PA,and the diameter of inhibition zone was about 19 mm.The inhibition of SA was slightly poor,and the diameter of antibacterial zone was 3-4 mm smaller than that in shaker.Fructozine content was determined by LC-MS,and the highest content in 60 h fermentation broth was 62 ug/L.In addition,guloglinolactone,2-hydroxy-4-methylpentanoic acid,phenylacetic acid,coumaric acid,2-ethylacetic acid,n-decanoic acid,pentadecanoic acid,aminophenyl formate and phenylpropionic acid were significantly increased in 5 L fermentor fermentation broth.