| Humic acids(HAs)are a class of polymer compounds rich in active groups such as phenolic hydroxyl groups and carboxyl groups,which have excellent functions in repairing intestinal damage and treating intestinal diseases.Unfortunately,the mechanism of action of HAs is unknown.Starting from the molecular structure and properties of HAs,this study explores the mechanism of action of HAs through the following work:(1)Bibliometric analysis was applied to systematically evaluate HAs applications in the environmental field.In-depth analysis of the publication characteristics of 9145 articles published from 1980 to 2020 revealed that the number of articles published on HAs increased rapidly,with China having the largest number of publications(3109),followed by the United States(1949)and Germany(679).Specifically,the Chinese Academy of Sciences had the largest number of publications(647)and was the center of the most productive institutional cooperation network.The research related to HAs mainly focuses on the environmental field,and the application and mechanism of HAs in the treatment of heavy metal machines and pollutants are important research topics in the environmental field.HAs could reduce the toxicity of a variety of organic pollutants or eliminate pollutants through photodegradation and adsorption,and the carboxyl and phenolic hydroxyl groups in the HAs molecule are the key groups for its function.Additionally,HAs might be also used to treat heavy metal pollutants.HAs could adsorb heavy metals,thereby reducing their mobility in the environment and toxicity.In addition,HAs are used as antidiarrheal drugs in the field of traditional medicine;Used as a feed additive in the breeding industry to promote livestock and poultry growth rate and health;In agriculture,HAs can improve crop growth and quality by improving soil and fertilizer utilization;It can also be used for carbon sequestration to adapt to climate change.However,bibliometric analysis shows that the molecular structure of HAs remains unclear,thus limiting the elucidating and further application of the mechanism.These findings highlight the interaction between HAs and the environment and provide key insights that not only support the further development of HAs in environmental applications,but also provide new ideas for the research and application of HAs in other fields.(2)At present,the methods for quantifying acid groups in HAs are complicated or difficult to detect,or the measurement results are inaccurate.Therefore,a method for the rapid and accurate determination of phenolic hydroxyl and carboxyl content in HAs,nonhydrous conductivity titration(NACT),was constructed.By varying the p H,titration solution concentration and nitrogen flow rate of the HAs sample,the optimal titration conditions were determined as follows: sample p H 4,titrant concentration of 0.05 mol/L,nitrogen flow rate of 80 m L/min.The phenolic hydroxyl content of parahydroxybenzoic acid determined by NACT was 758.82±111.76 cmol/kg,and the carboxyl content was744.44±51.11 cmol/kg.The theoretical contents of phenolic hydroxyl groups and carboxyl groups in parahydroxybenzoic acid were 723.96 cmol/kg,indicating that this method could accurately quantify the carboxyl and phenolic hydroxyl groups in parahydroxybenzoic acid.The phenolic hydroxyl and carboxyl content in HAs standards determined using NACT did not differ significantly from the traditional method(barium hydroxide method),indicating that NACT can quickly and accurately determine the phenolic hydroxyl and carboxyl content in HAs.In addition,29 HAs samples from 8provinces/autonomous-region across the country were collected,and the content of acidic groups in HAs was analyzed by NACT.(3)Aflatoxin B1(AFB1)is a secondary metabolite produced by Aspergillus flavus and Aspergillus parasitic species,which is currently recognized as the most toxic natural organic pollutant,with strong carcinogenicity and widely present in food.Through the above research,we know that HAs have the potential to adsorb organic pollutants,and the carboxyl and phenolic hydroxyl groups in the molecule are the key groups for their functioning.Therefore,S6 HAs samples with high acid group content were selected in this part of the work,and their effect on AFB1-challenged mice was studied in depth.The results showed that HAs reduced AFB1-induced hepatocyte swelling and inflammatory cell infiltration.HAs also restored levels of various enzymes in the liver disturbed by AFB1.HAs significantly reduced the oxidative stress and inflammatory response caused by AFB1 by enhancing the immune function of mice.HAs repair intestinal barrier function damaged by AFB1 by increasing small intestinal length and villi height.In addition,HAs reprogram the gut microbiome,increasing the relative abundance of Desulfovibrio,Odoribacter,and Alistipes.Both in vitro and in vivo tests have shown that HAs can effectively remove AFB1 by adsorption.In summary,HAs can mitigate AFB1-induced liver damage by enhancing intestinal barrier function,regulating intestinal microbiota and adsorption.(4)The gut microbiome is highly correlated with inflammatory bowel disease(IBD).A total of 3890 articles related to these two terms from 2000 to 2020 were extracted from the Web of Science Core Collection database to study the association from a bibliometric perspective.Articles on this topic have grown rapidly since 2008.The United States and Harvard University are the countries and institutions with the largest number of articles,respectively.Inflammatory Bowel Diseases is the most prolific journal,with 211 articles.The most influential journal in this field is Gut,with 13,359 citations.Co-cited analysis showed that the most interesting topics related to IBD were “gut microbiota”,“metagenomics”,“bacterial community”,“fecal microbiota transplantation”,“probiotics”and “colitis-associated colorectal cancer”.Keyword cluster and keyword burst analyses showed that “gut microbiota”,“metagenomics”,and “fecal microbiota transplantation”are currently the most researched topics in the field of IBD.The literature in this field is mainly distributed in the direction of intestinal microbiota,microbial metabolites and related host signaling pathways,and probiotic therapy has also appeared in the literature many times.This bibliometric analysis can guide future research and promote research in the field of gut microbiome and IBD.(5)This section explores the effects of HAs on E.coli and various intestinal probiotics.The results showed that HAs inhibited the growth of E.coli by destroying the E.coli cell membrane,while HAs promoted the growth and reproduction of Lactobacillus plantarum and shortened its lag period.Scanning electron microscopy and fluorescence staining showed that Lactobacillus plantarum adsorbed and formed HAs-Lactobacillus plantarum biofilm complexes on the surface of HAs.In order to explore the mechanism of complex formation,HAs and diatomaceous earth surface charges were determined.The results showed that the biofilm formation process was independent of the positive and negative surface charge of HAs.We further observed the film-forming effect of Lactobacillus plantarum on weathered coal and biochar,respectively.The results showed that Lactobacillus plantarum had the best film-forming effect on the surface of HAs,followed by weathered coal,and no film-forming phenomenon on the surface of activated carbon,which was positively correlated with the phenolic hydroxyl content in the three carriers.Therefore,we hypothesize that the hydrogen bond interaction force between phenolic hydroxyl groups in the carrier and carbonyl groups in biofilm extracellular proteins may be a key mechanism for biofilm formation.In addition,the results of filmforming phenomena on the surface of HAs by E.coli,Lactobacillus rhamnosus,Lactobacillus fermentum and Streptococcus thermophilus showed that except for E.coli,the other three bacteria formed biofilms on the surface of HAs.This study not only provides strong evidence for the conclusion that HAs use the gut microbiome as a vector for the treatment of intestinal diseases,but also provides a new solution to the problem of poor practical application of probiotic therapy. |
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