| Marine fouling organisms do great harm to offshore platforms and navigationequipment, severely affecting human economic and military activities at sea. Someresearchers find that the attachment of fouling microorganisms is a key link to thedamage of marine fouling, so it’s significant to study the antifoulant against them. Theuses of the available antifoulants have been gradually restricted due to their defectssuch as high toxicity and low degradability and so on, while the antifouling of naturalproduct has many advantages such as nonhazardous and environmentally friendly andso on, and has become a research hotpot. However, most of the active naturalproducts have a low yield with a high cost for preparation and a lack of stability. Soit’s urgent to search for the antimicrobial antifoulant with high production, low costand stabilityBacillus amyloliquefaciens can produce various kinds of active substances andhas a broad antibacterial spectrum. They have received more and more attention fortheir simple production process, high production and high activity and stability. In thisstudy, the results showed that B. amyloliquefaciens anti-CA strain had potent activityagainst Bacillus cereus, Vibrio anguillarum, Vibrio splendidus, Vibrio harveyi,Pseudoalteromonas and Pseudomonas aeruginosa which have been confirmed to befouling microorganisms.After TLC, column chromatography, HPLC and mass spectrometry were used toseparate and purify the active substances and analyze their structures from B.amyloliquefaciens anti-CA strain, the results showed that the active substances werecomposed of five chemical compounds whose molecular weight were994,1009,1022,1036and1050. We also analyzed the chemical structure of each compoundusing HPLC-MS and the tandem mass spectrum technique. After compared with the reference compounds available, these chemical compounds were found to be a seriesof homolog of cyclic lipopeptide with the difference of two adjacent homologs inmolecular weight being14. The main bioactive substance was confirmed to be acyclic lipopeptide containing a heptapeptide, the number of the carbon in two adjacenthomologs of lipid hydrocarbon chain differed by one and the fatty acid was a part ofthe cycle. The chemical compounds were named cyclic lipopeptide6-2belonging tothe class of surfactins, and their yield was0.6g/L.The methods of crystal violet staining and confocal sanning microscope (CLSM)were used to research the inhibition and scavenging effect of the cyclic lipopeptide6-2on the formation of fouling bacteria biofilms, and the result showed that it couldeffectively inhibit biofilm formation of bacterial strains of P. aeruginosa PAO1aswell as B. cereus. After the real–time quantitative PCR was used to analyze theexpression of the key gene PslC involved in polysaccharide biosynthesis in P.aeruginosa PAO1, the results showed that the cyclic lipopeptide6-2could decreasethe relative expression of the PslC gene.After the effects of the cyclic lipopeptide6-2on inhibition of kelp sporegermination and growth of barnacle larvae were also studied, the results showed thatthe cyclic lipopeptide6-2could effectively inhibit the germination of kelp spore andthe EC50was0.11μg/mL. Furthermore, it also had a certain toxicity against barnaclelarvae and the LC50was2.1μg/mL. After field test panel experiment, the resultsshowed that the test panel which containd12.0%of the cyclic lipopeptide6-2hadsome bioactivity against marine fouling organisms. |
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