Study On The Antibacterial Activity And Mechanism Of Alkaloids From Konjac Fly Flour

Konjac is a traditional Chinese medicine and food plant with various biological activities such as blood sugar lowering,blood fat lowering and anti-obesity.Konjac produces a lot of fly powder when it produces konjac flour(glucomannan).A lot of researches focus on how to use glucomannan in konjac flour,and rarely pay attention to the development and utilization of konjac fly powder.Our research found that the ethanol extract of konjac fly powder has a significant antibacterial activity.Further studies have found that this antibacterial activity is mainly due to the influence of alkaloids in konjac fly powder.In order to make full use of konjac powder,this study evaluated the extraction and purification conditions,antimicrobial activity and mechanism of Alkaloid from konjac powder(AKP).The main results are as follows:Ultrasonic-assisted methodology was used to extract the alkaloids from konjac powder.Based on the results of four single factor tests,including extraction temperature,ratio of material-to-liquid(g/mL),extraction time,and ultrasonic power.At this time,the yield of Alkaloid from konjac powder reached 1.986 mg/g,which was 18.9%higher than that before optimization.The extract of Alkaloid from konjac powder was purified with macroporous resin.Screening and comparing the adsorption and desorption capacities of five macroporous resins,we finally choosed NKA-II resin and optimize the purification conditions of its alkaloid extract.NKA-II reached adsorption and desorption equilibrium at 7 h and 9 h in static adsorption and desorption experiments,respectively.In the dynamic adsorption equilibrium experiment,the purification conditions of AKP were:the concentration of alkaloid was 2.56 mg/mL,the sampling rate was 4.0 BV/h,loading volume was 7 BV,and the elution concentration were 75%ethanol.Under this condition,the purity of Konjac fly alkaloid increased from the previous 44.85±0.57%to 71.78±0.52%,which was 1.6 times higher than that before purification.Determined the Minimum Inhibitory Concentration(MIC)of Escherichia coli CICC 10899,Staphylococcus aureus CICC 23656,Bacillus subtilis CICC 10732 and Salmonella typhimurium CICC 21483 by liquid ratio dilution method,AKP has obvious antimicrobial activity on four tested bacteria and the susceptibility tests of these four bacteria were studied by the Oxford Cup method.It was confirmed by the determination of the number of bacteria in the bacteria that the AKP can inhibit the growth of the colonies of the test bacteria.To investigate the antibacterial activity and mechanism of Escherichia coli CICC 10899 and Staphylococcus aureus CICC 23656 of alkaloid from konjac powderm,the growth curve was determined and the cell morphology and structure were observed.The conductivity of the culture medium changed significantly after treated with AKP.The tested bacterial has no full cell morphology under the scanning electron microscopy.And the contents of the test bacteria leaked under transmission electron microscope.After staining with Syto 9/PI,the tested bacterial detected red signal of membrane damage under laser confocal microscopy,determined the mechanism of AKP inhibit Escherichia coli CICC 10899 and Staphylococcus aureus CICC 23656 was that the integrity of the cell membrane was broken to achieve the bacteriostatic effect.