Purification Of Mutacin

The bacteriocins produced by the Mutans streptococci(MS),were designated mutacins which could kill or inhibit the growth of the closely related strains.As bacterial antibiotic resistance was on the rise,it became urgent to find new and better antimicrobial agents.Mutacins have extended-spectrum antimicrobial and low minimum inhibitory concentrations(MIC),and effective to multidrug-resistant bacterial strains(such as penicillin).So mutacin may be employed as a new source of antibiotics,which has an enormous potential prospect in the future.Numerous studies about mutacin have gradually become popular in the field of oral microbiology,and recent years some new mutacins were found constantly.In this study,to obtain the greatest active mutacin produced by Mutans streptococci(MS) isolates,we compared the mutacin activity of mutans streptococci isolates from caries-free and caries-active children against Streptococcus oralis ATCC 10557.Then we crude extracted and purified this selected mutacin by chloroform,SPE and RP-HPLC method.This study could contribute to exploratory development about mutacin.Methods 80 isolated strains were selected and identified from the dental plaque of 10 caries-active(dmft≥8) and 10 caries-free(dmft=0) children(3~5 years old), of which the antibacteria activity were tested by the stab culture technique against Streptococcus oralis ATCC 10557.The greatest active mutacin produced by"1G"from these MS isolates,was crude extracted by chloroform and initial purified by SPE(Solid-phase extraction).And then the tested active substances were purified by twice RP-HPLC.The second RP-HPLC was purified under the same eluent system as the first RP-HPLC(Reversed-phase high performance liquid chromatography).Then eluents of target peptide were collected and freezedried for further study.Results 1.The biggest inhibition zone size of caries-activity group was 21mm,the smallest one was 6mm, the average size was 10.4mm,while the biggest inhibition zone size of caries-free group was 28mm, the smallest one was 0mm, the average size was 6.9mm. The diffrerence between the two groups was statistically siginificant (t=3.098,P<0.05).2. The strain"1G"was the greatest active one of these mutans streptococci isolates,of which the average inhibition zone size was 20mm.Roughly 15ug crude mutacin was extracted from 200ml liquid medium of this strain 1G.Then we obtained the purified mutacin through SPE and twice RP-HPLC.Conclusion The production of mutacin has extensiveness in the MS isolates (88% MS isolates produce inhibition zone against ATCC 10557 in this study),and has differences between MS strains and isolates.The molecular mass of mutacin was very small,so it is much complex that mutacin was separated and purified.We mastered basic extracting and initial purifying methods of mutacin,which may make an important contribution to the further research about mutacin.