The Detection Of Oenococcus Oeni Producing Biogenic Amines And Biogenic Amines In Red Wine

Biogenic amines are small nitrogenous molecules frequently found in fermented food.Excessive intakes of biogenic amines have undesirable physiological effect on human bodies.The decarboxylation reaction of the free amino acids generated biogenic amines. Based onconsideration about food safety, it is necessary to detect lactic acid bacteria producingbiogenic amines and biogenic amines qualitatively and quantitatively in wine.In this paper, PCR methods were used to amplify the genomic DNA of40Oenococcusoeni strains for detecting amino acid decarboxylase gene. The molecular biology methoddetected potential risk of biogenic amine formation in food. Then RP-HPLC methods wereused to analyze biogenic amines produced in amino acid decarboxylase medium and in redwine. The results provided a reliable method to evaluate the safety of Oenococcus oenistrains.1. The genes responsible for encoding the corresponding amino acid decarboxylases,including histidine decarboxylase, tyrosine decarboxylase and ornithine decarboxylase, weredetected through single and multiplex PCR assays in use of primers JV16HC/JV17HC,TD2/TD5and AODC1/16in40strains of Oenococcus oeni. The results showed that33strains possessed histidine decarboxylase gene,22strains possessed tyrosine decarboxylasegene and16strains possessed ornithine decarboxylase gene among the40tested strains. ThePCR methods described in this study are simple and rapid to detect bacteria with amine aciddecarboxylase genes.2. RP-HPLC methods were used to analyze biogenic amines produced by40Oenococcusoeni strains in amino acid decarboxylase medium. Biogenic amines were derived by dansylchloride before testing. The standard curves of tryptamine, putrescine, histamine and tyraminewere seted up and their linear regression equation is y=63791x+24132, y=70589x+24148,y=119166x+37018and y=134566x+54018, respectively. The correlation coefficientswere over0.9986and showed a good Linear correlation. The detection limits of tryptamine,putrescine, histamine and tyramine were0.15,0.1,0.2,0.2mg/L, respectively. The results ofRP-HPLC verified the accuracy of PCR methods.3. RP-HPLC methods were used to analyze biogenic amines in8red wine samples. Tryptamine, putrescine, histamine and tyramine were derived by dansyl chloride beforeanalysed by RP-HPLC with UV detection. The results show that the concentrations oftryptamine, putrescine, histamine and tyramine ranged from:6.54to33.44mg/L,0.9to290.71mg/L,0to8.83mg/L,0to7.91mg/L, respectively. None of the samples exceeded thetoxicity limit of50mg/L,100mg/L set for histamine, tyramine by FDA in wine, respectively.