Screening Of Lactic Acid Bacteria With Angiotensin-converting Enzyme Inhibitory Activity And Study On Fermentation Characters

Hypertension is a clinical syndrome which is characterized by arterial systolic blood pressure or diastolic blood pressure.It often leads to various complications such as heart,brain,kidneys,eyes and blood vessels diseases,which seriously threatens to the health of human.Angiotensin-converting enzyme(ACE)which is closely related to blood pressure has been considered to be a main cause of hypertension.The main function of ACE is to catalyze the conversion of angiotensin I into angiotensin Ⅱ and inactivates the vasodepressor compound bradykinin,thus makes the blood pressure increase.Angiotensin-converting enzyme inhibitors(ACEI)can restrain the conversion of between angiotensin Ⅰ and angiotensin Ⅱ,and prevent ACE hydrolyzing antihypertensive substances-bradykinin.Therefore,ACEI can reduce blood pressure.However,antihypertensive drugs may cause secondary actions,such as kidney damage,so food with ACEI activity attract people’s attention.Studies have found that,during the fermentation process,some precursor of the bioactive substance were released under the action of a variety of enzyme which were secreted by the lactic acid bacteria,including ACEI.Hypotensive activity of fermented dairy product is directly related to the species of lactic acid bacteria used in fermentation.In this study,31 strains of lactic acid bacteria with ACEI activity were evaluated.Two strains of Lactobacillus plantarum with high ACEI activity were screened through preliminary screening,further screening experiments and optimization of fermentative conditions.Moreover,a new method of two-step liquid-state mixed fermentation was used to fermenting skim milk,Bacillus subtilis SP8-5,Bacillus subtilis LC-2-1,Lb.plantarum YM-4-3,and Lb.plantarum AY01 with fermented skim milk,to explore the ACE inhibitory activity.The main results were as follows:Firstly,ACEI activity of 31 strains of lactic acid bacteria in pure culture of fermented milk were detected by UV spectrophotometer.Fifteen strains with relatively high ACEI activity were screened out.These strains were Lb.plantarum YM-4-3,Lb.plantarum YML-4-3,Lb.plantarum GBLT-3-1,Lb.plantarum QBLB-3-1,Lb.plantarum AY01,Lb.plantarum KY-6-3L,Lb.plantarum KY-6-2L,Lb.plantarum KY-1-2L,Lb.plantarum SP-8-7L,Pediococcus pentosaceus YY-1-3L,P.pentosaceus YY-1-7L,P.pentosaceus GJ-4-2L,P.pentosaceus GJ-4-10L,P.pentosaceus GJ-5-6L,and Weissella confusa JS4.The content of free amino acids and peptides of 31 strains’ fermented milk were also investigated by o-pHthalaldehyde method.It was found that the content of free amino acids and peptides showed relative with ACEI ratio of fermentation milk.Secondly,the acid and bile salt tolerance characteristics were assessed for potential probiotic properties,antibacterial properties test and sensitivity test were used in this study.Through those tests,Lb.plantarum YM-4-3,P.pentosaceus GJ-5-6L,P.pentosaceus YY-1-7L and Lb.plantarum AY01 with probiotic properties were screened out.High performance liquid chromatography(HPLC)method was also used to precisely determining the ACEI activity of 15 lactic acid bacteria strains.Results showed Lb.plantarum YM-4-3 and Lb.plantarum AY01 with high ACE inhibitory activity.ACEI ratio of Lb.plantarum YM-4-3 and Lb.plantarum AY01 were 80.8%and 78.2%,respectively.The relationship between ACEI activity and temperature,pH,viable cells of Lb.plantarum YM-4-3 and Lb.plantarum AY01 fermented milk were investigated.It was found that ACEI activity of both strains had a great relationship with fermentation temperature and the optimal fermentation temperature was 37℃.It was also found that the ACEI activity increased with the number of live bacteria cells increased,and with the pH value decreased.When fermentation time was 48 h,the ACEI activity was the highest,so it can be determined that the optimal fermentation time was 48 h.Finally,a two-step liquid-state mixed fermentation was used in this study.B.subtilis SP8-5 and B.subtilis LC-2-1 were used,because of their high-yield producing of proteinase.B.subtilis SP8-5,B.subtilis LC-2-1 were mixed with Lb.plantarum YM-4-3,Lb.plantaru AY01 fermented skim milk.It was found that the ACEI activity of fermented milk was higher than pure Lb.plantarum strain fermentation.And the ACEI ratio was more than 90%,which is 15%higher than that of pure Lb.plantarum strain fermentation.