I nt r ac e l l ul arox i da v edamageduet oan bi oc songutbac t e r i a r e duc e dbyan ox i dantmol e c ul eGM15 Dhr ubj yot iSha r ma
De p a r t me n t o f Bi o t e c h n o l o g y , S RM I n s t i t u t eo f S c i e n c ea n dTe c h n o l o g y , Ka t t a n k u l a t h u r , Ta mi l Na d u , I n d i a
RESULTS 1.Humangutc ons i s t sof>1000s pe c i e sofbac t e r i awhi c har er e qui r e df or s moot hf unc t i oni ngoft hehumangut . 2.Br oads pe c t r um ant i bi ot i c sar eknownt ohaves i dee f f e c t s .Whe nant i bi ot i c s ar ei nt r oduc e di nt ot hebody,i ti nduc e sROSl e adi ngt ooxi dat i ves t r e s s ,whi c h l e adst oki l l i ngoft hec e l l s . 3.I tbe c ome sdi f f i c ul tf ort henat ur al l ypr e s e ntant i oxi dants ys t e mi nt hebody t obal anc et hee f f e c tofoxi dat i ves t r e s st ot hec e l l s ,s oi ti sr e qui r e dt oi nt r oduc e ant i oxi danti nt ot hebodyar t i f i c i al l y. 4. Thec hangei nt hee qui l i br i um ofoxi dant sandant i oxi dant s ,i nf avoroft he oxi dant swhi c hl e adst odamageofc e l l s ,i sknownas‘ oxi dat i ves t r e s s ’[ 1] . 5.I nt hi ss t udy, ant i oxi dantpe pt i dei s ol at e df r om A. pl ant e s i swe r et r e at e dwi t h ant i bi ot i ct r e at e dc e l l s . andi twasobs e r ve dt hatt heGM15pe pt i dec ans c ave nge t hee f f e c tofROSont hec e l l s[ 2] .
Ant i bi ot i c:GentamycinandStreptomycin St r ai ns:Bacillussubtilis(MN726522)andBacilluscereus(MN726923) Pe pt i de:GM15(GGTCVIRGCVPKKLM)isolatedfrom spirulina,A.plantesis. FACSAnal ys i s:
1 .Mi dl o g a r i t hmi cpha s eBa c i l l uss p.wa st r e a t e dwi t ha nt i bi o t i c sa ndi nc uba t e dwi t h GM1 5f o r2ha t3 7 º C. Thec e l lmi x t ur ea ndH2 O2we r eus e da sapo s i t i v ec o nt r o l . 2 . 5µlo f2 ′ , 7 ′ di c hl o r o f l uo r e s c e i ndi a c e t a t e( DCFDA)we r emi x e dwi t he a c hs a mpl e . I ti s o neo ft hemo s twi de l yus e ddy ef o rme a s ur i ngt her e do xs t a t eo ft hec e l l . 3 .Re l e a s eo ff l uo r e s c e nc ec a nbeo bs e r v e dbyt hepr e s e nc eo fpe a ki nt heFACS.The pe a kswe r et he nc o mpa r e dbe t we e nt hepo s i t i v ec o nt r o l a ndt hes a mpl ec o nt a i ni ngba c t e r i aa nda nt i bi o t i c s[ 3 ] .
FACSAnal ys i s:
1. Whe nbac t e r i alc e l l swe r et r e at e dwi t hhydr oge npe r oxi deandant i bi ot i c s , ani nc r e as ei nf l ur e s c e nc ec anbeobs e r ve dwhi c hc anbei de nt i f i e dbyt he he i ghtofpe akandt her e gi oni nwhi c ht hepe akbe l ongs . 2.I n Fi g14,i ti sc l e ar l yvi s i bl et hatt hegr aphi ss hi f t e dt owar dst hel e f t s i de whi c hde not e st hes i gni f i c antr e duc t i on i nt heoxi dat i ves t r e s st ot he c e l l s . 3.Thusi tc l e ar l yi ndi c at e st hatGM15pe pt i des ubs t ant i al l yr e duc e dt he ROSl e ve loft hec e l l s .
Fi gur e14.FACSanal ys i sofe f f e c tofGM15pe pt i deonbac t e r i alc e l l s ( B.s ubt i l i sandB.c e r e us )t r e at e dwi t hGe nt amyc i nandSt r e pt omyc i n
Re s az ur i nDyeTe s t: 1.Thec hangei nc ol ourofRe s az ur i nwe r eobs e r ve d. 2.Thec hangei nc ol ouri ndi c at e st hepr e s e nc eofme t abol i cac t i vi t yoft he c e l l s . 3.Theme t abol i c al l yac t i vec e l l swe r eabl et or e duc et her e s az ur i nc hangi ngt hec ol ourf r om Bl uet opi nkwhe r e asunc hangeofc ol ouri ndi c at e st he pr e s e nc eofi nac t i vec e l l sorde adbac t e r i alc e l l s .
Re s az ur i nDyeTe s t:
1 . Thi sa s s a yhe l psi nt hes t udyo fba c t e r i a lme t a bo l i ca c t i v i t y . 2 . I ti sar e do xi ndi c a t o rwhi c ht ur nsi tc o l o urf r o m bl uet opi nkwhe nt r e a t e dwi t ha c t i v e ba c t e r i a lc e l l s .Co l o urc ha ng eo c c ur sduet or e duc t i o no fr e s a z ur i nt or e s o f ur i na ndf ur t he ri nt ohy dr o r e s o f ur i n[ 4 ] . 3 .Di f f e r e ntc o nc e nt r a t i o nso fGM1 5( 2 5μM,5 0μM a nd1 0 0μM)) we r eus e df o rt hi s a s s a ya ndwa spe r f o r me do n9 6we l lpl a t e .
Fi gur e5. Changeofc ol ourofRe s az ur i nf r om Bl uet oPi nki nt hepr e s e nc e ofGM15pe pt i dei ndi c at i ngt hepr e s e nc eofme t abol i c al l yac t i vec e l l s .
CONCLUSI ON: he ngt ngi ave c sbys s e r t ves i dat heoxi et duc e or et deGM15wasabl i pt dantpe oxi i heant hatt udet l onc anc wec , nt me i r xpe heabovee om t Fr he soft e i t r ope dantpr oxi i heant dt me r i onf aysc s aras ul c e Mol . s l l e c al i r e t dbac e at e r ct i ot bi i nant si s e r t ves i dat ooxi dduet e oduc spr dant oxi . s s e r t ves i dat toxi ns dagai e ugsbas onofdr i t oduc hepr dont e us oc obef dt e onne i nt e t eat ur ut arf nne ,i .Thus de i pt GM15pe
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