Inqaku
I-Methylene Blue neVithamini C: Kutheni Isisombululo Sinokungabi Nambala
Ufaka ipilisi yevithamini C kwiglasi yesisombululo se-methylene blue esiluhlaza okwesibhakabhaka, uxovule, uze ubukele umbala oluhlaza uphela. Emva komzuzu, ulwelo lukhangeleka njengamanzi aqhelekileyo. Kulula ukugqiba kwelokuba ivithamini C iwutshabalalisile okanye iwenzile awasebenza lo mbala.
Loo ndlela yokucinga ayichananga, kwaye loo mpazamo ibalulekile. Into oyibukeleyo lutshintsho lwemeko ye-oxidation olunokubuyiselwa umva, hayi ukutshatyalaliswa. Into enika umbala isekhona. Phantsi kweemeko ezifanelekileyo, umbala uyabuya.
Iimeko ezimbini zento enika umbala
I-methylene blue ineemeko ezimbini eziqondwa kakuhle. Uhlobo olwenziwe i-oxidation, oludla ngokubhalwa njenge-MB+, lunenkqubo yamakhonkco adityaniswe ngendlela ye-conjugation enabela kude, efunxa ukukhanya ngamandla kufutshane ne-664 nm. Oko kufunxa kukhanya kuko okwenza umbala oluhlaza okwesibhakabhaka. Uhlobo oluncitshisiweyo, i-leucomethylene blue (LMB), alusenalo elo bhanti linamandla lokufunxa ukukhanya okubonakalayo, ngoko isisombululo esixutywe namanzi amaninzi sikhangeleka singenambala.
Cinga ngento enika umbala njengendawo encinane yokugcina ii-electron enokutshajwa kwakhona. Ukuyitshaja ngee-electron neproton kuyawususa umbala wayo. Ukukhupha ezo electron kubuyisela umbala:
MB+ + H+ + 2e- ukuya kwi-LMB (engenambala)
Ivithamini C ibonelela ngee-electron zokuyitshaja. I-ascorbate yenziwa i-oxidation isiya kwi-dehydroascorbate ize inikele ngezilingani ezimbini zokunciphisa, ngoko kubalo olwenziwe lula, imole enye ye-ascorbate inciphisa malunga nemole enye ye-MB+. Kwizisombululo zokwenene zaselabhoratri, olo balo luntsonkothile ngakumbi kuba ioksijini enyibilikileyo ikhuphisana ngokufumana kwaezo electron. Yiyo loo nto kudla ngokufuneka i-ascorbate engaphezulu ukuze isisombululo sihlale singenambala.
Le nkqubo ayinyanzelekanga ukuba ibe linyathelo elinye elicocekileyo lee-electron ezimbini. Ii-radical neentlobo eziphakathi ezincitshiswe ngokuyinxenye ziyathatha inxaxheba, kwaye indlela echanekileyo iyatshintsha ngokwesinyibilikisi, ubumuncu, noxinaniso. Isiphumo sokugqibela siyafana noko. I-MB+ eluhlaza okwesibhakabhaka iba yi-LMB engenambala, kwaye ivithamini C iyasetyenziswa iphele kule nkqubo.
Kutheni ukushukumisa kubuyisela umbala oluhlaza okwesibhakabhaka
Uhlobo oluqhelekileyo lweli linge olwenziwa eklasini lunenxalenye yesibini. Yiyeke ingqayi encinane engenambala ingashukumi, ize ihlale ingenambala. Yishukumise, ubuye umbala oluhlaza okwesibhakabhaka. Linda, uze uphele kwakhona.
Into ebuyisela umbala yioksijini. I-O2 enyibilikileyo yenza i-oxidation kwi-LMB iyibuyisele kwi-MB+, ngoko ukushukumisa akuyenzi isebenze into enika umbala. Kuzalisa kwakhona ioksijini enyibilikileyo ngokuxuba umoya okwisithuba esingaphezu kolwelo ungene kulwelo, nto leyo ekhawulezisa inkqubo ebuyela umva. Logama kusekho i-ascorbate eseleyo, i-MB+ eyenziwe kwakhona iyancitshiswa kwakhona. Oku kuvelisa umjikelo oqhelekileyo: ukungashukumi kuzisa ukungabikho kombala, ukushukumisa kuzisa umbala oluhlaza okwesibhakabhaka, ukulinda kuzisa ukungabikho kombala kwakhona. Yakuphela i-ascorbate, ioksijini iyoyisa ngokusisigxina kwaye isisombululo sihlala siluhlaza okwesibhakabhaka.
Yiyo loo nto umboniso we-methylene blue nevithamini C uchaphazeleka yindlela esenziwe ngayo isitya. I-cuvette evaliweyo enesithuba esincinane somoya ngaphezu kolwelo ihlala ingenambala ixesha elide kunebeaker ebanzi. Ukufaka umoya ngamandla kunciphisa ixesha lokungabikho kombala. Nabani na onika ingxelo ngeli linge kufuneka ke achaze iimeko zeoksijini, hayi nje izixa zeerejenti.
Umboniso olawulwayo neemeko zawo
Umfanekiso weqweqwe weli nqaku ubonisa uthelekiso olujoliswe kulo: isampulu yolawulo ye-MB+ eluhlaza okwesibhakabhaka ecaleni kwesampulu ye-LMB encitshiswe nge-ascorbate, phantsi kokukhanya okufanayo. Ukuze isiphumo sikwazi ukuphindwa, gcina yonke into etshintsha isantya ikwimeko emiselweyo:
| Into eguquguqukayo | Imeko ecetyiswayo | Isizathu sokubaluleka kwayo |
|---|---|---|
| I-methylene blue | 10 ukuya kuma-90 micromolar kwisithinteli sotshintsho lwe-pH se-phosphate, pH 7 | Ihambelana noxinaniso olusetyenziswa kwizifundo ze-redox ezipapashiweyo kwaye igcina umlinganiselo wokufunxwa kokukhanya phakathi kwemida yokulinganisa |
| I-ascorbate | 1 mM sodium ascorbate, esandul’ ukulungiswa | Ibonelela ngezilingani zokunciphisa ezingaphezulu ukuze ukususwa kombala kugqitywe |
| Isithinteli sotshintsho lwe-pH ne-pH | 50 mM phosphate, pH 7.0, kubushushu begumbi | Imisela iintlobo zekhemikhali ze-ascorbate nozinzo lwe-LMB |
| Ukukhanya | Ukukhanya okungephi kwendawo, qaphela ixesha lokuchanabeka | Ukukhanya okuqaqambileyo kongeza iindlela ze-photoredox ne-peroxide |
| Ioksijini | Ingqayi enye izaliswe yioksijini yomoya enyibilikileyo, enye isuswe umoya ngenitrogen | Yahlula umjikelo we-redox ococekileyo kwi-oxidation ephindayo |
| Iintsimbi | Iirejenti zodidi lohlalutyo, kunye nesampulu enye yolawulo ene-chelator | Intsimbi ekhoyo ngezixa ezincinane ivula ikhemistri ye-Fenton (jonga ngezantsi) |
Fota zombini iingqayi ngamaxesha amiselweyo, umzekelo kwimizuzu eyi-0, 1, 5, ne-15, uze uzishukumise kwaye uzifote kwakhona. Ingqayi ekususwe kuyo umoya ngenitrogen kufanele ihlale ingenambala elona xesha lide. Ingqayi efakwe umoya kufanele ibuye ibe luhlaza okwesibhakabhaka ngokukhawuleza emva kokushukunyiswa. Ukuba isampulu ayiphindi ibe luhlaza okwesibhakabhaka nokuba iyashukunyiswa kwaye ifakwe umoya, krokrela ukonakala okungenakubuyiselwa umva kunokuncitshiswa nje.
Abenzi bemixube basebenzisa kwaale ndlela yokucinga kumlinganiselo omkhulu. Indlela i-methylene blue eziphatha ngayo kumxube ixhomekeke kuzo zonke iimeko ze-redox nezokwenziwa komxube. Uvavanyo lozinzo kufuneka lurekhode i-pH, ioksijini enyibilikileyo, isixa sokukhanya esifunyenweyo, neentsimbi ezikhoyo ngezixa ezincinane, kunye noko kubonwayo kumbala.
I-pH, ukukhanya, noxinaniso zityhalela kumacala ahlukeneyo
Abaqalayo badla ngokubuza ukuba yiasidi okanye sisiseko esikhawulezisa le nkqubo. Impendulo echanekileyo kukuba i-pH ichaphazela iinxalenye ezimbini ezahlukeneyo zomjikelo. Phantsi kweemeko zamanzi aneasidi, ukufumaneka kweeproton kunokukhawulezisa inyathelo lokuncitshiswa kwe-MB+ yi-ascorbate. Kodwa i-LMB ngokwayo iyaxhathisa ngakumbi kwi-oxidation ebangelwa ngumoya xa ikwiasidi, kwaye iphinda yenziwe i-oxidation ngokukhawuleza ngakumbi njengoko i-pH inyuka. Ngoko isampulu eneasidi inokuphulukana nombala ngokukhawuleza ize ihlale ingenambala, ngoxa isampulu enealkali iphulukana nombala ize ibuye ibe luhlaza okwesibhakabhaka ngokukhawuleza. Zombini ezi zinto zibonwayo zichanekile phantsi kweemeko zazo.
Ukukhanya kongeza enye indlela eyahlukileyo. I-methylene blue evuselelwe kukukhanya ifikelela kwimeko ye-triplet ehlala ixesha elide, apho idlulisa ii-electron kunye ne-ascorbate neoksijini. Le khemistri ibangelwa kukukhanya inokukhawulezisa ukususwa kombala, ibuyisele umbala, okanye ivelise iintlobo zeoksijini ezisebenzayo, ngokuxhomekeke kubude bamaza, inqanaba leoksijini, noxinaniso. Thatha ukukhanya njengento eguquguqukayo elingweni kwaye ukurekhode.
Uxin aniso lumisela ukuba leliphi icala eloyisayo. I-ascorbate eninzi ithetha ukususwa kombala ngokukhawuleza kunye nemijikelo emininzi yokushukumisa nokuphela kombala. Into enika umbala eninzi ifuna izilingani zokunciphisa ezingaphezulu ngokomlinganiselo ofanayo. Ioksijini enyibilikileyo eninzi ikhuthaza umbala oluhlaza okwesibhakabhaka. Kuxinaniso oluphezulu lwento enika umbala kuvela obunye ubuntsonkothe, kuba iimolekyuli ze-methylene blue ziyadibana zibe ngamaqela ezibini namaqela amakhulu aneembonakalo zokufunxa ukukhanya ezitshintshileyo. Ngoko ukuphinda kabini isixa sento enika umbala akusaphindi kabini indlela efanayo yokuziphatha.
Ngaba ivithamini C iyenza ingasebenzi i-methylene blue?
Lo ngumbuzo osisiseko sophando oluninzi nge-methylene blue nevithamini C, kwaye ufuna impendulo echanekileyo. Hayi, ukuphela kombala akuthethi ukuba into yenziwe ayasebenza. Inxalenye yemolekyuli enika umbala itshintshe imeko, kodwa undoqo we-phenothiazine nomsebenzi wayo we-redox zisekho kwisisombululo. Le nkqubo inokubuyiselwa umva ngumoya, kwaye uhlobo oluncitshisiweyo ngokwalo luyakwazi ukusabela: i-LMB ayinantlawulo yombane kwaye ithanda amafutha ngakumbi kune-MB+. Kuphando lweeseli, iwela iinwebu ngaphambi kokuba iphinde yenziwe i-oxidation ngaphakathi.
Uncwadi lophando lweeseli luyenza icace le nto. Kwiiseli ze-endothelium ezikhuliswe elabhoratri, i-methylene blue ekuxinaniso oluphantsi lwe-micromolar ikhuthaza umsebenzi wendlela ye-pentose phosphate ngoxa isenza i-oxidation kwi-NADPH ne-NADH. Xa ingaphezu malunga ne-5 micromolar, yonyusa iziphawuli zezinto ezenza i-oxidation ngaphakathi kweeseli xa inkxaso yeglucose ingekho. Iiseli ezifakwe i-ascorbate ziyenza i-oxidation ngokuthe ngcembe loo ascorbate xa zichanabeke kwinto enika umbala. Isixa se-ascorbate esifakiweyo sinciphisa kuphela inxalenye yesiginali yezinto ezenza i-oxidation, njengoko kuchaziwe kuphononongo lwe-redox yale nto inika umbala kwiiseli ze-endothelium. Ngamanye amazwi, i-ascorbate ithatha inxaxheba ekuhambeni kwee-electron kwi-redox. Ayiyicimi into enika umbala.
Ukukhanya kuyenza icace ngakumbi le ngxelo. Xa kukho ukukhanya, imixube ye-methylene blue, i-ascorbate, neoksijini isebenzisa ioksijini, iqokelela i-hydrogen peroxide, kwaye xa kukho intsimbi esebenza njengesikhawulezisi, ivelisa i-radical ye-hydroxyl ngekhemistri ye-Fenton, njengoko kubonisiwe ngamalinge okubamba ii-spin nawokulinganisa ukusetyenziswa kweoksijini. Ii-radical ze-hydroxyl zinokonakalisa iimolekyuli zebhayoloji ngendlela engenakubuyiselwa umva kwaye zonakalise nento enika umbala ngokwayo. Ngoko isampulu ephelelwe ngumbala ngokusisigxina phantsi kokukhanya nongcoliseko lwentsimbi ayibonisi ukuncitshiswa okucocekileyo. Isenokuba ibonisa ukutshatyalaliswa okubangelwa zezinye iinkqubo zekhemistri.
Izinto ezingangqinwa sisisombululo esingenambala
Izigqibo ezintathu eziqhelekileyo zidlulela ngaphaya kobungqina kwaye kufuneka ziphetshwe kwiingxelo zelabhoratri, kwiileyibhile, nakwizilayidi zokufundisa.
Okokuqala, ukungabi nambala akuthethi ukungabi nabungozi. I-LMB kunye neoksijini kunye neentsimbi ezikhoyo ngezixa ezincinane zinokuxhasa iindlela ze-peroxide nee-radical, ngakumbi phantsi kokukhanya. Ukuqinisekisa ukungabi nabungozi kufuna uphando lwetyhefu, hayi ukulinganiswa kombala.
Okwesibini, ukungabi nambala akuthethi ukususwa kongcoliseko. Into enika umbala kunye nezinye izinto ezisebenzisana nayo kwi-redox zisekhona. Izinto ezichithekileyo, ulwelo olulahlwayo, nezinto ezingcoliswe ngamabala azilungiswa ngokongeza i-ascorbate. Kuyafuneka ukuqinisekiswa ngohlalutyo, umba oxoxwa kwiindlela zovavanyo lweentsalela nokususwa kongcoliseko.
Okwesithathu, ukungabi nambala akubonisi indlela eluncedo yokusebenzisa ezi zinto zimbini kunye. Inkqubo eyenzeka kwityhubhu yovavanyo ayithethi nto malunga nokufunxwa emzimbeni, ukusasazeka kwizicubu, izixa ezisetyenziswayo, okanye ukhuseleko lokusebenzisana kwazo ebantwini. Nabani na ocinga ukudibanisa ezi zinto zimbini ngenjongo yezempilo kufuneka aqale afunde uncwadi lokusebenzisana kwazo nolokhuseleko, aze adibane nemithombo efanelekileyo kunokuba athathe izigqibo ezibanzi koko kubonwa kwingqayi encinane.
Ukutolikwa okuchanekileyo kuncinci kodwa kuluncedo ngakumbi. I-methylene blue kunye ne-ascorbate ngumzekelo olula womjikelo we-redox: ivithamini C inikela ngee-electron, into enika umbala iba ingenambala, ioksijini ithatha ezo electron kwakhona, umbala uyabuya, kwaye ukukhanya, i-pH, uxin aniso, nentsimbi zimisela isantya neemveliso ezisecaleni. Yakuba iqondakele loo ngcamango, umboniso uyayeka ukuba liqhinga uze uqale ukuba sisixhobo sokuhlola. Ukuba isampulu yakho iziphatha ngokwahlukileyo, hlola ukufikelela kweoksijini, isixa sokukhanya esifunyenweyo, i-pH, nongcoliseko lweentsimbi ngaphambi kokuthandabuza into enika umbala.