Bakan shanyewar haske na shuɗin methylene da ikon shanyewa na molar: bayanin tunani mai alamta yanayi

Bakan shuɗin methylene yana kama da abu mai sauƙi har sai ka yi ƙoƙarin sake amfani da wata lamba daga gare shi. Wani ɗakin gwaje-gwaje yana bayar da ƙididdigar kashe haske ta molar kusa da 82,000. Wani yana bayar da 67,000. Na uku yana bayar da sama da 100,000. Dukkansu ukun na iya zama bayanai ingantattu na ma'aunin da suka yi, amma babu ɗayansu da yake tabbatacciyar ƙima ta bai ɗaya ga wannan rini.

Ruɗanin yana fitowa ne daga haɗa ra'ayoyi biyu daban. Ma'aunin shanyewar haske sakamakon awo ne a kan wani takamaiman narkakke a cikin takamaiman sel. Ikon shanye haske na molar, wanda ake yawan rubutawa da epsilon, wata ƙima ce a cikin tsarin lissafin wannan narkakken. Idan shuɗin methylene ya sauya yanayin haɗuwar ƙwayoyinsa, tsarin lissafin yana sauyawa, don haka ƙimar ma tana sauyawa. Hanyar da ta fi amfani ita ce ɗaukar kowane ƙididdiga a matsayin ƙima mai bayanin yanayi: tsawon igiyar haske, sinadarin narkewa, pH ko buffer, zafin jiki, zangon taro, da tsawon hanyar haske su kasance tare da lambar.

Yankin monomer a ruwa mai ƙarancin taro

A narkakken ruwa mai ƙarancin taro, babban yankin shanyewar haske na cation ɗin shuɗin methylene yana kusa da 664 zuwa 665 nm, kuma ana yawan bayar da shi a tsakanin 664 zuwa 668 nm bisa bambancin na'urori da zaɓin daidaita ma'auni. Wannan yankin shi ne zaɓin da ya dace don ƙayyade adadi idan narkakken yana da ƙarancin taro da ya isa ya sa yawancin rinin ya kasance a matsayin monomer.

Ƙimomi uku daga wallafe-wallafen bincike suna nuna dalilin da ya sa yanayin gwaji ya fi muhimmanci fiye da babbar lambar da aka bayar. Masu binciken yumɓu waɗanda suka riƙe samfuran ruwa ƙasa da kusan 7 micromolar a cikin sel na 1 cm sun bayar da epsilon na kusan 82,000 M-1 cm-1 a 665 nm. Wannan ƙimar tana da cikakken bayani fiye da yadda aka saba, a matsayin gangaren daidaita ma'auni da aka samo daga gwaji ga samfuran ruwa masu ƙarancin taro sosai. Wani binciken daidaita ma'auni na asali a ruwan da aka cire ions da aka yi daga baya, wanda ya yi amfani da 2.34 zuwa 37.52 micromolar da hanyar haske ta 1 cm, ya bayar da kusan 67,100 M-1 cm-1 a 665 nm ga jimillar rini. Ana sa ran wannan ƙaramin sakamakon; ba saɓani ba ne, domin rabin sama na wannan zangon daidaita ma'auni ya riga ya ƙunshi dimer mai yawa. Ƙima ta uku, kusan 73,460 M-1 cm-1 a 664 nm, ta fito ne daga aikin da aka yi a pH 6.86 da 293 K, wanda ya rarraba bakan da aka gani zuwa ɓangarorin monomer da dimer, kamar yadda aka taƙaita a wani sabon nazari mai tantancewa kan ilimin haske na shuɗin methylene. Wannan lambar ɓangaren monomer ne da aka ware ta lissafi, ba gangaren kai tsaye na ma'aunin shanyewar haske da jimillar rini ba.

Abin da ya kamata a yi a aikace shi ne: zaɓi ƙididdigar da yanayinta ya dace da samfuran da kake da su, ko ka daidaita ma'auninka da kanka. Kada ka ɗauki 82,000 ka yi amfani da shi a gwajin buffer na phosphate mai 30 micromolar, kana zaton alaƙar madaidaiciyar layi tana nan.

Kafadar gajeren tsawon igiyar haske gauraye ce

Yayin da taro yake ƙaruwa, wani ɓangaren gefe mai kama da kafada kusa da 600 zuwa 615 nm yana ƙaruwa idan aka kwatanta shi da kololuwar 664 nm. Ayyukan zamani galibi suna sanya gudummawar dimer na nau'in H kusa da 605 zuwa 610 nm. Cikakken nazarin bakan shuɗin methylene a ruwa ya ware kololuwar dimer a 607 nm da kololuwar babban taron ƙwayoyi a 600 nm, tare da ɓangarorin resonance na monomer a 664 da 650 nm. Ana iya samun cikakken rubutunsa ta bayanin buɗaɗɗen damar karatu na PMID 33251415.

Bai kamata a yi wa wannan kafadar alama a matsayin ma'aunin shanyewar haske na dimer kaɗai ba. Tsarin vibronic na monomer ma yana ba da gudummawa a wannan yankin, don haka tsayin da aka auna haɗuwar gudummawa da dama ne. Wani daidaitawar lissafi ta gama-gari ya gano cewa tsarin monomer da dimer ya kasa aiki a duk zangon 1.1 micromolar zuwa 3.4 millimolar, amma ƙara tetramer ya ba da daidaitawa mafi kyau, tare da siginar tetramer da ake iya ganowa ko da ƙasa da 34 micromolar. Ƙarfin ɓangarorin da aka daidaita ya yi yawa: kusan 107,800 M-1 cm-1 a 664 nm ga babban ɓangaren monomer, kusan 101,400 ga dimer a 607 nm, da kusan 138,500 ga tetramer a 600 nm, a ruwan da aka cire ions wanda ba shi da buffer, tare da sodium chloride daga 0 zuwa 0.15 M da zafin jiki daga 282 zuwa 333 K. Waɗannan ɓangarori ne da tsarin lissafi ya ware, don haka bai kamata a musanya su da gangaren Beer Lambert da aka samu daga gwaji ba.

An auna ƙarfin samuwar dimer kai tsaye. A pH 6.86 da 293 K, wani bincike ya samu tabbatacciyar ƙimar samuwar dimer kusa da 3,900 M-1. Wannan girman yana bayyana dalilin da ya sa narkakken aiki mai 5 micromolar yake nuna alaƙar kusan madaidaiciyar layi, yayin da narkakken asali mai 50 micromolar ba ya yin haka. Wani bincike mai amfani yayin haɓaka hanyar gwaji shi ne rabon ma'aunin shanyewar haske kusa da 610 nm da ma'aunin shanyewar haske kusa da 665 nm. Idan wannan rabon yana ƙaruwa tare da taro, epsilon a 664 nm ba ya sake bayyana nau'in ƙwaya guda da ba ya sauyawa.

A wani zangon taro da nau'o'in ƙwayoyi biyu masu sauyawa zuwa juna suka mamaye, bakuna na iya haɗuwa a wani wurin isosbestic, wanda aka gani a gwaje-gwajen jerin narkakken ruwa kusa da 625 nm. Wuri bayyananne da bai sauya ba ya dace da kasancewar manyan nau'o'in ƙwayoyi biyu. Idan wurin yana matsawa ko ya ɓace yayin da taro yake ƙaruwa, ɗauke shi a matsayin gargaɗi cewa manyan tarukan ƙwayoyi ko sauyawar daidaiton sinadarai sun shiga, ba kuskuren na'ura ba.

Sinadarin narkewa, pH, da zafin jiki suna matsar da bakan

Ruwa yana ƙarfafa haɗuwar ƙwayoyi. Ƙara dimethyl sulfoxide yana rage samuwar dimer, yana matsar da kololuwar shanyewar haske da fluorescence zuwa tsawon igiyar haske mafi tsawo, kuma yana ƙara yawan fitowar fluorescence ta quantum da kusan ninki huɗu, bisa ga binciken sinadarin narkewa na shuɗin methylene a ruwa da gaurayen DMSO, wanda aka sanya a kundin bayanai a matsayin PMID 30933781. Wannan aikin yana ɗaukar bakunan ruwa a matsayin daidaiton monomer da dimer, tare da ma'aunin shanyewar haske na monomer kusa da 650 zuwa 670 nm da na dimer kusa da 580 zuwa 615 nm, kuma yana samo tabbatattun ƙimomin samuwar dimer daga bakunan a 293.15 K.

Alcohols suna sauya matsayi da ƙarfin da ake gani, amma ba sa kawar da taruwar ƙwayoyi. A wani kwatancen bincike na asali a 31.3 micromolar da zafin ɗaki, wanda aka bayar a wani bincike kan sinadarin narkewa na shuɗin methylene, kololuwar dogon tsawon igiyar haske ta kasance 665 nm a ruwa amma 655 nm a methanol, tare da ƙimomin da ake gani na jimillar narkakken kusa da 37,100 a ruwa da 60,800 a methanol. Wani binciken tabbatarwa daban a ethanol marar ruwa a 298 K ya bayar da kololuwa a 655 nm tare da epsilon kusa da 68,000.

Sauye-sauyen pH na yau da kullum suna matsar da bakan ƙasa da yadda hanyoyin gwaji da yawa suke tsammani. Nazarin tantancewa da aka ambata a sama ya bayar da cewa daga kusan pH 2 zuwa pH 12, kololuwar ta kasance kusa da 665 nm, tare da ƙananan sauye-sauyen ƙarfi kawai har zuwa pH 11. Nau'in buffer da ƙarfin ions na iya sauya sakamako gwargwadon ƙimar pH da aka ambata, don haka a rubuta ainihin buffer da gishirin da aka yi amfani da su, ba karatun na'urar pH kaɗai ba. Acid mai ƙarfi ko alkali mai ƙarfi yanayi ne daban, domin rinin na iya sauyawa ta fuskar sinadarai maimakon haɗuwar ƙwayoyi da za ta iya komawa baya. Zafin jiki da gishiri su ma suna sauya daidaiton, shi ya sa binciken ruwa na 2020 ya sauya dukkansu da gangan.

Yin awo ta yadda lambar za ta ci gaba da inganci

Aikin ƙayyade adadi yana dogara da alaƙar Beer Lambert: ma'aunin shanyewar haske yana daidai da epsilon sau taro sau tsawon hanyar haske, tare da hujjar cewa zangon alaƙar madaidaiciyar layi yana aiki da gaske a yanayin da kake amfani da shi.

Yi amfani da sel na 1 cm wajen aiki da ƙaramin taro na micromolar. Idan epsilon yana kusa da 82,000, 5 micromolar yana ba da ma'aunin shanyewar haske kusa da 0.41, kuma 10 micromolar yana ba da kusan 0.82. Don awun da aka mayar da hankali kan monomer a ruwa kaɗai, kasancewa kusa da ko ƙasa da kusan 5 zuwa 7 micromolar ya fi aminci fiye da daidaita ma'auni har zuwa taron da ya kai gomomin micromolar. Ga narkakkun asali masu babban taro, rage tsawon hanyar haske maimakon amincewa da ma'aunin shanyewar haske da ya zarce 1 sosai.

Yi awun blank da gauraye iri ɗaya: ruwa mai matakin inganci iri ɗaya, kaso iri ɗaya na ƙarin sinadarin narkewa, buffer da gishirai iri ɗaya, amma babu rini. Shirya narkakkun daidaita ma'auni a cikin wannan gaurayen iri ɗaya, a zafin jiki iri ɗaya. A lokacin haɓaka hanyar gwaji, bincika duk zangon hasken da ake iya gani, mafi dacewa kusan 550 zuwa 750 nm, maimakon karanta tsawon igiyar haske guda. Bibiyi siffa, ba tsayi kaɗai ba. Idan jerin rage taro yana sauya siffar da aka daidaita zuwa ma'auni ɗaya, tsarin yana dogara da taro ta fuskar sinadarai. Kololuwa mai ɗorewa da rabon 610 zuwa 665 nm mai ɗorewa sun fi babban R squared kaɗai zama hujjar zangon da ya dace.

Rubuta nau'in sinadarin gwaji. Kayan da ake sayarwa na iya kasancewa methylthioninium chloride marar ruwan kristal ko hydrate kamar trihydrate, don haka 질ar molar da ake amfani da ita wajen lissafin narkakken asali dole ta dace da abin da yake cikin kwalbar a zahiri. Wannan mataki shi kaɗai yana bayyana wani ɓangare na saɓanin ƙimomin taron molar da aka ambata tsakanin ɗakunan gwaje-gwaje. Irin wannan tsantseni yana da muhimmanci idan ana amfani da sakamako a lissafin gwajin ƙayyade adadi, idan bakuna suna taimakawa wajen duba kwanciyar haɗin sinadarai, da kuma idan bayanan ƙazantu suna buƙatar nazarin tsabta da ƙayyadaddun halaye.

Taƙaitawa mai alamta yanayin gwaji

Yi amfani da wannan tebur a matsayin samfurin farawa don rubutun ɗakin gwaje-gwaje, ba a matsayin tushen tabbatattun ƙimomi da za a iya musanyawa ba.

Saboda haka, tsawon igiyar shanyewar haske na shuɗin methylene amsa ce gajera mai dogon ƙarin bayani. Bayar da kololuwar da ka auna, a cikin narkakken da ka auna, da taron da ka auna. Wannan ɗabi'ar tana hana wata ƙima ta narkakken ruwa mai ƙarancin taro kusa da 664 nm zama tabbatacciyar ƙima ta bai ɗaya ta ƙarya.