Njengesixhobo sokuchaneka okuphezulu, i-HPLC inokukhokelela ngokulula kwiingxaki ezincinci ezincinci ukuba ayisetyenziswanga ngendlela echanekileyo ngexesha lokusetyenziswa.Enye yeengxaki eziqhelekileyo yingxaki yoxinzelelo lwekholamu.Uyicombulula njani ngokukhawuleza ingxaki yechromatograph engalunganga.Inkqubo ye-HPLC iquka ibhotile yokugcina amanzi, impompo, i-injector, ikholamu, igumbi lobushushu lekholomu, i-detector kunye nenkqubo yokucubungula idatha.Kuyo yonke inkqubo, iintsika, iimpompo kunye nee-detectors zizinto eziphambili kunye neendawo eziphambili ezijongene neengxaki.
Isitshixo soxinzelelo lwekholomu yindawo efuna ingqalelo esondeleyo xa usebenzisa i-HPLC.Ukuzinza koxinzelelo lwekholomu kuhambelana ngokusondeleyo kwimilo ye-chromatographic peak, ukusebenza kakuhle kwekholomu, ukwahlula kakuhle kunye nexesha lokugcinwa.Uzinzo loxinzelelo lwekholomu aluthethi ukuba ixabiso loxinzelelo lizinzile kwixabiso elizinzile, kodwa kunoko ukuba uluhlu lokutshintsha koxinzelelo luphakathi kwe-345kPa okanye i-50PSI (ukuvumela ukusetyenziswa kwe-gradient elution xa uxinzelelo lwekholomu luzinzile kwaye lutshintsha ngokukhawuleza).Uxinzelelo oluphezulu okanye oluphantsi kakhulu yingxaki yoxinzelelo lwekholomu.
Eyona nto ithandwa kakhulu kwiintsilelo ze-HPLC kunye nezisombululo
I-1, uxinzelelo oluphezulu yeyona ngxaki ixhaphakileyo ekusebenziseni i-HPLC.Oku kuthetha ukunyuka ngokukhawuleza koxinzelelo.Ngokubanzi, kukho ezi zizathu zilandelayo: (1) Ngokuqhelekileyo, oku kungenxa yokuvaleka komjelo wokuhamba.Ngeli xesha, kufuneka sihlolisise ngokweenkcukacha.a.Okokuqala, cima indawo yokungena yempompo yokufunxa.Ngeli xesha, ityhubhu ye-PEEK yazaliswa ngolwelo ukwenzela ukuba ityhubhu ye-PEEK yayincinci kunebhotile ye-solvent ukuze ibone ukuba i-liquid iyancipha ngokuthanda.Ukuba ulwelo aluvuzi okanye luvuze kancinane, intloko yokucoca isinyibilikisi ivalekile.Unyango: Gxila kwi-30% ye-nitric acid kwisiqingatha seyure kwaye uhlambulule ngamanzi e-ultrapur.Ukuba ulwelo luthontsiza ngokungakhethiyo, intloko yokucoca isinyibilikisi iqhelekile kwaye iyajongwa;b.Vula i-valve ye-Purge ukwenzela ukuba isigaba seselula singadluli kwikholamu, kwaye ukuba uxinzelelo aluncitshiswanga kakhulu, intloko emhlophe yokucoca ivaliwe.Unyango: I-whiteheads ehlungiwe yasuswa kwaye i-sonicated nge-10% isopropanol kwisiqingatha seyure.Ukucinga ukuba uxinzelelo lwehla ngaphantsi kwe-100PSI, intloko emhlophe ehluziweyo iyinto eqhelekileyo kwaye ihlolwe;c.Susa ukuphela kokuphuma kwekholamu, ukuba uxinzelelo alunciphisi, ikholamu ivaliwe.Unyango: Ukuba sisithinteli setyuwa evalekileyo, hlambulula ama-95% de uxinzelelo luqhelekile.Ukuba isithintelo sibangelwa yinto ethile egcinwe kakhulu, ukuhamba okunamandla kunenqanaba elihambayo elikhoyo kufuneka kusetyenziswe ukuleqa kuxinzelelo oluqhelekileyo.Ukuba uxinzelelo lwexesha elide lokucoca alunciphisi ngokwendlela engentla, i-inlet kunye ne-outlet yekholomu inokuthathwa njengento edibeneyo nesixhobo ngokuchaseneyo, kwaye ikholamu inokucocwa kunye nesigaba seselula.Ngeli xesha, ukuba uxinzelelo lwentsika lungekancitshiswa, ipleyiti yokungena kwikholam inokutshintshwa kuphela, kodwa emva kokuba umsebenzi ungalunganga, kulula ukukhokelela ekunciphiseni umphumo wekholamu, ngoko zama ukusebenzisa ngaphantsi.Kwiingxaki ezinzima, ukutshintshwa kwekholomu kunokuqwalaselwa.
(2) Ukumiswa kwesantya sokuhamba okungalunganga: Isantya sokuhamba esichanekileyo sinokusetwa kwakhona.
(3) Umlinganiselo wokuhamba ngendlela engafanelekanga: isalathisi se-viscosity semilinganiselo eyahlukeneyo yokuhamba iyahluka, kwaye ingcinezelo yenkqubo ehambelanayo yokuhamba kunye ne-viscosity ephezulu nayo inkulu.Ukuba kunokwenzeka, i-solvents ye-viscosity ephantsi inokutshintshwa okanye iphinde isetyenziswe kwaye ilungiswe.
(4) Uxinzelelo lwenkqubo zero drift: lungelelanisa i-zero yenzwa yenqanaba lolwelo.
2, uxinzelelo luphantsi kakhulu (1) ludla ngokubangelwa kukuvuza kwenkqubo.Omawukwenze: Fumana unxibelelwano ngalunye, ngakumbi ujongano kuzo zombini iziphelo zekholamu, kwaye uqinise indawo evuzayo.Susa isithuba kwaye uqinise okanye ulandele ifilimu ye-PTFE ngamandla afanelekileyo.
(2) Igesi ingena kwimpompo, kodwa uxinzelelo luhlala lungazinzanga ngeli xesha, luphezulu kwaye luphantsi.Okubaluleke ngakumbi, impompo ayiyi kukwazi ukufunxa ulwelo.Indlela yonyango: vula i-valve yokucoca kwaye uhlambuluke kwinqanaba lokuhamba kwe-3 ~ 5ml / min.Ukuba akunjalo, amaqamza omoya ayenqwenelwa kwivalve yokukhupha kusetyenziswa ityhubhu yenaliti ezinikeleyo.
(3) Akukho ukuphuma kwesigaba esihambahambayo: khangela ukuba kukho isigaba esihambayo kwibhotile ye-reservoir, ukuba i-sink ifakwe kwisigaba esihambayo, nokuba ipompo iyasebenza.
(4) Ivalve yereferensi ayivalwanga: ivalve yereferensi ivaliwe emva kokuncipha.Ngokuqhelekileyo yehla ukuya kwi-0.1.~ 0.2mL/ min emva kokuvala ivalve yereferensi.
Isishwankathelo:
Kweli phepha, kuphela iingxaki eziqhelekileyo kwi-chromatography yolwelo zihlalutywa.Kakade ke, kwindlela esisebenza ngayo, siya kuhlangabezana nezinye iingxaki ezingakumbi.Ekuphatheni impazamo, kufuneka silandele le migaqo ilandelayo: tshintsha kuphela into enye ngexesha ukugqiba ubudlelwane phakathi kwe-hypothetical factor kunye nengxaki;Ngokubanzi, xa sitshintsha iinxalenye zokusombulula ingxaki, kufuneka sinikele ingqalelo ekubuyiseleni ezo ndawo zicoliweyo endaweni yokuthintela inkunkuma;Ukwenza umkhwa wokurekhoda olungileyo ngundoqo kwimpumelelo yokuphatha iimpazamo.Ukuqukumbela, xa usebenzisa i-HPLC, kubalulekile ukunikela ingqalelo kwisampulu yonyango lwangaphambili kunye nokusebenza ngokufanelekileyo kunye nokugcinwa kwezixhobo.
Ixesha lokuposa: Sep-18-2023