Ezinye izihloko zokuxazulula izinkinga ze-LC aziphelelwa yisikhathi, njengoba kunezinkinga ekusebenzeni kwe-LC, njengoba ubuchwepheshe bezinsimbi buyathuthuka ngokuhamba kwesikhathi.Kunezindlela eziningi lapho izinkinga ezingavela khona ohlelweni lwe-LC futhi zigcine zisesimweni esibi kakhulu.
Kube mnandi ukubhala le kholomu ethi “LC Troubleshooting” kanye nokucabanga ngezihloko inyanga ngayinye, ngoba ezinye izihloko aziphumi esitayeleni.Ngenkathi emkhakheni we-chromatography icwaninga izihloko ezithile noma imibono iyaphelelwa yisikhathi njengoba ithathelwa indawo imibono emisha nengcono, emkhakheni wokuxazulula izinkinga, njengoba isihloko sokuqala sokuxazulula izinkinga savela kule jenali (i-LC ver 8 izihloko kusasele iminyaka engu-3 edlule) , ngigxile ezigabeni ezimbalwa zokuxazulula izinkinga ze-LC kumathrendi esimanje athinta i-liquid chromatography (LC) (isibonelo, ukuqhathanisa okuhlobene kokuqonda kwethu umthelela wengcindezi ekugcineni [2] Intuthuko Entsha) Ukuhumusha kwethu imiphumela ye-LC kanye nendlela yokuxazulula inkinga ngamathuluzi e-LC yesimanje. s of LC yokuxazulula izinkinga — izakhi ezinhle kunoma yisiphi isixazululi sezinkinga zibalulekile, kungakhathaliseki ubudala besistimu esiyisebenzisayo.Isihloko esiwumgogodla salolu chungechunge sihambisana kakhulu neshadi lodonga le-LCGC elidumile elithi “LC Troubleshooting Guide” (4) elilenga ezindaweni eziningi zokucwaninga.Engxenyeni yesithathu yalolu chungechunge, ngikhethe ukugxila ezindabeni ezihlobene nokubunjwa kodonga okungaba khona; t sicabangele zonke lezi zinkinga ngokuningiliziwe esihlokweni esisodwa, ngakho kulesi sitolimende sokuqala esihlokweni, ngizogxila kwezinye engizibona kaningi.Ngithemba ukuthi abasebenzisi be-LC abancane nabadala bazothola amathiphu nezikhumbuzi eziwusizo ngalesi sihloko esibalulekile.
Ngizithola ngiya ngokuya ngiphendula imibuzo yokuxazulula inkinga ngokuthi “kungenzeka noma yini”. Le mpendulo ingase ibonakale ilula uma ngicabangela ukuqaphela okunzima ukukuhumusha, kodwa ngikuthola kufaneleka kakhulu. Ngezizathu eziningi ezingaba khona zokuma okubi kakhulu, kubalulekile ukugcina umqondo ovulekile lapho ucabangela ukuthi inkinga ingaba yini, futhi sikwazi ukubeka phambili izimbangela ezingaba khona ukuze siqale imizamo yethu yokuxazulula izinkinga, ukugxila kulawo maphuzu abaluleke kakhulu, okungenzeka kakhulu.
Isinyathelo esibalulekile kunoma yimuphi umsebenzi wokuxazulula inkinga - kodwa engicabanga ukuthi sisezingeni eliphansi - ukuqaphela ukuthi kunenkinga okufanele ixazululwe. Ukubona ukuthi kunenkinga ngokuvamile kusho ukuqaphela ukuthi okwenzeka ethuluzini kuhlukile kulokho esikulindele, okubunjwa ngethiyori, ulwazi olunohlonze, nolwazi (5) .I-"peak shape" okubhekiselwa kuyo kuphela, i-supeal fluetric, i-symmetrically ebushelelezi kuphela lapha, i-supeal fluetric ffy, unqenqema oluphambili, umsila, njll.), kodwa futhi ububanzi.Okulindelekile kwethu kokuma kwesiqongo sangempela kulula.Ithiyori (6) isekela kahle okulindelekile kwencwadi yokufunda ukuthi, ezimweni eziningi, iziqongo zechromatographic kufanele zilingane futhi zivumelane nokuma kokusatshalaliswa kwe-Gaussian, njengoba kukhonjisiwe kuMfanekiso 1a.Yini esiyilindele ekuphakameni okukhulu kwesihloko esizoxoxa ngayo ngalesi sihloko esiyinkimbinkimbi nakakhulu. umdwebo 1 ukhombisa amanye amathuba angabonwa—ngamanye amazwi, ezinye zezindlela izinto ezingase zingahambi kahle.Engxenyeni esele yalesi sitolimende, sizochitha isikhathi sixoxa ngezibonelo ezithile eziqondile zezimo ezingaholela kulezi zinhlobo zomumo.
Kwesinye isikhathi iziqongo azibonwa nhlobo ku-chromatogram lapho okulindeleke ukuthi zikhishwe khona.Ishadi lodonga elingenhla libonisa ukuthi ukungabikho kwenani eliphakeme (ngokucabangela ukuthi isampula empeleni iqukethe i-analyte eqondiwe ekugxiliseni okufanele yenze impendulo yomtshina yanele ukuyibona ngaphezu komsindo) ngokuvamile ihlobene nenkinga yensimbi ethile noma izimo zesigaba seselula ezingalungile (uma zibonwa nhlobo).iziqongo, ngokuvamile kakhulu “zibuthakathaka”).Uhlu olufushane lwezinkinga ezingaba khona nezisombululo kulesi sigaba zingatholakala kuThebula I.
Njengoba kushiwo ngenhla, umbuzo wokuthi kungakanani ukunwetshwa kwesiqongo okufanele kubekezelelwe ngaphambi kokunaka nokuzama ukusilungisa kuyisihloko esiyinkimbinkimbi engizoxoxa ngaso esihlokweni esizayo.Okuhlangenwe nakho kwami ​​ukuthi ukunwetshwa okubalulekile kwesiqongo kuvame ukuhambisana noshintsho olubalulekile ekubunjweni kwepeak, futhi umsila we-peak uvame kakhulu kunokuqala noma ukuhlukaniswa.
Ngayinye yalezi zindaba ixoxwe ngokuningiliziwe ezindabeni zangaphambilini zokuxazulula izinkinga ze-LC, futhi abafundi abanentshisekelo kulezi zihloko bangabheka lezi zihloko zangaphambilini ukuze bathole ulwazi ngezimbangela kanye nezixazululo ezingaba khona kulezi zinkinga.Imininingwane eyengeziwe.
Umsila ophakeme, ukugqama okuphambili, nokuhlukanisa konke kungabangelwa izenzakalo zamakhemikhali noma ezibonakalayo, futhi uhlu lwezixazululo ezingaba khona kulezi zinkinga luyahlukahluka kakhulu, kuye ngokuthi sibhekene nenkinga yamakhemikhali noma engokomzimba. Ngokuvamile, ngokuqhathanisa iziqongo ezihlukene zechromatogram, ungathola izinkomba ezibalulekile mayelana nokuthi iyiphi imbangela. iziqongo ziyathinteka, kodwa ezinye zibukeka zizinhle, imbangela cishe ingamakhemikhali.
Izimbangela zamakhemikhali ze-peak tailing ziyinkimbinkimbi kakhulu ukuba zingaxoxa ngazo kafushane lapha.Umfundi onentshisekelo ubhekiselwa omagazini bakamuva "I-LC Troubleshooting" ukuze uthole ingxoxo ejulile (10). Nokho, into elula ukuyizama ukunciphisa isisindo sohlazi olujovwe futhi ubone ukuthi ukwakheka okuphezulu kuyathuthuka yini.Uma kunjalo, khona-ke lokhu kuyinkomba enhle yokuthi le nkinga yokulayisha ngokweqile, kufanele ibe yindlela encane yokulayisha "i-analyte" encane. noma izimo ze-chromatographic kufanele zishintshwe ukuze kutholwe amajamo aphezulu aphezulu noma ngabe kujovwe ubuningi obukhulu.
Kukhona futhi izizathu eziningi ezingaba khona ezithinta umsila omkhulu. Abafundi abanentshisekelo yengxoxo enemininingwane yamathuba badluliselwa kolunye udaba lwakamuva lwe-“LC Troubleshooting” (11).Enye yezimbangela ezivame kakhulu zomsila we-peak uxhumo olungeluhle endaweni ethile phakathi komjovo nomtshina (12).Isibonelo esidlulele sikhonjiswe kuMfanekiso 1d, esiwuthole emasontweni ambalwa edlule umjovo wami owenziwe nge-valve entsha. ngaphambili, futhi yafaka iluphu yomjovo wevolumu encane ene-ferrule eyayibunjwe ku-capillary yensimbi engagqwali.Ngemva kokuhlola okuthile kokuxazulula inkinga, saqaphela ukuthi ukujula kwembobo ku-stator ye-valve yomjovo kwakujule kakhulu kunalokho esasikujwayele, okubangele ivolumu enkulu efile ezansi kwembobo.Le nkinga ixazululwa kalula ngokufaka enye indawo ye-injection ye-injection ye-injection yembobo ukuze ulungise i-loop yembobo efile .
Iziqongo ezifana nalezo ezikhonjiswe kuMfanekiso 1e nazo zingabangelwa izinkinga ezingokwenyama noma zamakhemikhali. Imbangela evamile yomzimba yonqenqema oluholayo ukuthi umbhede wezinhlayiyana wekholomu awugcwele kahle, noma ukuthi izinhlayiya zihlele kabusha ngokuhamba kwesikhathi. Njengokungathi umsila ophakeme obangelwa yile nto ebonakalayo, indlela engcono kakhulu yokulungisa lokhu ukufaka esikhundleni sekholomu futhi siqhubeke nalokho okushiwo yikhemikhali okuholela ekuphakameni okuvame ukuvela, okuvame ukubizwa ngokuthi "i-pemental-pemental" ebizwa ngokuthi "imvelaphi" ngaphansi kwezimo ezikahle (eziqondile), inani le-analyte eligcinwe yisigaba esimile (ngakho-ke, isici sokugcina) lihlobene ngokuqondile nokugxilisana kwe-analyte kukholamu. Ngokwe-Chromatographically, lokhu kusho ukuthi njengoba inqwaba ye-analyte ejovwe kukholamu iyanda, inani eliphakeme liba lide, kodwa alibi banzi. Uyajova.Ngaphezu kwalokho, umumo ongewona umugqa unquma ukuma kweziqongo zechromatographic, okuholela emaphethelweni aholayo noma alandelanayo.Njengoba ngomthwalo omningi obangela umsila ophezulu (10), ukuhola okuphezulu okubangelwa ukugcinwa okungewona umugqa kungaxilongwa ngokunciphisa isisindo se-analyte esijovwe. ukuziphatha.
Kwesinye isikhathi sibona lokho okubonakala kuwukuqothuka “kokwehlukana”, njengoba kuboniswe kuMfanekiso 1f. Isinyathelo sokuqala ekuxazululeni le nkinga siwukuba sinqume ukuthi ukuma okuphezulu kubangelwa yini ukuhlangana okuyingxenye (okungukuthi, ukuba khona kwezinhlanganisela ezimbili ezihlukene kodwa ezisondelene kakhulu). Uma empeleni kukhona abahlaziyi ababili abahlukene abasondelana ndawonye, ​​kusho ukuthi kuyindaba yokuthuthukisa ipuleti, ukukhetha nokwandisa ukuxazulula, ukukhetha ipuleti, ipuleti le-applitation (noma ukukhetha) iziqongo zihlobene nokubonakalayo Ukusebenza akuhlangene nekholomu ngokwayo.Isikhathi esiningi, umkhondo obaluleke kakhulu kulesi sinqumo ukuthi zonke iziqongo zechromatogram zikhombisa ukuhlukana komumo, noma okukodwa noma okubili nje.Uma kukodwa noma kubili nje, cishe kuwudaba lokusebenzisana;uma zonke iziqongo zihlukaniswa, mhlawumbe inkinga engokomzimba, okungenzeka ukuthi ihlobene nekholomu ngokwayo.
Iziqongo zokuhlukaniswa ezihlobene nezici ezibonakalayo zekholomu ngokwayo ngokuvamile ngenxa yokuvinjelwa kancane kwe-inlet noma i-outlet frits, noma ukuhlelwa kabusha kwezinhlayiya zekholomu, okuvumela isigaba seselula ukuthi sigeleze ngokushesha kunesigaba seselula ezindaweni ezithile zokwakheka kwesiteshi sekholomu .kwezinye izifunda (11).I-frit evaleke kancane ngezinye izikhathi ingasulwa ngokuhlehlisa; ukugeleza kwekholomu;nokho, ngokuhlangenwe nakho kwami, lokhu kuvame ukuba yisixazululo sesikhashana kunesikhathi eside.Lokhu kuvame ukubulala ngamakholomu anamuhla uma izinhlayiya ziphinde zihlangane ngaphakathi kwekholomu.Kuleli qophelo, kungcono ukufaka esikhundleni sekholomu futhi uqhubeke.
Isilinganiso esiphezulu ku-Figure 1g, futhi kusukela kusenzakalo sakamuva elebhu yami, ngokuvamile kubonisa ukuthi isignali iphakeme kangangokuthi isifinyelele ekupheleni okuphezulu kwebanga lokuphendula.Kuma-optical absorbance detectors (i-UV-vis kulesi simo), lapho ukugxila kwe-analyte kuphezulu kakhulu, i-analyte idonsa iningi lokukhanya okudlula emtshinini wokugeleza kweseli, ishiya izimo ezincane kakhulu zokubona ukukhanya okuvela ngaphansi kwethonya likagesi. imithombo ehlukahlukene yomsindo, njengokukhanya okunhlanhlathayo kanye “nobumnyama bamanje”, okwenza isiginali “ifiphele” kakhulu ngokubukeka futhi izimele ekugxilweni kwe-analyte.Uma lokhu kwenzeka, inkinga ngokuvamile ingaxazululwa kalula ngokunciphisa umthamo womjovo we-analyte—ukunciphisa umthamo womjovo, ukuhlambulula isampula, noma kokubili.
Kwi-Chromatography School, sisebenzisa isinali ye-Detector (ie, i-y-axis ku-chromatograph) njengenkomba yokuhlaziya okuhlangenwe nakho kwe-zero.
Kulesi simo, ukuphakama okunegethivu kusho ukuthi ama-molecule aphumayo kukholomu amunca ukukhanya okuncane kunesigaba seselula ngokwaso ngokushesha ngaphambi nangemuva kwesiqongo.Lokhu kungenzeka, isibonelo, uma kusetshenziswa amaza amaza okubona aphansi (<230 nm) kanye nezithasiselo zesigaba seselula ezimunca iningi lokukhanya kulawa maza obude.Izithasiselo ezinjalo zingaba izingxenye zesigaba seselula ze-solvent noma izakhi ezifana ne-peak thayitha ye-peak noma i-peak thaw ukuze kulungiswe ijika lokulinganisa nokuthola ulwazi lobuningi olunembile, ngakho-ke asikho isizathu esibalulekile sokuzigwema ngesekhondi ngalinye (le ndlela ngezinye izikhathi ibizwa ngokuthi “ukutholwa kwe-UV engaqondile”) (13). Nokho, uma ngempela sifuna ukugwema ukuqothuka okungekuhle ngokuphelele, esimweni sokutholwa kokumunca, ikhambi elingcono kakhulu ukusebenzisa ukutholwa okuhlukile kwe-wavelength ukuze i-wavenge yokukhanya imunce kancane kunokushintsha kwesigaba seselula ukuze i-analyse imunce kakhulu kunokwakheka kwe-mobile tes.
Iziqongo ezingezinhle zingavela futhi uma kusetshenziswa ukutholwa kwe-refractive index (RI) uma inkomba ye-refractive yezingxenye ngaphandle kwe-analyte kusampula, njenge-solvent matrix, ihlukile kunkomba ye-refractive yesigaba seselula.Lokhu kwenzeka futhi ngokutholwa kwe-UV-vis, kodwa lo mphumela uvame ukuncishiswa ngokuhlobene nokutholwa kwe-RI. Kuzo zombili izimo, ukuphakama okunegethivu okufinyezwayo okucishe kube yisampula ye-matrix yokwakheka kweselula okuncane kakhulu.
Engxenyeni yesithathu esihlokweni esiyisisekelo sokuxazulula inkinga kwe-LC, ngixoxile ngezimo lapho umumo wesiqongo esibhekiwe uhluka khona esimeni esiphezulu esilindelekile noma esivamile.Ukuxazulula inkinga okuphumelelayo kwezinkinga ezinjalo kuqala ngolwazi lokwakheka okuphezulu okulindelekile (okusekelwe kumbono noma ulwazi lwangaphambilini ngezindlela ezikhona), ngakho ukuphambuka kulokhu okulindelekile kusobala.Izinkinga zomumo ophakeme zinezimbangela eziningi ezihlukene ezingaba khona, okuholela ekuxazululeni okuningi, njll. kaningi.Ukwazi le mininingwane kunikeza indawo enhle yokuqala ukuxazulula inkinga, kodwa akubambi konke okungenzeka.Abafundi abathanda uhlu olujulile lwezimbangela nezisombululo bangabhekisela eshadini lase-LCGC elithi “LC Troubleshooting Guide”.
(4) Ishadi lodonga le-LCGC “LC Troubleshooting Guide”.https://www.chromatographyonline.com/view/troubleshooting-wallchart (2021).
(6) A. Felinger, Ukuhlaziywa Kwedatha kanye Nokucubungula Kwezimpawu ku-Chromatography (Elsevier, New York, NY, 1998), amakhasi 43-96.
(8) Wahab MF, Dasgupta PK, Kadjo AF kanye no-Armstrong DW, Anal.Chim.Journal.Rev.907, 31–44 (2016).https://doi.org/10.1016/j.aca.2015.11.043.