Hongcheng Wang, Bo Xu
Codsiga Xarunta R&D
Hordhac
Marka loo eego jaangooyooyinka qaraabada ah ee marxaladda taagan iyo marxaladda wareegta, chromatography dareeraha waxaa loo qaybin karaa marxaladda caadiga ah ee chromatography (NPC) iyo chromatography wejiga rogan (RPC).RPC, kala-soocida wajiga moobilka ayaa ka xoog badan kan wajiga taagan.RPC waxay noqotay midka ugu badan ee loo isticmaalo hababka kala soocida chromatography dareeraha ah sababtoo ah waxtarkeeda sare, xallinta wanaagsan iyo habka haynta cad.Sidaa darteed RPC waxay ku habboon tahay kala-soocidda iyo nadiifinta xeryahooda kala duwan ee polar ama non-polar, oo ay ku jiraan alkaloids, carbohydrates, acids fatty, steroids, acids nucleic, amino acids, peptides, borotiinada, iwm. In RPC, marxaladda ugu badan ee la isticmaalo. matrix-ka silica jel kaas oo ku xidhan kooxo kala duwan oo shaqaynaya, oo ay ku jiraan C18, C8, C4, phenyl, cyano, amino, iwmWaxaa lagu qiyaasaa in in ka badan 80% ee RPC ay hadda isticmaalayaan wejiga isku xidhan ee C18.Sidaa darteed tiirka chromatography C18 wuxuu noqday tiir caalami ah oo laga rabo shaybaar kasta.
Inkasta oo tiirarka C18 loo isticmaali karo codsiyo aad u ballaaran, si kastaba ha ahaatee, shaybaarada qaarkood kuwaas oo ah polar ama aad u sarreeya, tiirarka caadiga ah ee C18 waxay yeelan karaan dhibaatooyin marka loo isticmaalo si loo nadiifiyo shaybaaradaas.Gudaha RPC, dareerayaasha caadiga ah ee la isticmaalo ayaa lagu dalban karaa iyadoo loo eegayo polaritykooda: biyaha
Jaantuska 1. Jaantuska jaantuska ee wejiyada isku xidhan ee dusha sare ee jelka silica ee tiirka C18 ee caadiga ah (bidix) iyo C18AQ (midig).
Si wax looga qabto dhibaatooyinka kor ku xusan, soosaarayaasha qalabka wax lagu xidho chromatographic ayaa sameeyay horumarin farsamo.Mid ka mid ah hagaajintaasi waxay samaynaysaa qaar ka mid ah isbeddelada dusha sare ee silica matrix, sida soo bandhigida kooxaha cyano hydrophilic (sida ku cad qaybta saxda ah ee Jaantuska 1), si loo sameeyo dusha sare ee silica jel ka badan.Sidaa darteed silsiladaha C18 ee dusha sare ee silica ayaa si buuxda loogu kordhin karaa xaaladaha aqueous aadka u sarreeya iyo burburka wejiga hydrophobic waa laga fogaan karaa.Tiirarkan C18 ee wax laga beddelay waxaa loo yaqaan tiirarka C18 aqueous, kuwaas oo ah tiirarka C18AQ, kuwaas oo loogu talagalay xaaladaha biyo-biyoodka aadka u sarreeya waxayna u dulqaadan karaan 100% nidaamka aqueous.Tiirarka C18AQ ayaa si weyn loogu dabaqay kala-soocidda iyo nadiifinta xeryahooda polar ee xooggan, oo ay ku jiraan asiidhyada organic, peptides, nucleosides iyo fiitamiinnada biyaha-milmi kara.
Desalting waa mid ka mid ah codsiyada caadiga ah ee tiirarka C18AQ ee nadiifinta tooska ah ee shaybaarada, kaas oo meesha ka saaraya milixda ama qaybaha kaydka ee dareeraha muunada si loo fududeeyo codsiga muunada ee daraasadaha xiga.Boostada dhexdeeda, FCF-da buluugga ah ee buluuga ah ee leh polarity xooggan ayaa loo adeegsaday muunada oo lagu safeeyey tiirka C18AQ.Muunada dareeraha ah waxaa lagu beddelay dareere organic ah oo ka soo baxay xalka kaydinta, sidaas darteed fududaynta uumiga wareegga soo socda iyo sidoo kale badbaadinta dareeraha iyo wakhtiga shaqada.Intaa waxaa dheer, nadiifinta shaybaarka ayaa la wanaajiyay iyadoo laga saaray xoogaa wasakh ah muunada.
Qaybta Tijaabada
Jaantuska 2. Qaab dhismeedka kiimikada ee muunada.
FCF-ga buluuga ah ee quruxda badan ayaa loo adeegsaday muunad ahaan boostadan.Nadiifinta muunada cayriin waxay ahayd 86% qaabka kiimikaad ee muunada ayaa lagu muujiyay Jaantuska 2. Si loo diyaariyo xalka muunada, 300 mg budada cayriin ee adag ee Brilliant Blue FCF ayaa lagu milmay 1 M NaH2PO4 xal baraf ah oo si fiican u ruxay si uu u noqdo xal dhamaystiran oo cad.Xalka muunada ayaa markaa lagu duray tiirka flashka iyadoo la durayo.Habaynta tijaabada ah ee nadiifinta tooshku waxay ku taxan tahay shaxda 1.
| Qalabka | SepaBean™ Machine2 | |||
| Kartridges | 12 g SepaFlash C18 RP flash cartridge (silica wareegsan, 20 - 45 μm, 100 Å, Lambarka dalbashada: SW-5222-012-SP) | 12 g SepaFlash C18AQ RP flash cartridge (silica wareegsan, 20 - 45 μm, 100 Å, Lambarka dalbashada: SW-5222-012-SP(AQ))) | ||
| Mawjada dhererka | 254 nm | |||
| Marxaladda mobilada | Walxaha A: Biyaha Miraha B: Methanol | |||
| Heerka socodka | 30 ml/daqiiqo | |||
| Muunad rarista | 300 mg (FCF buluug ah oo dhalaalaya oo nadiif ah 86%) | |||
| Gradient | Waqtiga (CV) | Miraha B (%) | Waqtiga (CV) | Miraha B (%) |
| 0 | 10 | 0 | 0 | |
| 10 | 10 | 10 | 0 | |
| 10.1 | 100 | 10.1 | 100 | |
| 17.5 | 100 | 17.5 | 100 | |
| 17.6 | 10 | 17.6 | 0 | |
| 22.6 | 10 | 22.6 | 0 | |
Natiijooyinka iyo dood
A SepaFlash C18AQ RP flash kartridge ayaa loo isticmaalay muunad nadiifinta iyo nadiifinta.Talaabada jaranjarada ayaa la adeegsaday kaas oo biyo saafi ah loo isticmaalay sidii wejiga wareega ee bilawga elution oo loo ordayay 10 tiir mug (CV).Sida ku cad Jaantuska 3, marka loo isticmaalayo biyaha saafiga ah sida marxaladda wareega, muunada si buuxda ayaa loogu hayaa sanduuqa flash-ka.Marka xigta, methanol-ka wareega wareega ayaa si toos ah loo kordhiyey 100% iyo gradient-ka waxaa lagu sii hayay 7.5 CV.Muunadda ayaa laga saaray min 11.5 ilaa 13.5 CV.Jajabyada la ururiyey, xalka muunada waxa laga beddelay NaH2PO4 xal kaydiyaha oo loo beddelay methanol.Marka la barbar dhigo xalka aqueous aadka u sarreeya, methanol aad bay u fududayd in laga saaro uumiga wareega ee tallaabada xigta, taas oo sahlaysa cilmi-baarista soo socota.
Jaantuska 3. Kormatogaraamka toosh ee muunada ee ku dul yaal kartoonka C18AQ.
Si loo barbardhigo hab-dhaqanka sii-haynta kartoojka C18AQ iyo kartoojka caadiga ah ee C18 ee muunadaha polarity xooggan, tijaabada isbarbardhigga ayaa la sameeyay.Kartridge SepaFlash C18 RP ah ayaa la isticmaalay iyo chromatogram-ga muunada ee muunada ayaa lagu muujiyay Jaantuska 4. Kartijyada caadiga ah ee C18, saamiga wajiga biyaha ee ugu sarreeya ee loo dulqaadan karo waa ilaa 90%.Sidaa darteed jilbaha bilawga ah ayaa lagu dejiyay 10% methanol 90% biyaha ah.Sida ku cad Jaantuska 4, sababtoo ah burburka wejiga hydrophobic ee silsiladaha C18 ee ay sababtay saamiga biyaha sare, muunada ayaa si dirqi ah loogu hayaa kartuunada caadiga ah ee C18 waxaana si toos ah uga baxay wajiga wareega.Natiijo ahaan, hawlgalka nadiifinta saamiga ama nadiifinta lama dhammaystiri karo.
Jaantuska 4. Karamatogaraamka toosh ee muunada ee ku jira kartoonka C18 ee caadiga ah.
Marka la barbar dhigo gradient toosan, isticmaalka jaranjarada jaranjarada waxay leedahay faa'iidooyinka soo socda:
1. Isticmaalka xalinta iyo wakhtiga socodsiinta ee muunad nadiifinta waa la dhimay.
2. Badeecada bartilmaameedka ahi waxay ku dhalataa meel sare oo fiiqan, taas oo yaraynaysa mugga jajabyada la ururiyey oo sidaas awgeed fududeynaya uumiga wareega ee soo socda iyo sidoo kale kaydinta wakhtiga.
3. Alaabta la ururiyey waxay ku jirtaa methanol taas oo si sahlan loo uumi-baxsan yahay, sidaas awgeed wakhtiga qalajinta ayaa la dhimay.
Gebogebadii, sifaynta muunada oo ah mid aad u xoog badan ama hydrophilic, SepaFlash C18AQ RP flash cartridges oo lagu daro nidaamka chromatography ee diyaarinta ah ee SepaBean ™ Mashiinka ayaa bixin kara xal degdeg ah oo hufan.
Ku saabsan SepaFlash Bonded Taxanaha C18 RP flash cartridges
Waxaa jira taxane ah kartoonada flash-ka ee SepaFlash C18AQ RP oo leh tilmaamo kala duwan oo ka yimid Tignoolajiyada Santai (sida ku cad Shaxda 2).
| Lambarka Shayga | Cabirka tiirka | Heerka socodka (ml/min) | Ugu badnaan.cadaadiska (psi/bar) |
| SW-5222-004-SP(AQ) | 5.4 g | 5-15 | 400/27.5 |
| SW-5222-012-SP(AQ) | 20 g | 10-25 | 400/27.5 |
| SW-5222-025-SP(AQ) | 33 g | 10-25 | 400/27.5 |
| SW-5222-040-SP(AQ) | 48 g | 15-30 | 400/27.5 |
| SW-5222-080-SP(AQ) | 105 g | 25-50 | 350/24.0 |
| SW-5222-120-SP(AQ) | 155 g | 30-60 | 300/20.7 |
| SW-5222-220-SP(AQ) | 300 g | 40-80 | 300/20.7 |
| SW-5222-330-SP(AQ) | 420 g | 40-80 | 250/17.2 |
Shaxda 2. SepaFlash C18AQ RP flash cartridges.
Qalabka wax lagu baakadeeyo: Waxtarka spherical C18 (AQ) ee silica isku xiran, 20 - 45 μm, 100 Å.
logy (sida ku cad shaxda 2).
Waqtiga boostada: Agoosto-27-2018