Rui Huang, Bo Xu
Daim ntawv thov R&D Center
Taw qhia
Lub peptide yog ib qho kev sib xyaw ua ke ntawm cov amino acids, txhua tus uas muaj lub cev thiab tshuaj lom neeg tshwj xeeb vim muaj ntau hom thiab kev txiav txim ntawm cov amino acid residues tsim nws cov kab ke.Nrog rau txoj kev loj hlob ntawm cov khoom theem tshuaj synthesis, cov tshuaj synthesis ntawm ntau yam active peptides tau ua tau zoo.Txawm li cas los xij, vim muaj qhov nyuaj ntawm cov peptide tau los ntawm cov khoom sib txuas ua ke, cov khoom kawg yuav tsum tau ua kom huv los ntawm kev sib cais txhim khu kev qha.Cov kev siv purification feem ntau rau peptides suav nrog ion txauv chromatography (IEC) thiab thim rov qab-theem siab ua kua chromatography (RP-HPLC), uas muaj qhov tsis zoo ntawm cov qauv qis qis, tus nqi siab ntawm kev sib cais tawm, cov khoom siv sib cais nyuaj thiab kim, thiab lwm yam. Rau kev ua kom huv sai ntawm me me molecule peptides (MW < 1 kDa), ib daim ntawv thov kev vam meej yav dhau los tau luam tawm los ntawm Santai Technologies, uas yog siv SepaFlash RP C18 cartridge rau kev ua kom huv sai ntawm thymopentin (TP-5) thiab cov khoom lag luam ua tau raws li qhov yuav tsum tau ua.
Daim duab 1. 20 hom amino acids (reproduced from www.bachem.com).
Muaj 20 hom amino acids uas muaj nyob rau hauv muaj pes tsawg leeg ntawm peptides.Cov amino acids no tuaj yeem muab faib ua pawg hauv qab no raws li lawv cov polarity thiab acid-puag khoom: non-polar (hydrophobic), polar (uncharged), acidic lossis yooj yim (raws li qhia hauv daim duab 1).Nyob rau hauv ib theem peptide, yog tias cov amino acids tsim cov kab ke feem ntau yog cov polar sawv daws yuav (raws li cim xim liab hauv daim duab 1), xws li Cysteine, Glutamine, Asparagine, Serine, Threonine, Tyrosine, thiab lwm yam, cov peptide no yuav muaj zog. polarity thiab soluble heev hauv dej.Thaum lub sij hawm purification txheej txheem rau cov muaj zog polar peptide cov qauv los ntawm reversed-theem chromatography, ib tug tshwm sim hu ua hydrophobic theem vau yuav tshwm sim (saib rau ib tug yav tas los daim ntawv thov daim ntawv sau npe los ntawm Santai Technologies: Hydrophobic Phase Collapse, AQ Reversed Phase Chromatography Kab thiab Lawv daim ntawv thov).Piv nrog rau cov kab C18 li niaj zaus, cov kab C18AQ txhim kho zoo tshaj plaws rau kev ua kom huv ntawm cov polar muaj zog lossis cov qauv hydrophilic.Hauv cov ntawv tshaj tawm no, muaj zog polar peptide tau siv los ua tus qauv thiab ua kom huv los ntawm kab C18AQ.Yog li ntawd, lub hom phiaj cov khoom ua tau raws li qhov yuav tsum tau muaj thiab tuaj yeem siv rau hauv kev tshawb fawb thiab kev tsim kho hauv qab no.
| Ntsuas | SepaBean™tshuab 2 | |||
| Cartridges | 12 g SepaFlash C18 RP flash cartridge (spherical silica, 20 - 45 μm, 100 Å, Order nunber: SW-5222-012-SP) | 12 g SepaFlash C18AQ RP flash cartridge (spherical silica, 20 - 45 μm, 100 Å, Order naj npawb: SW-5222-012-SP(AQ)) | ||
| Lub wavelength | 254 nm, 220 nm | 214nm ua | ||
| Mobile theem | Solvent A: Dej Cov kuab tshuaj B: Acetonitrile | |||
| Tus nqi ntws | 15 ml / min | 20 ml / min | ||
| Sample loading | 30 mg | |||
| Gradient | Lub sijhawm (CV) | Cov kuab tshuaj B (%) | Sijhawm (min) | Cov kuab tshuaj B (%) |
| 0 | 0 | 0 | 4 | |
| 1.0 | 0 | 1.0 | 4 | |
| 10.0 | 6 | 7.5 | 18 | |
| 12.5 Nws | 6 | 13.0 Nws | 18 | |
| 16.5 ib | 10 | 14.0 Nws | 22 | |
| 19.0 Nws | 41 | 15.5 ib | 22 | |
| 21.0 Nws | 41 | 18.0 Nws | 38 | |
| / | / | 20.0 Nws | 38 | |
| 22.0 Nws | 87 | |||
| 29.0 Nws | 87 | |||
Cov txiaj ntsig thiab kev sib tham
Txhawm rau sib piv cov kev ua kom huv huv rau cov qauv polar peptide nruab nrab ntawm kab C18 ib txwm thiab C18AQ kem, peb siv C18 kem tsis tu ncua rau flash purification ntawm cov qauv ua qhov pib.Raws li pom nyob rau hauv daim duab 2, vim yog lub hydrophobic theem vau ntawm lub C18 chains tshwm sim los ntawm high aqueous ratio, tus qauv twb NW nyuam qhuav khaws cia nyob rau hauv ib txwm C18 cartridge thiab tau ncaj qha eluted tawm los ntawm lub mobile theem.Raws li qhov tshwm sim, tus qauv tsis zoo sib cais thiab huv si.
Daim duab 2. Lub teeb chromatogram ntawm cov qauv ntawm lub cartridge C18 tsis tu ncua.
Tom ntej no, peb siv C18AQ kem rau flash purification ntawm tus qauv.Raws li pom nyob rau hauv daim duab 3, peptide tau zoo khaws cia rau ntawm kab thiab ces eluted tawm.Lub hom phiaj khoom raug cais tawm ntawm cov impurities hauv cov qauv nyoos thiab sau.Tom qab lyophilization thiab tom qab soj ntsuam los ntawm HPLC, cov khoom ua kom huv si muaj purity ntawm 98.2% thiab tuaj yeem siv ntxiv rau kev tshawb fawb thiab kev txhim kho cov kauj ruam tom ntej.
Daim duab 3. Lub teeb chromatogram ntawm tus qauv ntawm C18AQ cartridge.
Hauv kev xaus, SepaFlash C18AQ RP flash cartridge ua ke nrog lub flash chromatography system SepaBean™tshuab tuaj yeem muab kev daws teeb meem sai thiab zoo rau kev ua kom huv ntawm cov polar muaj zog lossis cov qauv hydrophilic.
Muaj ib tug series ntawm SepaFlash C18AQ RP flash cartridges nrog txawv specifications ntawm Santai Technology (raws li qhia nyob rau hauv Table 2).
| Yam khoom naj npawb | Kem Loj | Tus nqi ntws (ml / min) | Max. Siab (psi/bar) |
| SW-5222-004-SP(AQ) | 5,4g e | 5-15 | 400/27.5 |
| SW-5222-012-SP(AQ) | 20 g ib | 10-25 | 400/27.5 |
| SW-5222-025-SP(AQ) | 33g ua | 10-25 | 400/27.5 |
| SW-5222-040-SP (AQ) | 48g ua | 15-30 | 400/27.5 |
| SW-5222-080-SP(AQ) | 10 5g | 25-50 | 350/24.0 Nws |
| SW-5222-120-SP(AQ) | 15 5g | 30-60 | 300/20.7 |
| SW-5222-220-SP(AQ) | 300 g | 40-80 : kuv | 300/20.7 |
| SW-5222-330-SP(AQ) | 420g ua | 40-80 : kuv | 250/17.2 |
Table 2. SepaFlash C18AQ RP flash cartridges.Cov ntaub ntawv ntim: High-efficiency spherical C18 (AQ)-bonded silica, 20 - 45 μm, 100 Å.
Yog xav paub ntxiv txog cov lus qhia ntxaws ntxaws ntawm SepaBean ™ tshuab, lossis cov ntaub ntawv xaj ntawm SepaFlash series flash cartridges, thov mus saib peb lub vev xaib.
Post lub sij hawm: Oct-12-2018