Rui Huang, Bo Xu
Ka Papahana R&D Center
Hoʻolauna
ʻO ka peptide kahi pūhui i haku ʻia me nā waikawa amino, ʻo kēlā me kēia mea he mau waiwai kino a me nā kemika ma muli o nā ʻano like ʻole a me ke ʻano o nā koena amino acid i hoʻokumu ʻia i kāna kaʻina.Me ka hoʻomohala ʻana i ka synthesis chemical phase paʻa, ua holomua nui ka synthesis chemical o nā ʻano peptides ikaika.Eia nō naʻe, ma muli o ka paʻakikī paʻakikī o ka peptide i loaʻa e ka solid phase synthesis, pono e hoʻomaʻemaʻe ʻia ka huahana hope e nā ʻano hoʻokaʻawale hilinaʻi.ʻO nā ʻano hoʻomaʻemaʻe maʻamau i hoʻohana ʻia no nā peptides ʻo ia ka ion exchange chromatography (IEC) a me ka reversed-phase high performance liquid chromatography (RP-HPLC), nona nā hemahema o ka haʻahaʻa haʻahaʻa haʻahaʻa haʻahaʻa, ke kumu kūʻai kiʻekiʻe o ka media kaʻawale, paʻakikī a me nā lako hoʻokaʻawale kumu kūʻai. etc. No ka hoʻomaʻemaʻe wikiwiki ʻana i nā peptides liʻiliʻi liʻiliʻi (MW <1 kDa), ua paʻi mua ʻia kahi hihia noi kūleʻa e Santai Technologies, kahi i hoʻohana ʻia ai kahi pahu pahu SepaFlash RP C18 no ka hoʻomaʻemaʻe wikiwiki ʻana o thymopentin (TP-5) a me ka ua loaʻa ka huahana pahuhopu i nā koi.
Kiʻi 1. 20 mau amino acids maʻamau (i hoʻopuka hou ʻia mai www.bachem.com).
Aia he 20 mau ʻano amino acids maʻamau i ka haku ʻana o nā peptides.Hiki ke hoʻokaʻawale ʻia kēia mau ʻakika amino i loko o nā pūʻulu e like me ko lākou polarity a me ka waiwai kumu waiwai: non-polar (hydrophobic), polar (uncharged), acidic a kumu paha (e like me ka mea i hōʻike ʻia ma ke Kiʻi 1).I loko o kahi kaʻina peptide, inā he polar ka nui o nā amino acids e hoʻokumu ana i ke kaʻina (e like me ka mea i hōʻailona ʻia i ka waihoʻoluʻu ʻulaʻula ma ke Kiʻi 1), e like me Cysteine, Glutamine, Asparagine, Serine, Threonine, Tyrosine, etc. polarity a hiki loa i ka wai.I loko o ke kaʻina hana hoʻomaʻemaʻe no kēia mau laʻana peptide polar ikaika e ka reversed-phase chromatography, e hiki mai ana kahi mea i kapa ʻia ʻo hydrophobic phase collapse (e nānā i kahi palapala noi i paʻi mua ʻia e Santai Technologies: Hydrophobic Phase Collapse, AQ Reversed Phase Chromatography Columns a me kā lākou mau noi).Hoʻohālikelike ʻia me nā kolamu C18 maʻamau, ʻoi aku ka maikaʻi o nā kolamu C18AQ i hoʻomaikaʻi ʻia no ka hoʻomaʻemaʻe ʻana i nā polar ikaika a i ʻole hydrophilic samples.Ma kēia pou, ua hoʻohana ʻia kahi peptide polar ikaika e like me ka laʻana a hoʻomaʻemaʻe ʻia e kahi kolamu C18AQ.ʻO ka hopena, ua loaʻa ka huahana i hoʻokō ʻia i nā koi a hiki ke hoʻohana ʻia i ka noiʻi a me ka hoʻomohala ʻana.
| Mea hana | SepaBean™mīkini 2 | |||
| Nā pahu pahu | 12 g SepaFlash C18 RP pahu pahu uila (spherical silica, 20 - 45 μm, 100 Å, Order nunber:SW-5222-012-SP) | 12 g SepaFlash C18AQ RP pahu pahu uila (spherical silica, 20 - 45 μm, 100 Å, Helu kauoha:SW-5222-012-SP(AQ)) | ||
| Loʻihi nalu | 254 nm, 220 nm | 214 nm | ||
| Māhele kelepona | Solvent A: Wai Mea hoʻoheheʻe B: Acetonitrile | |||
| Kahe kahe | 15 mL/min | 20 mL/min | ||
| Hoʻouka laʻana | 30 mg | |||
| Gradient | Manawa (CV) | Mea hoʻoheheʻe B (%) | Manawa (min) | Mea hoʻoheheʻe B (%) |
| 0 | 0 | 0 | 4 | |
| 1.0 | 0 | 1.0 | 4 | |
| 10.0 | 6 | 7.5 | 18 | |
| 12.5 | 6 | 13.0 | 18 | |
| 16.5 | 10 | 14.0 | 22 | |
| 19.0 | 41 | 15.5 | 22 | |
| 21.0 | 41 | 18.0 | 38 | |
| / | / | 20.0 | 38 | |
| 22.0 | 87 | |||
| 29.0 | 87 | |||
Nā hualoaʻa a me ke kūkākūkā
No ka hoʻohālikelike ʻana i ka hana hoʻomaʻemaʻe no ka laʻana peptide polar ma waena o ke kolamu C18 maʻamau a me ke kolamu C18AQ, ua hoʻohana mākou i kahi kolamu C18 maʻamau no ka hoʻomaʻemaʻe uila o ka hāpana ma ke ʻano he hoʻomaka.E like me ka mea i hōʻike ʻia ma ke Kiʻi 2, ma muli o ka hāʻule ʻana o ka pae hydrophobic o nā kaulahao C18 i hoʻokumu ʻia e ka ratio wai kiʻekiʻe, ua paʻa ʻole ka hāpana ma ka pahu pahu C18 maʻamau a ua hoʻokaʻawale pololei ʻia e ka pae kelepona.ʻO ka hopena, ʻaʻole i hoʻokaʻawale ʻia a hoʻomaʻemaʻe ʻia ka hāpana.
Kiʻi 2. ʻO ka chromatogram flash o ka hāpana ma kahi pahu pahu C18 maʻamau.
A laila, ua hoʻohana mākou i kahi kolamu C18AQ no ka hoʻomaʻemaʻe uila o ka hāpana.E like me ka mea i hōʻike ʻia ma ka Figure 3, ua mālama maikaʻi ʻia ka peptide ma ke kolamu a laila eluted.Ua hoʻokaʻawale ʻia ka huahana i hoʻopaʻa ʻia mai nā haumia i loko o ka hāpana maka a hōʻiliʻili ʻia.Ma hope o ka lyophilization a nānā ʻia e HPLC, ʻo ka huahana i hoʻomaʻemaʻe ʻia he 98.2% ka maʻemaʻe a hiki ke hoʻohana hou ʻia no ka noiʻi a me ka hoʻomohala ʻana.
Kiʻi 3. ʻO ka chromatogram flash o ka hāpana ma kahi pahu pahu C18AQ.
I ka hopena, ua hui pū ʻia ka pahu pahu uila SepaFlash C18AQ RP me ka ʻōnaehana chromatography flash SepaBean™Hiki i ka mīkini ke hāʻawi i kahi hopena wikiwiki a maikaʻi no ka hoʻomaʻemaʻe ʻana i nā polar ikaika a hydrophilic paha.
Aia kekahi pūʻulu o nā pahu uila SepaFlash C18AQ RP me nā kikoʻī like ʻole mai Santai Technology (e like me ka hōʻike ʻana ma ka Papa 2).
| Helu 'ikamu | Ka nui o ke kolamu | Kahe Kahe (mL/min) | Max.Pressure (psi/pa) |
| 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 |
Papa 2. SepaFlash C18AQ RP pahu pahu flash.Mea hoʻopili: High-efficiency spherical C18(AQ)-hoʻopaʻa silica, 20 - 45 μm, 100 Å.
No ka ʻike hou aku e pili ana i nā kikoʻī kikoʻī o ka mīkini SepaBean™, a i ʻole ka ʻike kauoha ma nā pahu pahu uila SepaFlash series, e ʻoluʻolu e kipa i kā mākou pūnaewele.
Ka manawa hoʻouna: Oct-12-2018