これでレベルアップ ★逆相HPLC分析における メソッド開発 …

1 0 JASIS 2013 hplc 1 JASIS 2013 hplc 1. hplc 2. 2 JASIS 2013 hplc 3 JASIS 2013 hplc pH 4 JASIS 2013 hplc pKa log Pow 5000 30 nm 5 JASIS 2013 PDA UV : ODS 3 m or 5 m : 150 mm : + : 10 50 mM : 10 50 mM pH7 : 1 mL/min : 1 mg/mL 1 L : PDA 210 400 nm ODS 2 m 3 50 mm 50 mm 6 JASIS 2013 ODS (ODS ) ( ) ( ) ( ) ( ) 7 JASIS 2013 ODS ODS ODS L-column2 ODS 8 JASIS 2013 ( ) 01020304050L-column2 ODSB rand A ODS Brand B ODS Analytical conditions Column: C18(ODS), 5 mColumn size: 150 mmMobile phase: CH3CN/25 mM Phosphate buffer pH 7(60/40)Flow rate.

2 1 mL/minTemp.: 40 CDetection: UV 220 nmSample: 1. 2. 3. 4. 5. 6. 7. 8. 9. 1 2 3 4 5 6 7 8 9 1 2 3 4 5 7 6 8 9 1 2 3 4 5 6 7 8 9 9 JASIS 2013 UHPLC 10 JASIS 2013 : 5 m : 150 mm : mL/min 2 m5 m (3)Rs(1,2)N(3) Analytical conditions Column: L-column2 ODSM obile phase: CH3OH/25 mM Phosphate buffer pH 7 (15/85)Temp.: 40 CDetection: UV 260 : 1 LSample: 1. (60 mg/L)2.

3 (30 mg/L)3. (30 mg/L)in mobile phaseSystem: Agilent 1200SL051015123123 : 2 m : 75 mm : mL/min 1/6 tR: ; N: ; Rs: 11 JASIS 2013 ODS C8 C6-Phenyl 12 JASIS 2013 ODS ( ) C8 ( ) C6-Phenyl ( )+ - ( )C8 C6-Phenyl ODS C8 C6-Phenyl 13 JASIS 2013 C8 Analytical conditions Column: 5 mColumn size: 150 mmMobile phase: CH3CN/25 mM Phosphate buffer pH 7 (30/70)Flow rate: 1 mL/minTemp.: 40 CDetection: UV 220 : 1 LSample: 1. 2. 3. 4. 5.

4 6. System: LC-10 ADvp series( ) 010203040 (min)L-column2 C8L-column2 ODSRs(2,3)= (2,3)= : 14 JASIS 2013 C6-Phenyl 123123 Analytical conditions Column: 5 mColumn size: 150 mmMobile phase: CH3OH/H2O (50/50)Flow rate: 1 mL/minTemp.: 40 CDetection: UV 210 : 2 LSample: 1. 2. m- 3. 1, 3, 5- System: LC-10 ADvp series( )051015 (min)NO2NO2O2N 1, 3, 5- L-column2 ODSL-column2 C6-Phenyl15 JASIS 2013 C6-Phenyl 01020 (min)NONNCH3 FCH3 OHOOHODS 123123 Analytical conditions Column: 5 mColumn size: 150 mmMobile phase: CH3OH/20 mM H3PO4(10/90)Flow rate: 1 mL/minTemp.

5 : 40 CDetection: UV 294 : 2 LSample: 1. A 2. B3. System: LC-10 ADvp series( )3. L-column2 ODSL-column2 C6-Phenyl16 JASIS 2013 LC-MS/MS LC-MS ODS C8 C6-Phenyl 18 ( )L-column2 ODSL-column2 C6-PhenylL-column2 C817 JASIS 2013 hplc 18 JASIS 2013 : UV 3 m : UV UV250nm : PEEK : hplc BHT 19 JASIS 2013 /20 mM (35/65) /20 mM (35/65)0246802468 ( ) ( ) Analytical conditions Column: L-column ODS, 5 m; Column size: 150 mm; Flow rate: 1 mL/min; Temp.

6 : 30 C20 JASIS 2013 pH (buffer solution) H3PO4 H2PO4-( NH3 NH4 ) : (pH= )H3PO4 H2PO4-+H+ (1:1 ) pH pKa 1 21 JASIS 2013 02040608010012345678910111213 pKa ( )(pH)H3PO4H2PO4-HPO42-PO43- HO P OHOH O pH 4 pH9 22 JASIS 2013 hplc 5 50 mM1 10 5 10 (HCO3) (NH4+) (CO32-) pKaMS MS pKa 1 10 mM( ) ( 5 )23 JASIS 2013 A) 20 mM KH2PO4(pH ) (75/25) B) 20 mM (pH ) (75/25) ( )OHOOCH3 Analytical conditions Column: L-column2 ODS, 5 m; Column size; 150 mmSample: 1.

7 (100 mg/L); 2. (100 mg/L) 02468100246810 1 L2 L5 L122. 1. OHO24 JASIS 2013 ( ) ( ) B)20 mM A) 20 mM KH2PO4 Analytical conditions Column: L-column2 ODS, 5 m; Column size: 150 mmMobile Phase: A)20 mM KH2PO4(pH )/CH3CN (75/25)B)20 mM Acetate buffer (pH )/CH3CN (75/25) : 2 LSample: 1. (100 mg/L); 2. (100 mg/L) 25 JASIS 2013 C4H9N+C4H9C4H9C4H9Br-RSO3-C4H9N+C4H9C4H9 C4H9 RSO3-Na+ R1N+R2R4R3X-RSO3-Na+R1N+R2R4R3 RSO3-pH (TBA) pH 26 JASIS 2013 NaClO4 C12H25 OSO3Na n- (SDS)Cl Br (C4H9)4N,H2PO4 (TBA-P)1 n 7 CnF2n+1 COOH 3 n 6(CnH2n+1)2NH 3 n 13 CnH2n+1SO3Na MS 27 JASIS 2013.

8 : : : : 28 JASIS 2013 05010012345678 pH ( )(pH)COOH pH : : pH pKa pH pH pKa 2 COO-29 JASIS 2013 pH pH min Analytical conditions Column: L-column ODS, 5 mColumn size: 150 mmMobile phase: CH3CN/25 mM (25/75) ( )Flow rate: 1 : 1 LSample: ( )pH pH 02468pH 30 JASIS 2013 Analytical conditions Column: L-column2 ODS, 5 mColumn size: 150 mmMobile phase: CH3CN/10 mM TBA-PO4in H2O(45/55)Flow rate: 1 mL/minDetection: VIS 430 : 1 LNNSO3 NaOHNNOHSO3 NaNNNHSO3Na1.

9 - 2. 73. 50510123 31 JASIS 2013 : : : : : : 32 JASIS 2013 246810 Analytical conditions Column: L-column2 ODS, 5 mColumn size: 150 mmMobile phase:CH3CN/20 mM H3PO4(30/70)Flow rate: 1 mL/minTemp: 40 : 1 LSample: OOHNHCH3CH3 pKa 1000 mg/L50 mg/LN=4000N=10000N: 33 JASIS 2013 Analytical conditions Column: L-column2 ODS, 5 mColumn size: 150 mmMobile phase:CH3CN/25 mM pH 7(30/70)Flow rate: 1 mL/minTemp: 40 : 1 LSample: 246810 OOHNHCH3CH3 pKa 1000 mg/L50 mg/LN=12200N=12000N: 34 JASIS 2013 Analytical conditions Column: ODS(C18), 5 mColumn size: 150 mmMobile phase:CH3CN/25 mM pH 7(30/70)Flow rate: 1 mL/minTemp: 40 : 1 LSample.

10 (50 mg/L)246810N=9600N=12000 L-column2 ODSOOHNHCH3CH3 pKa N: 35 JASIS 2013 Analytical conditions Column L-column2 150 mm (C18, 5 m)Mobile phase ACH3CN/20 mM H3PO4(30/70)BCH3CN/20 mM H3PO4 10 mM C5H11SO3Na(30/70)Flow rate 1 mL/minTemp. 40 1 LSample N+OOOCH3 OCH3minN=3136N=10826 BA024681036 JASIS 2013 ( ) 37 JASIS 2013 Analytical conditions Column: L-column ODS, 5 mColumn size: 150 mmFlow rate: 1 mL/minTemp: 40 CDetection: 225 : 1 LSample: (in CH3CN)6 8 10 12 14minCH3OH/25 mM pH 7 (80/20)CH3CN/25 mM pH 7 (55/45)S=.