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ODS固定相を用いた血漿試料直接注入による血中ランソプラゾール濃度測定法○ 立木秀尚1，矢澤到2
東和薬品株式会社1，インタクト株式会社2

概要
前処理に関するアンケート

Novel Hydrophobic/Hydrophilic Hybrid ODS Column

Direct Injection LC-MS/MS Analysis

Clinical Trial1前処理に関するアンケート
Q1. 前処理はやりたくないですか？

有効回答数 23
MS/MSユーザーズML 調査
2

前処理に関するアンケート
Q2. よく行なう前処理法は何ですか？

有効回答数 23
MS/MSユーザーズML 調査
3

前処理に関するアンケート
Q2. よく行なう前処理法は何ですか？

有効回答数 23
MS/MSユーザーズML 調査
4生体試料直接注入の問題点① 前処理カラムやカラムスイッチングが必要② 汎用分析に適用しにくい
生体試料分析専用カラム
有機溶媒濃度に制限
リン酸緩衝液を常用（LC-MS分析が困難）
ODSが存在しない

5生体試料直接注入の問題点 解決① 前処理用カラムやカラムスイッチングは不要② 汎用分析でも使用可能なODSカラムでの条件を満たす分析法開発

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固定相構造血清直接注入

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Compounds

Lansoprazole( Analyte )Omeprazole( Internal Standard )9Sample Preparation
Direct Injection↓Plasma↓Centrifugation↓Addition of I.S.
9:1 H2O ( I.S. ) to Plasma↓Injection

Direct Injection vs.
Protein Precipitation

Protein Precipitation↓Plasma↓Centrifugation↓Protein Precipitation
4:1 ACN ( IS ) to Plasma↓Filtration↓Injection

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Direct Injection vs.
Protein Precipitation

Analytical Conditions
Direct Injection

Protein Precipitation

ColumnColumn

Cadenza HS-C18ODS Column

20×4.6 mm, 3 µm ( Imtakt )with Guard Column
Mobile Phase
A, 10 mM ammonium formate( pH3.0 )B, Acetonitrile
100 % A( 0→0.5 min. )0→100 % B ( 0.5→2.5 min. )100 %B( 2.5→3.5 min. )20×4.6 mm, 5 µm
without Guard Column
IsocraticA, 10 mM ammonium formate( pH3.0 )B, Methanol
C, Acetonitrile
A/B/C = ( 27.5 : 35.0 : 37.5 )0 - 8.5 min.

Flow Rate 500 µL/min.
Temperature 40 deg. C
Injection Volume 10 µL
MS/MS ESI, Positive, MRM

Flow Rate 70 µL/min.
Temperature 40 deg. C
Injection Volume 1 µL
MS/MS ESI, Positive, MRM

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Direct Injection vs.
Protein Precipitation

Chromatogram
LOQ, plasma

Plasma Blank
Lansoprazole

Omeprazole

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Direct Injection vs.
Protein Precipitation

Linearityy = 0.0017x - 0.0072
R = 0.998

10 - 3000 ng/mL, Weight 1/x2, n = 3
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Direct Injection vs.
Protein Precipitation

Linearityy = 0.0017x - 0.0072
R = 0.998

y = 0.0016x - 0.0012
R = 0.999

10 - 3000 ng/mL, Weight 1/x2, n = 3
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Direct Injection vs.
Protein Precipitation

Accuracy and Precision

Nominal Concentration ( ng/mL )10
Accuracy ( Mean, % ) -1.9300

3000

5.5

4.3 -0.4-6.4

0.7Precision ( CV, % )Intra-batch ( n = 3 )

2.7

4.0

2.4

3.1

2.5

2.8

Inter-batch ( n = 6 )

5.0

6.1

4.0

5.1

6.6

4.8

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Quality Control

Clinical Trial

CV = 8.8 %Lansoprazole 30 mg Single Dosa ( One Healthy Adult male )● - IS Peak Area
◆ - QC Samples ( 10, 300 and 3000 ng/mL ); QC Range = ±15 %16

Longevity

Clinical Trial

17Plasma Concentrations

Clinical Trial●●AUC 2407
2304Cmax 1141.8 1127.2● - Direct Injection● - Protein Precipitation

AUC - ng·h/mLCmax - ng·/mL
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Metabolites

Clinical Trial

Lansoprazole

Lansoprazole Sulfide

5-hydroxy Lansoprazole

Lansoprazole Sulfone

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まとめ
Novel Hydrophobic/Hydrophilic Hybrid ODS Column
Cadenza HS-C18
Direct Injection LC-MS/MS Analysis of
Lansoprazole in human plasma
Method Validation
Clinical Trial
Direct Injection vs. Protein precipitation
20

まとめNovel Hydrophobic/Hydrophilic Hybrid ODS Column
Cadenza HS-C18
Direct Injection LC-MS/MS Analysis of
Lansoprazole in human plasma
Method Validation
Clinical Trial
Direct Injection vs. Protein precipitation

Satisfactory

21

まとめ
Novel Hydrophobic/Hydrophilic Hybrid ODS Column
Cadenza HS-C18
Direct Injection LC-MS/MS Analysis of
Lansoprazole in human plasma

ExMethod
Validation
cell Trial
ClinicalSatisfactory
en precipitation
Direct Injection vs. Proteint?
22

まとめNovel Hydrophobic/Hydrophilic Hybrid ODS Column
Cadenza HS-C18
Direct Injection LC-MS/MS Analysis of
Lansoprazole in human plasma

Ex Pe
MethodValidation
crfee
ClinicalTrial
Satisfactorylcle
Direct Injection vs. Protein
nttprecipitation?
?
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