Enantiomeric Determination ofd- andl-Lactate in Rat Serum Using High-Performance Liquid Chromatography with a Cellulose-Type Chiral Stationary Phase and Fluorescence Detection

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A method for the enantiomeric determination of 2,7,8-trimethyl-2-(beta-carboxyethyl)-6-hydroxy chroman (LLU-alpha, gamma-CEHC) in rat plasma was developed using high-performance liquid chromatography (HPLC) with a fluorimetric derivatization with 4-N, N-dimethylaminosulfonyl-7-piperazino-2,1,3-benzoxadiazole (DBD-PZ) followed by O-acetylation with acetyl chloride. The proposed HPLC system used two non-chiral columns (phenyl and octadecylsilica) and a chiral column (a modified cellulose type), which were connected via two column-switching valves. A derivatized sample prepared from rat plasma was first separated on the phenyl column, and the fraction including LLU-alpha derivative was introduced to the octadecylsilica column to quantify the concentration of the mixture of S- and R-LLU-alpha. Finally, the LLU-alpha derivative was directly injected into the chiral column to obtain the ratio of the enantiomers. The proposed HPLC system was applied to the enantiomeric determination of LLU-alpha in plasma after intravenous administration of racemic LLU-alpha. S-LLU-alpha was eliminated faster than R-LLU-alpha, and its concentration in plasma decreased to one-third at 2 min after dosing.

Section: Resultsmentioning

confidence: 99%

A fluorimetric, column‐switching HPLC and its application to an elimination study of LLU‐α enantiomers in rat plasma

Hattori

Fukushima

Hamamura

et al. 2001

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“…While DBD-PZ was used as a fluorescent reagent Ichihara et al, 1999), NBD-PZ was also employed as a fluorescent reagent in this study (Fig. 1), because NBD moiety exhibits a strong fluorescence intensity (Toyo'oka et al, 1991).…”

Section: Resultsmentioning

confidence: 99%

Simultaneous determination of D‐lactic acid and 3‐hydroxybutyric acid in rat plasma using a column‐switching HPLC with fluorescent derivatization with 4‐nitro‐7‐piperazino‐2,1,3‐benzoxadiazole (NBD‐PZ)

Fukushima

Lee

Korenaga

et al. 2001

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A highly sensitive method for the determination of D-lactic acid and 3-hydroxybutyric acid (3-HB) in rat plasma was developed using high-performance liquid chromatography with octadecylsilica (ODS) connected to a chiral column. At first, (D + L)-lactic acid and 3-HB in the plasma were derivatized with a fluorescent reagent, 4-nitro-7-piperazino-2,1,3-benzoxadiazole (NBD-PZ), separated on the ODS column and determined fluorimetrically at 547 nm with 491 nm of excitation wavelength. During the separation step on the ODS, the peak fraction of (D + L)-lactate derivative was introduced directly to a phenylcarbamoylated beta-cyclodextrin chiral column by changing the flow of the eluent via a six-port valve. Then, D-lactate derivative was separated enantiomerically from the L-lactate derivative, and the enantiomeric ratio was determined from the chromatogram. Intra- and inter-day accuracy values for the determination of D-lactic acid in 10 microL of rat plasma were 97.8-109.2 and 98.4-109.9%, and those for 3-HB were 99.8-108.4 and 99.8-103.8%, respectively. The intra- and inter-day precision values were within 4.6 and 5.1% for D-lactic acid, and 2.7 and 2.4% for 3-HB, respectively. The detection limits for D-lactic acid and 3-HB were approximately 2.0 and 0.04 microM, respectively (signal-to-noise ratio 3). The proposed method was applied to the plasma of diabetic rats induced by intraperitoneal administration of streptozotocin, and the significant increases of both D-lactic acid and 3-HB concentrations were observed in the diabetic rats as compared to the normal rats.

“…The detection limits for DBD-PZ-carboxylic acids derivatives were 3.2-4.7 fmol (S/N = 3) using HPLC (Toyo'oka et al, 1991a). DBD-PZ was applied to the determination of prostaglandins (Toyo'oka et al, 1992b), a novel H 1 -receptor antagonist HSR-609 (Hasegawa et al, 1996), 2-arylpropionic acids on reversedphase and chiral stationary phase , D-and L-lactate on reversed-phase and chiral stationary phase Ichihara et al, 1999), free fatty acids (Ueno et al, 1999), and 2,7,8-trimethyl-2-(b-carboxyethyl)-6-hydroxy chroman (LLUa) on reversed-phase and chiral stationary phase (Hattori et al, 2000).…”

Section: For Carboxylic Acids Nbd-pzmentioning

confidence: 99%

Fluorogenic and fluorescent labeling reagents with a benzofurazan skeleton

Uchiyama

Santa

Okiyama

et al. 2001

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151

101

Fluorogenic and fluorescent labeling reagents having a benzofurazan (2,1,3-benzoxadiazole) skeleton such as 4-fluoro-7-nitro-2,1,3-benzoxadiazole (NBD-F), 4-N,N-dimethylaminosulfonyl-7-fluoro-2,1,3-benzoxadiazole (DBD-F), 4-aminosulfonyl-7-fluoro-2,1,3-benzoxadiazole (ABD-F), ammonium 7-fluoro-2,1,3-benzoxadiazole-4-sulfonate (SBD-F), 4-hydrazino-7-nitro-2,1,3-benzoxadiazole (NBD-H), 4-N,N-dimethylaminosulfonyl-7-hydrazino-2,1,3-benzoxadiazole (DBD-H), 4-nitro-7-N-piperazino-2,1,3-benzoxadiazole (NBD-PZ), 4-N,N-dimethylaminosulfonyl-7-N-piperazino-2,1,3-benzoxadiazole (DBD-PZ), 4-(N-chloroformylmethyl-N-methyl)amino-7-N,N-dimethylaminosulfonyl-2,1,3-benzoxadiazole (DBD-COCl) and 7-N,N-dimethylaminosulfonyl-4-(2,1,3-benzoxadiazolyl) isothiocyanate (DBD-NCS) are reviewed in terms of synthetic method, reactivity, fluorescence characteristics, sensitivity and application to analytes.