Determination of serum D-lactic and L-lactic acids in normal subjects and diabetic patients by column-switching HPLC with pre-column fluorescence derivatization

Hisanori Hasegawa, Takeshi Fukushima, Jen Ai Lee, Kazuhisa Tsukamoto, Kyoji Moriya, Yoshikazu Ono, Kazuhiro Imai

Research output: Contribution to journalArticle

50 Citations (Scopus)

Abstract

d-Lactic and l-lactic acids were simultaneously determined by means of a column-switching high-performance liquid chromatography (HPLC) with fluorescence detection. As a fluorescence reagent, 4-nitro-7-piperazino-2,1,3-benzoxadiazole (NBD-PZ) was employed for the fluorescence derivatization of lactic acid. The proposed HPLC system adopted both octylsilica (Cadenza CD-C8) and amylose-based chiral columns (CHIRALPAK AD-RH), which proved to give a sufficient enantiomeric separation of the lactic acid derivatives with a separation factor (α) of 1.32 and a resolution (R s) of 1.98. Moreover, the features of the first elution of d-lactic acid peak in the proposed HPLC were convenient for the determination of trace amount of serum d-lactic acid, which is known to increase under diabetes. Intra-day and inter-day accuracies were in the range of 90.5-101.2 and 89.0-100.7%, and the intra-day and inter-day precisions were 0.3-1.2 and 0.4-4.8%, respectively. The proposed method was applied to determine d-lactic and l-lactic acids in human serum of normal subjects and diabetic patients, showing that both d-lactic and l-lactic acid concentrations were significantly increased in the serum of diabetic patients (n=31) as compared with normal subjects (n=21). This fact was found for the first time owing to the development of the proposed HPLC method which is able to determine d-lactic and l-lactic acid simultaneously. Finally, serum d-lactic acid concentrations determined by the proposed HPLC method were compared with those from a reported enzymatic assay, and the smaller p value between normal subjects and diabetic patients was shown by the proposed HPLC method.

Original languageEnglish
Pages (from-to)886-891
Number of pages6
JournalAnalytical and Bioanalytical Chemistry
Volume377
Issue number5
DOIs
Publication statusPublished - Nov 2003

Fingerprint

High performance liquid chromatography
Lactic Acid
Milk
Fluorescence
High Pressure Liquid Chromatography
Serum
Amylose
Enzyme Assays
Medical problems
Assays
Reference Values
Derivatives

Keywords

  • Amylose-based chiral column
  • Column-switching HPLC
  • D-Lactic acid
  • Diabetic patients
  • L-Lactic acid
  • NBD-PZ

ASJC Scopus subject areas

  • Analytical Chemistry
  • Clinical Biochemistry

Cite this

Determination of serum D-lactic and L-lactic acids in normal subjects and diabetic patients by column-switching HPLC with pre-column fluorescence derivatization. / Hasegawa, Hisanori; Fukushima, Takeshi; Lee, Jen Ai; Tsukamoto, Kazuhisa; Moriya, Kyoji; Ono, Yoshikazu; Imai, Kazuhiro.

In: Analytical and Bioanalytical Chemistry, Vol. 377, No. 5, 11.2003, p. 886-891.

Research output: Contribution to journalArticle

Hasegawa, Hisanori ; Fukushima, Takeshi ; Lee, Jen Ai ; Tsukamoto, Kazuhisa ; Moriya, Kyoji ; Ono, Yoshikazu ; Imai, Kazuhiro. / Determination of serum D-lactic and L-lactic acids in normal subjects and diabetic patients by column-switching HPLC with pre-column fluorescence derivatization. In: Analytical and Bioanalytical Chemistry. 2003 ; Vol. 377, No. 5. pp. 886-891.
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AU - Hasegawa, Hisanori

AU - Fukushima, Takeshi

AU - Lee, Jen Ai

AU - Tsukamoto, Kazuhisa

AU - Moriya, Kyoji

AU - Ono, Yoshikazu

AU - Imai, Kazuhiro

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KW - Diabetic patients

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