Determination of short-chain acyl-coenzyme A esters by high-performance liquid chromatography

Yoshitaka Hosokawa, Yoshiharu Shimomura, Robert Harris, Takayuki Ozawa

Research output: Contribution to journalArticle

46 Citations (Scopus)

Abstract

A method for the determination of short-chain acyl-CoA esters in tissue extracts by HPLC has been developed. The acyl-CoA esters were extracted from freeze-clamped rat livers with perchloric acid. The extract was applied to a Sep-Pak C18 cartridge. The cartridge was washed with acidic water, pH 3, followed by petroleum ether, chloroform, and methanol. Then the acyl-CoA esters were eluted from the cartridge with ethanol/water (65:35) containing 0.1 m ammonium acetate. By this procedure, the acyl-CoA esters were concentrated and partially purified. The eluate was analyzed by HPLC using reverse-phase columns of Develosil ODS (0.46 × 15 cm plus 0.46 × 25 cm). The separation of the acyl-CoA esters was conducted with a linear gradient (1.75 to 10%) of acetonitrile. The CoA compounds (malonyl-CoA, succinyl-CoA plus CoASH, methylmalonyl-CoA, 3-hydroxy-3-methylglutaryl-CoA, acetyl-CoA, acetoacetyl-CoA, and propionyl-CoA) were identified and determined by monitoring at 260 nm. Isobutyryl-CoA was used as an internal standard, since the content of this CoA ester was negligible in livers from rats with several metabolic conditions. The lower limit of detection of individual acyl-CoA esters was approximately 50 pmol. Using this analytical method, short-chain acyl-CoA esters were determined in livers from normal and fasted rats.

Original languageEnglish
Pages (from-to)45-49
Number of pages5
JournalAnalytical Biochemistry
Volume153
Issue number1
DOIs
StatePublished - Feb 15 1986

Fingerprint

Acyl Coenzyme A
High performance liquid chromatography
Esters
High Pressure Liquid Chromatography
Coenzyme A
Liver
Rats
Malonyl Coenzyme A
Acetyl Coenzyme A
Tissue Extracts
Water
Chloroform
Methanol
Limit of Detection
Ethanol
Monitoring

Keywords

  • acyl-CoA esters
  • HPLC
  • Sep-Pak C cartridge

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology

Cite this

Determination of short-chain acyl-coenzyme A esters by high-performance liquid chromatography. / Hosokawa, Yoshitaka; Shimomura, Yoshiharu; Harris, Robert; Ozawa, Takayuki.

In: Analytical Biochemistry, Vol. 153, No. 1, 15.02.1986, p. 45-49.

Research output: Contribution to journalArticle

Hosokawa, Yoshitaka ; Shimomura, Yoshiharu ; Harris, Robert ; Ozawa, Takayuki. / Determination of short-chain acyl-coenzyme A esters by high-performance liquid chromatography. In: Analytical Biochemistry. 1986 ; Vol. 153, No. 1. pp. 45-49.
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