A Modified Method for the Determination of Plasma Total Homocysteine by High Performance Liquid Chromatography

Serhat TOKGÖZ

Makale detayları

A Modified Method for the Determination of Plasma Total Homocysteine by High Performance Liquid Chromatography

Yazarlar:

Serhat TOKGÖZ

Yayıncı:

TUBITAK

Dergi:

Turkish Journal of Medical Sciences 2003 Vol.33 Issue 2, pp.71-76

Belge Tipi:

Makale

Yayın Tarihi:

1/2003

Belge Dili:

Türkçe

Document ID (Org No.):

2000756801

Sıklık:

İki Aylık

eISSN:

1303-6165

pISSN:

1300-0144

Anahtar kelimeler:

Özet:

 

We have modified a high-performance liquid chromatography (HPLC) procedure for homocysteine assay using 7-fluoro-2, 1, 3-benzoxadiazole-4-sulfonamide (ABD-F) pre-column derivatization. In this study,  the thiol compounds were liberated from plasma proteins by reduction with 2-mercaptoethanol (BME) and derivatized with a thiol-specific fluoregenic marker,  ABD-F. The derivatives were separated by reversed-phase HPLC on a Hypersil C 18 column (4.6 x 150 mm; 5 µ) using isocratic elution with 0.1 M phosphate buffer pH 6.0 containing 8% methanol and fluorescence detection. Excitation and emission wavelengths were 386 and 516 nm. The peak of homocysteine was eluted at 413 ± 25 s. Mean within-day and between-day precisions were determined to be 1.96 and 4.34% CV,  respectively. The method was linear up to 100 µmol/L and proved to be sensitive with a detection limit of 1 µmol/L for homocysteine.  In plasma samples from healthy adult subjects,  the concentration of homocysteine was found to be higher in men than in women (11.69 ± 2.9 versus 9.45 ± 1.78 µmol/L,  p  =  0.002). The method is simple,  sensitive,  and reproducible and allows a rapid determination of total homocysteine in human plasma under routine conditions.

 

We have modified a high-performance liquid chromatography (HPLC) procedure for homocysteine assay using 7-fluoro-2, 1, 3-benzoxadiazole-4-sulfonamide (ABD-F) pre-column derivatization. In this study,  the thiol compounds were liberated from plasma proteins by reduction with 2-mercaptoethanol (BME) and derivatized with a thiol-specific fluoregenic marker,  ABD-F. The derivatives were separated by reversed-phase HPLC on a Hypersil C 18 column (4.6 x 150 mm; 5 µ) using isocratic elution with 0.1 M phosphate buffer pH 6.0 containing 8% methanol and fluorescence detection. Excitation and emission wavelengths were 386 and 516 nm. The peak of homocysteine was eluted at 413 ± 25 s. Mean within-day and between-day precisions were determined to be 1.96 and 4.34% CV,  respectively. The method was linear up to 100 µmol/L and proved to be sensitive with a detection limit of 1 µmol/L for homocysteine.  In plasma samples from healthy adult subjects,  the concentration of homocysteine was found to be higher in men than in women (11.69 ± 2.9 versus 9.45 ± 1.78 µmol/L,  p  =  0.002). The method is simple,  sensitive,  and reproducible and allows a rapid determination of total homocysteine in human plasma under routine conditions.

Makale detayları

A Modified Method for the Determination of Plasma Total Homocysteine by High Performance Liquid Chromatography

Yazarlar:

Serhat TOKGÖZ

Yayıncı:

TUBITAK

Dergi:

Turkish Journal of Medical Sciences 2003 Vol.33 Issue 2, pp.71-76

Belge Tipi:

Makale

Yayın Tarihi:

1/2003

Belge Dili:

Türkçe

Document ID (Org No.):

2000756801

Sıklık:

İki Aylık

eISSN:

1303-6165

pISSN:

1300-0144

Homocysteine

High performance liquid chromatography

ABD-F

Sulfamoylbenzofurazan

Mercaptoethanol

Sayı

Makale detayları

A Modified Method for the Determination of Plasma Total Homocysteine by High Performance Liquid Chromatography

Yazarlar: Serhat TOKGÖZ

Yayıncı: TUBITAK

Dergi: Turkish Journal of Medical Sciences 2003 Vol.33 Issue 2, pp.71-76

Belge Tipi: Makale

Yayın Tarihi: 1/2003

Belge Dili: Türkçe

Document ID (Org No.): 2000756801

Sıklık: İki Aylık

eISSN: 1303-6165

pISSN: 1300-0144

Homocysteine

High performance liquid chromatography

ABD-F

Sulfamoylbenzofurazan

Mercaptoethanol

Sayı

Yazarlar:

Serhat TOKGÖZ

Yayıncı:

TUBITAK

Dergi:

Turkish Journal of Medical Sciences 2003 Vol.33 Issue 2, pp.71-76

Belge Tipi:

Makale

Yayın Tarihi:

1/2003

Belge Dili:

Türkçe

Document ID (Org No.):

2000756801

Sıklık:

İki Aylık

eISSN:

1303-6165

pISSN:

1300-0144