کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
6488063 | 1416354 | 2018 | 8 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
A kinetic spectrophotometric method for the determination of pyridoxal-5â²-phosphate based on coenzyme activation of apo-d-phenylglycine aminotransferase
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موضوعات مرتبط
مهندسی و علوم پایه
مهندسی شیمی
بیو مهندسی (مهندسی زیستی)
پیش نمایش صفحه اول مقاله
چکیده انگلیسی
A new PLP assay method based on the coenzyme activation of apo-d-phenylglycine aminotransferase (apo-d-PhgAT) is reported. The assay process is comprised of two steps. First, PLP present in plasma samples is allowed to reconstitute apo-d-PhgAT, forming active holo-d-PhgAT. In the second step, the enzymatic activity of reconstituted d-PhgAT is determined using d-4-OH-phenylglycine as the amino donor substrate with 4-OH-benzoylformate (OH-BZF) as the reaction product. OH-BZF absorbs UV light strongly at 334â¯nm (molar absorption coefficientâ¯=â¯25.4â¯Ãâ¯103 Mâ1cmâ1) and its rate of formation is monitored spectrophotometrically. The rate of the transamination reaction catalyzed by the reconstituted d-PhgAT is directly proportional to the amount of PLP in the sample. The method is applicable for determining PLP in the concentration range from 5.2 to 250â¯nM and requires 50â¯Î¼L of plasma sample. The mean within- and between-run coefficient of variations (CVs) were 8.1% and 12.4%, respectively. Analytical recoveries ranged from 98 to 108%. The assay was specific and showed good correlation with the established method (CDC, Method No: 4002.05). The assay requires one reaction catalyzed by a single enzyme, does not require a radioactive substrate, and a derivatization reagent is not needed. This PLP determination process is relatively simple to perform and can be completed using common laboratory equipment.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Enzyme and Microbial Technology - Volume 117, October 2018, Pages 64-71
Journal: Enzyme and Microbial Technology - Volume 117, October 2018, Pages 64-71
نویسندگان
Juntratip Jomrit, Duangnate Isarangkul, Pijug Summpunn, Suthep Wiyakrutta,