VITAMIN C (Plasma/Serum) HPLC

Vitamın C (Plasma/Serum) For HPLC

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INTENDED USE

KR136 Reagent kit for the HPLC determination of Vitamın C in Plasma/Serum.

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KIT SPECIFICATION

Contents for 100 analyses

Cat no	Quantity	Reagents
KR1361	1000 mlx1	Mobile Phase
KR1362	100 mlx1	İnternal Standart
KR1363	0.5 mlx5	Serum Calibrator

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ACCESSORIES

KR1365	1 adet	HPLC column for the analysis of Vitamın C in Plasma/Serum.
KR1369	100 adet	Autosampler vial with teflon caps

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CONTROLS

KR1367	0.5 ml x5	Serum Control, Level I
KR1368	0.5 ml x5	Serum Control, Level II

INTRODUCTION

Vitamin C, called as L-ascorbate, is an essential nutrient for higher primates and a couple of other species. Ascorbate’s presence is needed for a series of essential metabolic reactions in plants and in all animals. Moreover, it is made by almost all organisms, humans being one notable expection. Scurvy is caused by the vitamin defiency. It is also widely used as a food additive.

The pharmacophore of Vitamin C is the ascorbate ion. In living organisms, ascorbate is an antioxidant, thanks to it, the body is protected against oxidative stress, and is a cofactor in several vital enzymatic reactions.

Vitamin C is purely the L-enantiomer of ascorbate; the opposite D-enantiomer has no physiological importance. Both of them are mirror images of the same molecular structure. When L-ascorbate, that is a strong reducing agent, conducts its reducing function, it is transformed to its oxidized form, L-dehydroascorbate. L-dehydroascorbate can then be reduced back to the active L-ascorbate form in the body by enzymes and glutathione.

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EQUIPMENT AND INSTRUMENTAL PARAMETERS

The analysis of 2,5 -hexanedıone requires a HPLC system including isocratic pump, autosampler and UV detector.

The use of a solvent degasser prevents air bubbles and ensure a stable baseline.

Sampler: Injection volume 20 μl, analytical run time 5 min..

HPLC pump: Flow rate 1.5 ml/min is recommended

Column temperature: Ambient (~25 °C)

FLD detector: 245 nm

COLLECTION AND STORAGE OF PATIENT SAMPLES

The stability for vitamin C in heparin plasma is 5 days at -20 °C.
At room temperature this is only up to two hours.

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SAMPLE PREPARATION PROCEDURE

100 µl sample is added to light protected vial with teflon cap

100 µl internal standard is added.

Vortex the mix for 10s.

Centrifuge at 14 000 rpm for 10 min.

Samples prepared as above can be stored at ambient temperature for about 12 hours, at +2 to +8 °C up to 36 h, frozen below -18 °C up to 3 months.