VMA, HVA and 5-HIAA (Urine) HPLC

VMA-5-HIAA-HVA (Urine) For HPLC

INTENDED USE

KR138 Reagent kit for the HPLC determination of VMA-5-HIAA-HVA in urine.

KIT SPECIFICATION

Contents for 100 analyses

ACCESSORIES

CONTROLS

INTRODUCTION

Catecholamines are dihydroxyl organic compounds that are characterized by a phenolic ring.
Epinephrine (E), Norepinephrine (NE), and Dopamine are the most significant members of this family.
Catecholamines’s biosynthetic pathway uses L-tyrosine as initial substratum. Chromaffin cells synthesize and store epinephrine in the adrenal medulla, while norepinephrine production happens in the sympathetic nerve endings. Dopamine is above all a neurotransmitter in the CNS.

CATABOLISM

The biological effects of catecholamines terminate rapidly by uptake into the symphathetic nerve endings. The major changes that occur in these sites contain their transformation into meta-O-methylated and deaminated metabolites due to Cathechol-O-methyltransferase (COMT) and monoamine Oxidase (MAO) respectively and, finally, their conjugation with sulfate and glucuronide.
Homovanillic Acid is the major metabolite of Dopamine while Vanillymandelic Acid is the main metabolite of Norepinephrine and Epinephrine.

CLINICAL BACKGROUND

Catecholamines are often dsignated in urine for neurological diagnosis and for observing the reply to therapy in illnesses like pheochromocytoma and neuroblastoma.

Pheochromocytoma is a catecholamine-producing tumor multiplied from adrenomedullary chromaffin cells. More than 90% appear to be benign. Because of their capacity to store and dismiss catecholamines, they are hazardous.

EQUIPMENT AND INSTRUMENTAL PARAMETERS

The analysis of VMA-5-HIAA-HVA requires a HPLC including gradient system with HPLC pump, autosampler, autoinjector and FLD detector.

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

Sampler: Injection volume 25 μl, analytical run time 22 min..
HPLC pump: Flow rate 0.9 ml/min is recommended
Column temperature: Ambient (~25 °C)
FLD detector: Ex:280,Em:330

COLLECTION AND STORAGE OF PATIENT SAMPLES

The sample material is 24 hours urine. Storage life at room temperature is about 1 day, refrigerated (+2 to +8 °C) up to 1 week and frozen below -20 °C up to 6 months.

SAMPLE PREPARATION PROCEDURE

50 µl urine sample is added to vial with teflon cap

500 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 24 hours, at +2 to +8 °C up to 36 h, frozen below -18 °C up to 3 months.