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INTRODUCTION

Approaches to evaluate vitamin status:

1. Biochemical methods

2. Clinical evaluation3. Dietary methods

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Why Biochemical assessment?

Clinical evaluation effective in diagnosis of late-

stage vitamin defficiencies- involving physiologic

dysfunction or morphological changes.

Dietary evaluation- diets and food habits likely to

provide insufficient amounts of vitamin intake

Biochemical assessment- Detection of early-stage

vitamin defficiencies.

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BIOCHEMICAL ASSESSMENT-

DEFINITION

Biochemical assessment is the analysis of specific

enzymes / substances that are produced as a resultof chemical reactions in the body.

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REQUIREMENTS OF USEFUL

BIOCHEMICAL METHODS

Correlate with the rate of vitamin intake and

respond to deprivation of the vitamin

Relate to a meaningful period of time

Relate to normal physiologic function

Be measurable in an accessible specimen

Be technically feasible, reproducible and affordable

Have an available base of normative data.

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BIOCHEMICAL ASSESSMENT

OF VITAMIN D

SERUM ALKALINE PHOSPHATASE

SERUM 25- HYDROXY VITAMIN D

SERUM VITAMIN D3

SERUM 1,25-DIHYDROXY VITAMIN D

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BIOCHEMICAL ASSESSMENT

OF VITAMIN D

Over 40 metabolites of vitamin D

Very short half life in the circulation

Vitamin D

half life of almost 24 hours 25 hydroxy vitamin D: half life: 21-30 days

1,25 dihydroxy vitamin D: half life: 4-15 hrs

Measurement of 25 hydroxy Vitamin D is a better

indicator of Vitamin D stores- obtained from sunlight

- obtained from dietary sources.

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Why 25-hydroxy vitamin D?

25 OH-D circulates in nmol/L concentrations while 25-

dihydroxy Vitamin D circulates in pmol/L concentrations.

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MAIN METHODS TO ESTIMATE 25OHD are

1. Immunoassay

2. HPLC attached to fluoroscence

3. Mass spectrometry detection

4. Tandem MS

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IMMUNOASSAYS Immunoassay techniques are especially suited for

the analysis of compounds at low concentration and

in samples with little or no preparation, since their

detection limits are usually within nanogram to

picogram range

they are analytical methods based on the specificimmuno-reaction between an antibody(Ab) and an

antigen(Ag), for the determination of the amount of

either reactant in the solution.

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IMMUNOASSAYS:

automated and incorporated into large commercial

analyser systems- excellent functionality and ability

to measure large numbers of samples routinely

Weakness: issues with calibration, standardisation,

inability to quantify vitamin D2 and D3 separately-

they give an estimation of total 25-OHD

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HPLC High-performance liquid chromatography, HPLC, is

a chromatographic technique used to separate the

components in a mixture, to identify each

component, and to quantify each component.

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MASS SPECTROMETRY Mass Spectrometry is an analytical laboratory

technique to separate the components of a sample

by their mass.

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CONCLUSION Measurement of serum 25OHD is the best way of

estimating Vitamin D status

Assay used should be able to recognise all forms of

25OHD [D2 and D3]- Tandem MS more

preferable.

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REFERENCES Biochemical assessments of vitamins and trace

minerals Robert.A.Jacob and David. E.Milne

Vitamins : Fundamental Aspects in Nutrition

and Health (3rd Edition)

Combs, Gerald F.

Nutrition and Diet Research Progress : Vitamin

D Deficiency

Lerner, Vladimir Miodownik, Chanoch

Vitamin D and Bone Health: A Practical

Clinical Guideline for Patient Management:National Osteoporosis Society, UK

Nutritional Biochemistry of Vitamins: 2ndedition:

David.A.Bender.

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