OF SERUM VITAMIN D IN PRE MENOPAUSAL INDIAN WOMEN”

immunoassay technique. Result: 26 ± 8.5ng/mL was the biological reference interval (BRI) for Vitamin D, established by NIH but the in-house obtained BRI was 18.2 11.5ng/mL. The obtained BRI was significantly decreased ( p <0.001) than NIH established values. Conclusion : 18.2 ± 11.5ng/mL of Serum Vitamin D was observed in the adult premenopausal Indian sub population.

Vitamin D promotes absorption of calcium in the intestine as well as renal tubules and maintains adequate levels of calcium and phosphate in serum to maintain normal bone mineralization and prevents osteoporosis. This vitamin can be produced endogenously when ultraviolet (UV) rays in sunlight hit the skin and promotes its synthesis [1]. Deficiency of this vitamin is common in India, the rate of prevalence range from 40 -90 percent [2]! Geographically India is partly tropical and subtropical and receives sunlight round the year. More than 30ng/mL was the biological reference interval for vitamin D but it was derived from western population and it is using by the diagnostic laboratories across the country. Because of this scale majority of healthy individuals shows insufficient Vitamin D levels [3]. Objectives: To assess the status of vitamin D in the premenopausal women. To evolve a reference range in Indian sub-population and to correlate the findings with the reference interval established by National institutes of health (NIH), USA. Methodology: A cross sectional study consisting of 127 adult premenopausal women (29 -45 years of age) was conducted. Their total Vitamin D levels were measured by Chemiluminescence immunoassay technique.
Result: 26 ± 8.5ng/mL was the biological reference interval (BRI) for Vitamin D, established by NIH but the in-house obtained BRI was 18.2 ± 11.5ng/mL. The obtained BRI was significantly decreased (p <0.001) than NIH established values. Conclusion: 18.2 ± 11.5ng/mL of Serum Vitamin D was observed in the adult premenopausal Indian sub population.

…………………………………………………………………………………………………….... Introduction:-
India is well-known for its tradition, culture and lingual diversity and it is a wide tropical country located from 8.4º N to 37.6° N latitude and 68.7 o E to 97.25 o E longitude [4]. Deficiency of Vitamin D (Vit.D) in Indians is expected to be uncommon because the population gets the adequate sun light throughout the year [5]. However from the data available in the published literature, Vit.D deficiency is common in India in all the age groups and both the genders across the country [6,7]. Currently in the country, there is no health program that fulfills the specific health need of premenopausal women. Around 103 million Indian women are expected to attain post menopausal state by the year 2026 [8]. Therefore there is an urgent need to focus on the pre menopausal women health care. The present study is therefore expected to unmask the magnitude of suffering due to health issues in pre menopausal women associated with vit.D status. Because of the health risks associated with low levels of vit.D in postmenopausal women, the

Specimen handling and analysis
The blood specimen received in the laboratory was centrifuged (1800 x g /15mins) to separate the cellular components and the cell free serum processed for the analysis of routine biochemical parameters sought by the treating clinicians. Remaining specimens were aliquoted, labeled and stored at -20ºC till further analysis. The Aliquots of samples, once thawed were analyzed on the same day. Repeated freezing and thawing was not done to avoid any pre-analytical variables.

Type of study:
Randomized control study

Research design: Non Interventional
Analytical Method: Total serum Vit.D level was estimated by Chemiluminescence immunoassay technique.

Sources of Data
Available literature information from recent publications was updated during the course of study. The study design to be employed was standardized /modified depending upon the situation before applying the same for the sample analysis. Information with respect to study outcome was procured from the patient medical records and clinical expertise opinion was sought before relating the study outcome.

Statistical analysis:
SPSS Version 19 Data analysis package was used and applied to analyze the obtained data after discussion with the Biostatistician. The values were expressed in terms of mean ± SD. For the statistical comparison Student "t" test was used. The student"s "t" test *p< 0.05; **p< 0.01; ***p< 0.001 was considered as significant.
Analytical Method: Device Description: ADVIA centaur -XP -Chemiluminescence immunoassay system was used for the estimation of Vit.D levels. Assay Protocols Performance characteristics : The assay was highly specific and sensitive for the estimation. Sensitivity and Detection limit : 4.2 -150 ng/mL Specificity : 0.3 % cross reactivity Precision : It was evaluated according to the CLSI guidelines.
Quality control: Randox company controls were used. Two levels of controls were run (normal and abnormal) before analyzing the samples. Values of control samples were with in acceptable limits (± 2SD).
Procedural steps: 1. Load primary tube container into rack, ensuring that the barcode labels were clearly visible through the slot in the rack. 2. Place the rack in the sample compartment. 3. Press/clik the Start option. 4. The instrument was fully automated and analyzes the sample automaticly. 5. Values released from the instrument were interphased through LIS. 6. The values were noted.

Results:-
The obtained serum Vit.D Reference interval was 18.2 ± 11.5 ng/mL, which was significantly lower (***p < 0.001) compared to the reference interval established by NIH ie 26 ± 8.5ng/mL.  Thus, in present study measured values of vit.D in premenopausal women was compared with NIH reference interval (Fig 1). The observed finding suggests that, the measured Vit.D in premenopausal women significantly lower (***p < 0.001) than the NIH reference interval .   Fig 2:-Histogram represents the Vitamin D reference interval of premenopausal women from 3 sets age group Expressed as Mean ± SD. Student"s "t" test: *: p < 0.05; **: p < 0.01: ***: p < 0.001.

Serum vitamin D level in Premenopausal women
In the present study, the premenopausal healthy subjects (n = 117) have provided the reference interval of vit.D for Indian sub-population with distinct aged premenopausal groups range from 29 -45 years. Many factors involves in vit.D normal level including age, gender and body mass index(BMI). Hence, to minimize the bias the healthy subjects aged 29 -45years were further sub-divided into 3 groups (29 -35, 36 -40 and 41 -45 years). According to Fig 2, the histograms represents as 18.5 ± 12.1ng/mL, 17.6 ± 9.6ng/mL and 18.6 ± 13.1ng/mL between the 3 group respectively. The age related analysis did not showed any significant difference among the three groups.

Discussion:-
Serum Vit.D plays an important role in number of biological processes. Its deficiency also causes serious consequences. Inadequacy of the vitamin is the primary dilemma and inclined to osteoporosis in post menopausal women, in recent dogma. Vit.D became the most often sought after investigation to confirm or to rule out the suspected osteoporosis cases.
Vit.D is a unique nutrient because its requirement is met from diet and skin. In the year 1922, Dr. EV. Mc Collum et al were able to discover to prove that existence of second fat soluble vitamin [11], he did not called the common name Vit.D but he called as the Ca++ depositing vitamin. Nearing to a century of its existence in the field of medicine, still it is under stern scrutiny of defining the sufficiency and insufficiency for its optimum functional role in human beings.
In the past, Vit.D deficiency was defined as the concentration if less than 10ng/mL in serum because both the Vit.D and Ca++ absorption decline at this threshold [12]. In the 2003, the World health organization defined, Vit.D insufficiency as-lower than 20 ng/mL in serum and normal as 20 to 30 ng/mL [11]. The Institute of Medicine (IOM) reiterates that recommendation in 2011. The Endocrine Society proposed an alternative, defining insufficiency as less than 30 ng/mL and normal as higher than 30 ng/mL [13]. Why does it matter if the recommended cutoff for insufficiency is 20 ng/ mL or 30 ng/ mL? The reason is that meeting this target serum Vit.D changes the Recommended Dietary Allowance (RDA). The present study made an attempt to establish the in-house normative data (reference interval) of premenopausal women Vit.D value and it was compared with the available reference interval.

Summary and Conclusion:-
There had been an ample increase in measuring serum vit.D over the past decade. Hence, there is an urgency in developing our own reference interval of Vit.D to arrive a good indicator of osteoporosis to rule out its sufficiency and insufficiency in Indian sub-population. Our study involves obtaining normative Vit.D data from premenopausal women sub-population from India. Our study once again underscores the essentiality of our own in-house laboratory reference interval data for Vit.D may help us in identifying the individuals who are at increased risk for frequent bone fractures. In the present premenopausal women Vit.D level did not showed any astounding level which is normally called as sufficiency (>30ng/mL) by all available literature. It clearly indicates in our study that the pre menopausal women had 18.2 ± 11.5ng/mL concentration of Vit.D.