MASS SPECTROMETRIC DETERMINATION OF RETINOL IN EMIRATI POPULATION

This thesis is concerned with analyzing vitamin A deficiency. Vitamin A is crucial for several functions, including vision, growth, and cell differentiation. the main aim is to develop and validate an LC-MS/MS method for the determination of vitamin A in plasma matrix and use the new method for the determination of vitamin A levels in the obese Emirati population. Methods: A positive ion electrospray ionisation (ESI) LC-MS/MS method was used in the Multiple Reaction Monitoring (MRM) mode for quantification. It involved i) LC-MS/MS, ii) a guard column together with C18 Ascentis Express F5 column iii) Internal standard, 25-Hydroxyvitamin D3 (6, 19, 19-d3), and iv) identification via ESI and monitoring of three fragmentations of the parent ion. To demonstrate the practical value of this method, blood samples were collected from 452 Emirati participants (277 obese; 175 healthy). The method was validated according to FDA-US guidelines. The method has been applied to a sample of 277 Emirati obese patients, including 277 baselines, 277 follow-ups, and 175 health samples. The characteristics of the sample included: mixed-gender with 73 males in the age group of (18–82 years) and 204 females in the age group of (18–65 years). The results show that the concentrations of vitamin A for both females and males increased after supplementation (372 ng/mL to 440 ng/mL for the female samples, and 438 ng/mL to 540 ng/mL for the male sample). This has led to a sufficient level of vitamin A in participants. The new method allowed chromatographic separation and quantification of vitamin A. The new assay could detect 0.48 ng/mL of vitamin A in serum with the calibration curve ranging from 7.8 to 1000 ng/ml. The method validation parameters, including intra and inter-day precision, intra and inter-day accuracy, recovery, linearity, specificity, and stability, were within range. For example, the recovery percentage found were 99%, 96%, and 94% for QCH, QCM, and QCL respectively, while the percentage of change in the stability of vitamin A ranges between 0 and 3%. The applied LC-MS/MS method was intended to accurately detect Vitamin A in human plasma and has proven to be specific, reliable, and robust. The method can detect low levels of vitamin A. This analytical method does not require time-consuming derivatization and complex extraction techniques and could prove very useful in clinical studies