Comparing LC/MS with LC/MS/MS for the Simultaneous Extraction and Analysis of Water-Soluble Vitamins and Fat-Soluble Vitamins in Multivitamin Supplements

Importance of the topic

Dietary supplements are consumed by about half of adults in the USA. Multivitamin formulations are among the most popular. Precise quantification of both water soluble and fat soluble vitamins is critical to ensure product quality, regulatory compliance and consumer safety.

Objectives and study overview

This study evaluates and compares a single extraction workflow combined with either single quadrupole or triple quadrupole mass spectrometry for the simultaneous analysis of six water soluble B vitamins, vitamin C, and three fat soluble vitamins in gummy multivitamins. The goal is to assess sensitivity, selectivity, matrix effects and overall throughput.

Methodology and instrumentation

Sample preparation involves dissolving a weighed gummy in acidic aqueous solvent with acetonitrile, heating, adding ethanol, shaking and centrifugation followed by dilution. Chromatography uses an Agilent 1290 Infinity II UHPLC system with a Poroshell 120 Phenyl Hexyl column (3.0 x 100 mm, 2.7 µm) at 0.5 mL/min and a gradient of ammonium formate and formic acid in water against formic acid in methanol. Mass spectrometric detection was performed in positive electrospray ionization on an Agilent iQ LC MS in SIM mode and on an Agilent 6470 triple quadrupole LC MS MS in dynamic MRM mode. Key source and gas parameters were optimized to maximize sensitivity.

Main results and discussion

The Phenyl Hexyl column achieved clear separation of all analytes. MRM mode delivered superior selectivity and sensitivity, while SIM improved detection over UV methods but offered lower specificity. Recovery studies in a gummy matrix showed post spike recoveries of 91 to 125 for most vitamins, with vitamin D3 corrected using a deuterated internal standard to address lower recovery. Calibration curves met acceptance criteria of r2 ≥ 0.99, ion ratio consistency within 30 and coelution of analyte with internal standard.

Benefits and practical applications

• A unified extraction and LC MS workflow reduces assay complexity and cost by analyzing multiple vitamins in one run
• High throughput and rapid turnaround support quality control and regulatory testing
• Post matrix spike correction compensates for matrix effects ensuring accurate quantification
• MRM provides the highest confidence in complex supplement matrices

Future trends and potential applications

Advances may include integration of high resolution mass spectrometry, automation of sample preparation, expansion to a wider range of supplement formats, and implementation of machine learning for data interpretation and pattern recognition.

Conclusion

Combining UHPLC with single or triple quadrupole mass spectrometry enables comprehensive, reliable, and efficient profiling of water and fat soluble vitamins in multivitamin supplements. The validated method using MRM delivers outstanding sensitivity, selectivity, and accuracy, confirming compliance of the tested gummy product with label claims.