Fat- and Water-Soluble Vitamin Analysis in Foods and Supplements (Consumable Workflow Ordering Guide)

Significance of the Topic

Vitamins play a critical role in human health, regulating metabolic processes, immune function, and cellular repair. Accurate measurement of both fat-soluble (A, D2, D3, E, K) and water-soluble (C and B-complex) vitamins in foods and supplements is vital to ensure dietary adequacy, comply with regulatory standards, and guarantee product quality.

Objectives and Study Overview

This guide presents optimized analytical workflows for the simultaneous detection and quantitation of 13 essential vitamins across complex matrices. It demonstrates methods for:

• Separating and measuring fat-soluble vitamins and carotenoids in food and supplement samples.
• Analyzing trace levels of vitamin D isomers (D2, D3) without baseline chromatographic separation.
• Resolving water-soluble vitamins and caffeine in fortified products.

Methodology and Sample Preparation

Sample preparation approaches address the lipophilic and hydrophilic nature of target analytes:

• Fat-Soluble Vitamins – Saponification followed by supported liquid extraction (Agilent Chem Elut S) or SPE (Bond Elut Plexa) to remove lipids and concentrate analytes.
• Water-Soluble Vitamins – Simple dissolution or dilution followed by filtration using Captiva syringe filters or filter vials.

Used Instrumentation

Key instrumentation and consumables include:

• HPLC-DAD systems for vitamins A and E quantification using InfinityLab Poroshell 120 PFP columns.
• Triple quadrupole LC/MS/MS with Jet Stream ESI source for trace analysis of vitamins D2, D3, and B-complex compounds using Phenyl-Hexyl or Aq-C18 columns.
• Chromatographic conditions optimized with formic acid/ammonium salts in water and organic solvents (methanol, acetonitrile).
• Consumables: Max-Light DAD flow cells, Quick Connect/QT fittings, Captiva RC filters, and standard screw-cap vials.

Main Results and Discussion

Fat-soluble vitamins A and E were effectively quantified at low ppm levels by HPLC-DAD, avoiding MS detector saturation. LC/MS/MS provided sub-ppb sensitivity for vitamin D isomers. Poroshell column chemistries allowed:

• Baseline separation of vitamins A, K, E, and carotenoids in under 11 minutes.
• Resolution of D2 and D3 isomers for accurate potency assessment.
• Simultaneous profiling of vitamin E isoforms (α, β, γ, δ) within 9 minutes.

Water-soluble vitamins and caffeine exhibited sharp, well-resolved peaks using Aq-C18 and Phenyl-Hexyl phases, enabling multiplexed quantitation in a single run.

Benefits and Practical Applications

The described workflows deliver:

• High throughput with reduced run times and simplified sample processing.
• Improved reproducibility through robust extraction and cleanup protocols.
• Regulatory compliance by meeting FDA GMP requirements for identity, purity, and strength.
• Versatility to handle food matrices (infant formula, eggs, fish) and supplements (tablets, gummies, drinks).

Future Trends and Opportunities

Emerging advances may include ultra-high-throughput LC/MS platforms, automation of sample prep, and novel stationary phases for even faster separations. Integration of on-line extraction and real-time data processing could further streamline vitamin analysis and expand applications in personalized nutrition and quality control.

Conclusion

The combination of targeted sample preparation, state-of-the-art chromatography, and sensitive detection provides a comprehensive solution for vitamin analysis in foods and supplements. Implementing these methods supports accurate labeling, consumer safety, and regulatory compliance.

References

1. Determination of Fat-Soluble Vitamins in Foods using Agilent Chem Elut S Extraction with LC/DAD and LC/MS/MS Triple Quadrupole, 5994-5063EN
2. Simultaneous Detection and Quantitation of 14 Fat-Soluble Vitamins and Carotenoids by LC/MS/MS Triple Quadrupole, 5994-5064EN
3. Fat-Soluble Vitamins Analysis on an Agilent ZORBAX Eclipse PAH Polymeric C18 Bonded Column, 5990-5342EN
4. Determination of Fat-Soluble Vitamins in Infant Formula Using Agilent Bond Elut Plexa Polymeric SPE with HPLC and LC/MS/MS, 5994-2948EN
5. Separation of Water-Soluble Vitamins on the Agilent InfinityLab Poroshell 120 Aq-C18 Column, 5994-5535EN
6. Simultaneous Detection and Quantitation of 14 Water-Soluble Vitamins in a Supplement by LC/MS/MS Triple Quadrupole, 5994-3016EN