Analysis of Water-Soluble Vitamins in Foods by LCMS-8060NX Triple Quadrupole Mass Spectrometer

Importance of the Topic

Water-soluble vitamins play critical roles in human health and are commonly added to fortified foods and infant formulas. Accurate measurement of multiple vitamins in a single run improves laboratory efficiency, reduces sample handling errors, and supports quality control in food and nutrition research.

Goals and Overview of the Study

The primary objective was to develop and validate a simultaneous quantitative LC-MS/MS method for four water-soluble vitamins—biotin, nicotinic acid, nicotinamide, and pantothenic acid—using the Shimadzu LCMS-8060NX triple quadrupole mass spectrometer. The method was applied to complex matrices, namely modified milk powder and milk raw material, to demonstrate robustness and accuracy.

Methodology and Instrumentation

Sample Preparation

• Acid hydrolysis of modified milk powders and milk raw material to release bound vitamins.
• Filtration and dilution to obtain clear sample solutions.

Chromatographic Conditions

• Column: L-column3 C18, 150 mm×2.0 mm I.D., 3 µm.
• Mobile phase A: 10 mmol/L ammonium bicarbonate in water; B: acetonitrile.
• Gradient: B 2% (0–3 min) to B 60% (4–5 min), B 98% (5.01–9 min), return to B 2% (9.01–13 min).
• Flow rate: 0.35 mL/min; column temperature: 40 °C; injection volume: 1 µL.

Used Instrumentation

• Shimadzu LCMS-8060NX triple quadrupole mass spectrometer with IonFocus ionization interface.
• Electrospray ionization in positive mode with probe voltage −0.5 kV and optimized probe position at 4.5 mm.
• Multiple reaction monitoring (MRM) for targeted quantification.
• Gas flows: nebulizing 3.0 L/min, drying 25.0 L/min, heating 15.0 L/min; DL/heat block temperatures 250 °C/300 °C; interface temperature 400 °C.

Main Results and Discussion

Calibration and Linearity

• Excellent linearity achieved with correlation coefficients (R) ≥0.998 for all vitamins across the ranges: biotin 0.102–254 ng/mL, nicotinic acid 0.5–500 ng/mL, nicotinamide 0.605–605 ng/mL, pantothenic acid 1.02–510 ng/mL.
• Accuracy within 80–120% and repeatability ≤20% confirmed at each calibration level.

Sample Analysis and Comparison

• Quantitative results for milk raw material showed close agreement with traditional bioassay values: biotin (56.2 vs. 60.8 ng/mL), total niacin (242.5 vs. 274.7 ng/mL), pantothenic acid (1.14 vs. 1.33 ng/mL).
• Chromatograms exhibited clear peaks with no significant matrix interferences due to efficient neutral particle removal by the IonFocus unit.

Recovery and Robustness

• Spike-and-recovery tests in four modified milk powders and raw material yielded recoveries between 52.9% and 115.6%, demonstrating method accuracy across different matrices.
• The IonFocus interface significantly reduced contamination of the mass spectrometer, ensuring system robustness over extended runs.

Benefits and Practical Applications

• Simultaneous detection of multiple vitamins reduces analysis time and labor compared to single-analyte methods.
• High sensitivity and selectivity enable trace-level quantification in complex food matrices.
• Robustness of the IonFocus unit minimizes downtime and maintenance in routine QA/QC laboratories.

Future Trends and Opportunities

Continued advances may include expansion to additional vitamins and micronutrients, integration with high-resolution mass spectrometry for untargeted profiling, and automation of sample preparation. Data analytics and machine learning could further enhance method development and interpretation in nutritional and food safety studies.

Conclusion

The developed LCMS-8060NX method with IonFocus technology provides a fast, sensitive, and reliable protocol for simultaneous quantification of key water-soluble vitamins in complex food matrices. Its high throughput and robustness make it well suited for research, quality control, and regulatory compliance in the food industry.

References

1. Kawashima M. Analysis of Water-Soluble Vitamins in Foods by LCMS-8060NX Triple Quadrupole Mass Spectrometer. Shimadzu Application News. First Edition: July 2021.
2. Shimadzu Corporation. LCMS and IonFocus Technology Manuals.