Analysis of Vitamin K

Importance of the topic

Fat-soluble vitamin K comprises key isoforms involved in blood coagulation and bone metabolism. Accurate separation and quantification of vitamin K1 and menaquinones are critical for pharmaceutical quality control, nutritional research, and clinical diagnostics.

Objectives and study overview

This study demonstrates a straightforward reversed-phase liquid chromatography method to separate and quantify menaquinone-4 (MK-4), vitamin K1, and menaquinone-7 (MK-7) at 5 mg/L each, using Shim-pack GIST C8 in an isocratic system.

Methodology and instrumentation

• Column: Shim-pack GIST C8 (150 mm × 3.0 mm I.D., 5 µm)
• Mobile phase: acetonitrile (isocratic)
• Flow rate: 1.0 mL/min
• Column temperature: 40 °C
• Detection: UV at 270 nm
• Injection volume: 5 µL

Main results and discussion

The method achieved baseline separation of MK-4, K1, and MK-7 with sharp, symmetric peaks and consistent retention times. The isocratic acetonitrile mobile phase provided stable elution and rapid column reequilibration. The assay demonstrated reliable quantification of each vitamin at the target concentration.

Benefits and practical applications

• A single-solvent isocratic run simplifies setup and reduces analysis time
• High reproducibility and sensitivity support routine QC workflows
• Applicable to pharmaceutical formulations and dietary supplements

Future trends and potential applications

Further developments may include coupling to mass spectrometry for enhanced selectivity, transitioning to UHPLC for higher throughput, implementing green solvent systems, and extending the method to complex biological matrices. Advances in column chemistry, such as chiral separations and microflow formats, could broaden analytical scope.

Conclusion

The Shim-pack GIST C8 reversed-phase LC method provides a robust, reproducible, and efficient means to analyze vitamins K1, MK-4, and MK-7. Its simplicity and performance make it well suited for routine laboratory applications.

Used instrumentation

The analysis employed a Shim-pack GIST C8 column coupled with a UV detector set at 270 nm, an isocratic HPLC pump, and column oven maintained at 40 °C.