Improving the Analysis of Betamethasone and Phosphate, Acetate, and Dipropionate Derivatives Using CORTECS™ Premier C8 Columns

Significance of the Topic

Accurate chromatographic analysis of steroids and their derivatives is critical in pharmaceutical quality control and method development. Betamethasone and its phosphate, acetate, and dipropionate forms can exhibit undesirable interactions with metal surfaces in standard LC hardware, leading to peak tailing, reduced sensitivity, and compromised data quality. Employing inert high-performance surfaces addresses these challenges and supports robust, future-proof analytical methods.

Objectives and Study Overview

This study compares the chromatographic performance of three C8 column configurations—CORTECS Premier C8 with MaxPeak High-Performance Surfaces (HPS), standard CORTECS C8, and a solid-core C8 column from a different supplier—for the separation of four analytes: betamethasone phosphate, betamethasone, betamethasone acetate, and betamethasone dipropionate. Key metrics included peak area, USP tailing factor, and peak width at 4.4% height.

Experimental Methodology

Stock solutions (1 mg/mL) were prepared in 1:1 water:acetonitrile and diluted to 50 µg/mL of each analyte in 12.5% acetonitrile. A generic gradient from 5% to 95% acetonitrile (with 0.1% formic acid) over 6.86 min was employed on an ACQUITY UPLC H-Class PLUS System. Data were acquired at 254 nm and processed using Empower CDS.

Instrumental Setup

• LC System: ACQUITY UPLC H-Class PLUS with six column positions, PDA, QDa, and HPS flow path
• Detector: UV at 254 nm
• Columns:
  • CORTECS Premier C8, 2.1 × 50 mm, 2.7 µm
  • CORTECS C8, 2.1 × 50 mm, 2.7 µm
  • Vendor A C8, 2.1 × 50 mm, 2.7 µm
• Column Temperature: 30 °C
• Sample Temperature: 10 °C
• Injection Volume: 1.0 µL
• Flow Rate: 0.5 mL/min
• Mobile Phases: A – water; B – acetonitrile; D1 – 2% formic acid in water (5% constant)
• Gradient: 5% to 95% B over 6.86 min, hold 1.14 min, re-equilibrate 2.28 min (total 10.30 min)

Main Results and Discussion

Betamethasone phosphate exhibited significantly narrower peaks, improved symmetry, and higher area on the CORTECS Premier C8 column compared to both stainless steel columns. The other three derivatives, lacking strong acidic moieties, showed comparable performance across all columns. Quantitative data for peak area, USP tailing, and peak width confirmed that inert HPS hardware uniquely benefits analytes prone to metal-surface interactions.

Benefits and Practical Applications

• Enhanced resolution of four steroid compounds under a generic gradient.
• Improved peak shape and sensitivity for phosphate derivative, crucial for trace quantification.
• Mitigation of metal-analyte interactions ensures consistent method performance and reduces the need for revalidation.

Future Trends and Applications

Expansion of inert surface technology to other column chemistries and full LC systems will broaden its impact on acidic and phosphorylated analytes. Integration with high-throughput and two-dimensional separations promises further gains in sensitivity. Continued development of hybrid materials and surface coatings may extend these benefits to proteomics, metabolomics, and other challenging applications.

Conclusion

MaxPeak Premier C8 columns with inert HPS hardware deliver superior chromatographic performance for betamethasone phosphate without compromising separations of non-acidic derivatives. Adoption of this technology enhances data quality, streamlines method development, and future-proofs analytical workflows.

References

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