Assay for Citrate and Phosphate in Pharmaceutical Formulations Using a High-Pressure Compact Ion Chromatography System

Importance of the Topic

Accurate measurement of citrate and phosphate in pharmaceutical products is essential for ensuring medication stability, efficacy and patient safety. These anions function as buffering agents, flavor enhancers and anticoagulants, making reliable analytical methods critical for manufacturing quality control and regulatory compliance.

Objectives and Overview of the Study

The study aimed to modernize a previously published ion chromatography (IC) method (Application Note 164) by employing a Thermo Scientific™ Dionex™ Integrion™ HPIC™ system combined with a Dionex IonPac™ AS11-HC-4 µm microbore column. The primary goals were to halve the analysis time, improve sensitivity and reduce reagent consumption while maintaining or enhancing method performance in citrate and phosphate assays.

Methodology and Instrumentation

The method uses an electrolytically generated 60 mM KOH eluent at 0.35 mL/min on a 2 × 250 mm analytical IonPac AS11-HC-4 µm column with a 2 × 50 mm guard. Detection is by suppressed conductivity (AERS 500 suppressor in recycle mode at 52 mA). Key settings include a column temperature of 35 °C, detector compartment at 20 °C and a 2.5 µL injection volume. Standards and samples are prepared in 1 mM NaOH and deionized water. Calibration spans 0.5–200 mg/L for phosphate and citrate (0.5–50 mg/L linear region), and limits of quantitation (LOQs) are 0.03 mg/L and 0.06 mg/L, respectively.

Instrumentation

• Dionex Integrion HPIC system with high-pressure pump (≤ 5000 psi), column oven, detector compartment temperature control and tablet interface.
• Dionex AS-AP autosampler with 10 mL trays.
• EGC 500 KOH eluent generator cartridge, CR-ATC 600 trap column and HP EG degasser.
• Dionex AERS 500 electrolytically regenerated suppressor (2 mm, recycle mode).
• Dionex IonPac AS11-HC-4 µm analytical and AG11-HC-4 µm guard columns (2 mm format).

Main Results and Discussion

The updated method achieved baseline separation of citrate and phosphate within 5 min, reducing run time by 50% versus the original. Eluent consumption dropped from 20 mL to 1.75 mL per run. Calibration curves were linear or quadratic with correlation coefficients ≥ 0.9997. Intraday precision RSDs were below 0.6% (citrate) and 0.34% (phosphate); interday RSDs were 1.25% and 0.75%, respectively. Recoveries in spiked samples ranged from 95.2% to 105.5%. Robustness tests (± 10% variation in flow rate, temperature and eluent concentration) maintained adequate resolution (> 1.5) and minimal assay bias (< 2%). Analysis of a citrate-phosphate-dextrose solution confirmed assay accuracy against USP specifications.

Benefits and Practical Application

• Halved analysis time increases sample throughput and laboratory efficiency.
• Reduced eluent and sample volumes lower operating costs and waste.
• Enhanced sensitivity and precision support tighter quality control limits.
• Integrated system design with consumable tracking improves GMP compliance and ease of use.

Future Trends and Opportunities

Advances in high-pressure IC instrumentation and miniaturized columns are likely to further decrease analysis times and solvent consumption. Integration with automated data analytics and remote monitoring will streamline method validation and troubleshooting. Expanding electrolytic eluent generation to more complex gradients and coupling IC with mass spectrometry may broaden the range of detectable compounds in pharmaceutical and biological matrices.

Conclusion

The application demonstrates a robust, fast and sensitive IC method for simultaneous quantitation of citrate and phosphate in pharmaceutical formulations. The combination of the Integrion HPIC system and AS11-HC microbore column offers significant time and cost savings while meeting stringent performance criteria for precision, accuracy and robustness.

References

1. Thermo Scientific Application Note 164: Assay for Citrate and Phosphate in Pharmaceutical Formulations Using Ion Chromatography, 2004.
2. DeBorba B.M., Rohrer J.S., Bhattacharyya L.: J. Pharm. Biomed. Anal. 2004, 36, 517–524.
3. United States Pharmacopeia General Chapter <345>: Assay for Citric Acid/Citrate and Phosphate, USP 29–NF 24, 2006.
4. Thermo Scientific Application Proof Note 196: Assay for Citrate and Phosphate Using a Compact IC System, 2016.
5. United States Pharmacopeia General Chapter <1225>: Validation of Compendial Methods, USP 34, 2011.
6. Thermo Scientific Application Update 200: Fast Anion Determinations in Environmental Waters, 2016.
7. Thermo Scientific Technical Note 175: Configuring the Integrion HPIC System, 2016.
8. Anticoagulant Citrate Phosphate Dextrose Solution, USP 29 Monograph, 2016.