Agilent ZORBAX Rapid Resolution High Definition 300SB-C8 Threaded Column

Importance of the Topic

ZORBAX RRHD 300SB-C8 threaded columns represent a critical advancement for ultra-high performance liquid chromatography (UHPLC), enabling rapid, high-resolution reversed phase separations. Their ability to withstand elevated pressures and aggressive mobile phases makes them especially valuable for pharmaceutical, proteomic, and industrial analyses where stable, reproducible performance is essential.

Objectives and Overview

This data sheet aims to describe the design, performance characteristics, operating guidelines, and practical applications of the Agilent ZORBAX Rapid Resolution High Definition 300SB-C8 column. Key goals include demonstrating column efficiency, mechanical stability under high pressure, and compatibility with low-pH and protein-rich sample matrices.

Methodology and Instrumentation

• Stationary Phase: Spherical 1.8 µm particles of porous silica (300 Å) bonded with sterically protected diisopropyl n-octadecylsilane (C8).
• Column Dimensions: 2.1 mm × 50 mm threaded format, packed by high-pressure slurry loading for uniform bed density.
• Instrument Platform: Agilent 1290 Infinity LC system, capable of up to 1200 bar operation.
• Detection and Conditions: UV detection at 254 nm; typical mobile phase 35/65 acetonitrile/water; flow rate 0.5 mL/min; ambient temperature (23 °C).
• Sample Preparation: Filtration through 0.2 µm filters to protect 0.5 µm column frits.

Main Results and Discussion

Quality control chromatograms demonstrate sharp, well-resolved peaks for small organic compounds (uracil, phenol, 4-chloro-nitrobenzene, naphthalene) with high plate counts and minimal tailing. The columns maintain performance up to 80 °C and at pressures up to 1200 bar, although optimal lifetime is achieved below 1000 bar and under pH limits of 0.8–8.0. The silica-based support tolerates aggressive mobile phases including low pH and ion-pair additives, making it well suited for intact protein and peptide mapping applications.

Benefits and Practical Applications

• High resolution and speed for small molecules, peptides, and proteins.
• Low background for mass spectrometry when coupled to sensitive detectors.
• Broad chemical compatibility with a variety of organic solvents and buffers.
• Enhanced column longevity through steric protection of bonded phase.

Future Trends and Potential Uses

Advances may include further miniaturization of particle sizes for even faster separations, expanded temperature and pH robustness for challenging biologics, and integration with high-resolution mass spectrometry workflows. Continued optimization of surface chemistries could yield improved selectivity for post-translational modifications and complex biomolecules.

Conclusion

Agilent ZORBAX RRHD 300SB-C8 threaded columns deliver exceptional speed, efficiency, and stability for high-throughput reversed phase separations. Their robust design supports a wide range of analytical challenges, from small molecule assays to detailed peptide and protein mapping, making them a versatile tool for modern laboratories.

References

1. Claessens HA, van Straten MA, Kirkland JJ. J Chromatogr A. 1996;728:259.
2. Snyder LR, Kirkland JJ. Introduction to Modern Liquid Chromatography. 2nd ed. John Wiley & Sons; 1979.
3. Snyder LR, Kirkland JJ, Glajch JL. Practical HPLC Method Development. 2nd ed. John Wiley & Sons; 1997.