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Ultra High Performance Liquid Chromatography UPLC Beyond HPLC - The New Height of Chromatography Lab
Over the past three decades, high-performance liquid chromatography (HPLC) has become one of the most essential tools in laboratories worldwide. Its widespread use is due to its versatility and power, as it can handle a broad range of applications—from capillary to preparative scales. HPLC’s ability to detect various compounds makes it a go-to technique for many scientific fields. Today, the development of HPLC focuses on three main areas: chemistry, detectors, and data systems. These advancements have helped solidify HPLC’s role in modern science. However, there is still potential for further innovation in chromatographic techniques. So, what lies ahead for HPLC? How can future developments enhance its usefulness?
Enter UPLCTM, an advanced form of liquid chromatography that pushes the boundaries of traditional HPLC. Developed by researchers like Dr. J. Jorgensen and Dr. M. Lee, this technology uses particles smaller than 2 micrometers, leading to significant improvements in efficiency and speed. The Fandamt equation, which relates flow rate to column efficiency, plays a key role in understanding how these smaller particles improve performance.
As shown in Figure 1, reducing particle size below 2 μm leads to higher column efficiency without a drop in performance at higher flow rates. This allows for faster analysis, better resolution, and increased sensitivity. UPLCTM, or ultra-high performance liquid chromatography, represents a new era in separation science, where peak capacity—defined as the number of peaks separated per unit time—can be significantly enhanced.
Figure 2 illustrates the impact of using 1.7 μm particles instead of 5 μm ones. The results show a clear improvement in both resolution and peak capacity. Smaller particles allow for faster separations while maintaining high resolution, making UPLC ideal for complex samples.
Speed, sensitivity, and resolution are all improved with UPLC. Shorter columns and higher flow rates increase the speed of analysis, while smaller particles boost resolution. This combination allows for optimal performance in any given separation. As seen in Figure 3, UPLC can separate samples faster than traditional HPLC at the same level of resolution, while also using less solvent.
The challenge of producing and filling sub-2 μm particles into durable columns remains, but recent advances have made this possible. Porous particles are necessary to maintain sample loading and retention times, while the column design must accommodate smaller particles and ensure uniform packing. The inner surface of the column and the sieve plates must also be optimized to prevent blockages and maintain stability.
UPLC systems require higher pressure capabilities than standard HPLC instruments. For example, a 10 cm column with 1.7 μm particles can generate a pressure drop of around 15,000 psi. This demands pumps capable of delivering solvents under such conditions. Additionally, the solvent delivery system must account for compressibility effects, especially during gradient elution.
Sample injection is another critical component. UPLC requires injection systems that can withstand high pressures and minimize pulse interference. Small system volumes help reduce zone broadening, allowing for faster and more accurate injections. Low-volume injections also support higher sensitivity and reduce cross-contamination.
Detectors used in UPLC must have fast sampling rates to capture narrow peaks accurately. They also need minimal diffusion volume to preserve separation efficiency. For mass spectrometry, UPLC can significantly enhance sensitivity by concentrating analytes and reducing diffusion, leading to better ionization and detection.
A comprehensive system approach is essential for UPLC success. From the pump to the detector, every component must be optimized. The total system volume should be lower than that of traditional HPLC to maintain performance. Even small parts like tubing and fittings play a role in ensuring consistent results.
In practice, UPLC offers numerous benefits. It enables faster library screening, more accurate metabolite identification, and quicker peptide mapping. Quantitative analysis becomes more efficient, with better baseline resolution and reproducibility. Overall, UPLC redefines what is possible in chromatography labs, offering greater speed, resolution, and sensitivity.
UPLCTM is not just an upgrade—it's a transformation. It opens new possibilities for scientists working in analytical chemistry, pharmaceuticals, and bioanalysis. With continued innovation, UPLC will remain a cornerstone of modern separation science.
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