When it comes to polar organic compound purification, many chemists turn to normal-phase flash chromatography often utilizing dichloromethane and methanol as the eluting solvents. While this can work, it often can be challenging to optimize due to methanol’s high polarity and protic chemistry.
Improvements in solid phase peptide synthesis strategies and development of resin linkages susceptible to low acid cleavage conditions has enabled synthesis of long peptides while keeping the protecting groups intact. This strategy is now used for the preparation of chemically synthesized proteins, wherein shorter peptide fragments are ligated together. They are also found in the synthesis of peptide macrocycles that utilize head-to-tail cyclization strategies. Although linear synthesis of protected peptides is generally straightforward, purification of these compounds using traditional reversed phase methods is quite challenging. Herein we describe the use of normal phase chromatography for purification of fully protected peptides.
This brochure presents the extensive range of instrumentation for flash purification from Biotage. To complement these products Biotage also offers a complete range of flash consumables, including columns in a variety of sizes packed with irregular and spherical silica, making Biotage your one-stop partner for your flash purification needs. Keywords: Isolera™ Dalton, Isolera™ Spektra, 10 Isolera™ Spektra One and Four, Isolera™ Spektra LS (Large Scale), Isolera™ Prime, Isolera™ ELSD-1080, Biotage® Flash 75/150, Biotage® Flash 400
User Report: Isolera™ Dalton, Okayama University. The laboratory of Prof. Hiroyuki Miyachi at Okayama University installed Biotage’s Isolera Dalton, an automated mass-directed purification system for flash chromatography, allowing the fractionation and collection of optical isomers. “I felt like the world’s first mass-detection automated purification system had finally arrived.”
The term “Green Chemistry” has become a major part of the science community’s lexicon. In this application note we will look at two areas for flash chromatography: 1. Replacing chlorinated solvents with those considered more environmentally friendly. 2. Reducing solvent use and waste generation with more thoughtfully applied chromatography principles.
As reversed-phase flash chromatography gains traction in medicinal chemistry labs the need to monitor its cost and safety are becoming more important. Commonly used reversed-phase solvents typically include water with an organic solvent such as methanol or acetonitrile – each have advantages and disadvantages.
Reversed-phase chromatography is typically used when you need to separate several milligrams of relatively polar compounds that either are not soluble in normal-phase solvents or are not compatible with bare silica because they react, stick, or both. If you are currently using reversed-phase at preparative scale, such as flash chromatography, you know the mobile phase limitations – water with either methanol, acetonitrile, or THF. As with normal-phase flash chromatography, when it comes time to purify you want your crude sample fully solubilized in the weakest possible solvent at the highest possible concentration. ACS 2016
Although capable of very high resolution, RP-HPLC is often limited by low column loading capacity, therefore demanding a significant time investment for peptide purification. As an alternative strategy, reversed-phase flash chromatography can also be used to purify synthetic peptides. The larger particle size used in flash column chromatography enables much larger loading capacity, thereby significantly reducing the time required for peptide purification.
User report: Flash instruments. Chugai Pharmaceutical uses Biotage flash chromatography products for drug discovery research. When deciding to convert from manual open-column procedures to automated systems, they chose successive generations of Biotage products, ranging from the Flash+® packed column to the Biotage® Horizon, SP1, Isolera™ Spektra, and Isolera™ Dalton automated flash chromatography systems.
Peptide purification using standard reversed phase HPLC methods are hampered by low loading capacity, resulting in purifications that demand significant time investment. Recently, the use of reversed phase flash chromatography has increased in popularity for peptide purification due to the significant reduction of purification time, enabled by the increased loading levels of the larger stationary phase particles. Resolution, though, is somewhat diminished with the larger particle size, demanding creative techniques to retain a highly pure peptide product.
In medicinal chemistry, organic synthesis will generate by-products with similar chemistry to the product. Separating these impurities from the product often becomes a challenge that often only prep HPLC can achieve. In this poster, we discuss how using sample dry loading with a cation exchange scavenger media with flash chromatography improves product purity by removing many of the impurities.