Amino acid analysis is used throughout the world in many important areas including: clinical analysis, agriculture, food (human and animal), the environment, forensics, histochemistry and cell biology, pharmaceuticals, microbiology, medicine, nutrition, plant and protein sciences.
Amino Acid Analysis: a brief synopsis
JPP: simplifying the future
All JPP’s reagents used in analysis are ‘single-bottle’ with nothing else added at any stage. All have lives of more than three years in the presence of air at room temperature.
All other commercially available ninhydrin post-column reagents sold today for high performance ion chromatography determinations of amino acids are based on the original Spackman, Moore and Stein method, set out in a seminal work published in 1958. These Nobel Prize winning chemists found that the ninhydrin reagent used in this process required the presence of hydrindantin as an “accelerator”. This is because the reaction between just ninhydrin and amino acids/amine compounds to produce the intensely coloured Ruhemann’s Purple (RP) for spectrophotometric measurement is far too slow to be of any practical use in chromatographic systems.
In the presence of hydrindantin the ninhydrin reaction is considerably speeded up but still requires temperatures in the region of 130ºC to be close to completion in about one minute. Hydrindantin reacts quickly and easily with air so, in using the Moore & Stein method, all users try to handle solutions so as to rigidly exclude air. In practice this is all but impossible to achieve and solutions containing hydrindantin therefore inevitably slowly degrade. To offset this, most of the major manufacturers sell ninhydrin reagents in two bottles, one containing ninhydrin and the other containing hydrindantin. These constituents are then mixed when required and the manufacturers claim the resulting solution will then “last” from one to three months. It is widely known, however, that the mixed solutions only perform adequately for a few weeks at most and reagents left on the instrument have an even shorter lifespan. Therefore, even though it is nearly 60 years since the development of a ninhydrin post column reagent for the determination of amino acids by ion exchange chromatography by Moore and Stein, the reagent is still plagued by instability in the presence of air.
JPP: solving the problems
By altering the chemistry, JPP has developed ninhydrin reagents containing NO hydrindantin. These reagents work extremely effectively on all instruments in current manufacture. A high-speed reaction with amino acids and amines still occurs in the reaction coil at 130ºC to produce RP in about one minute or less but, because there is no hydrindantin involved in their preparation, JPP’s ninhydrin reagents are entirely stable and can be stored and used for periods of more than 3 years in the presence of air.
The new chemical and technical processes, which revolutionize ninhydrin post-column detection, are described in and protected by three patents in five key global territories. For further information see: JPP’s Patents.