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Application: ATP degradation products
Determination of ATP degradation products from fish decomposition. Technology: reversed phase. Present substances: inosine monophosphate, adenosine triphosphate, adenosine diphosphate, adenosine monophosphate, hypoxanthine, purine, inosine.
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When should a gradient be used?
More Questions and AnswersIn reverse phase HPLC it is recommended to run a scouting gradient if the conditions for a successful separation are unknown. Such a run is performed from ca 10-80% organic modifier during 30-50 min, using a linear increase in elution strength. Based on the result of this scouting gradient, either an appropriate mobile phase composition can be chosen for isocratic elution (small molecules < 1000 g/mol) or a suitable gradient for the separation of peptides or other larger molecules.
Gradient elution can also be chosen for separation of compounds that vary greatly in their polarity. As a rule of thumb, if peaks can be detected during more than 25% of the scouting gradient, gradient elution will likely be the best choice for that separation problem. If peaks appear for less than 25% of the scouting gradient, isocratic elution should be preferred, as the selectivity will always be superior under isocratic conditions.
If the sample consists of two distinct groups of compounds, step elution can lead to a good separation results. In this case one starts with isocratic mobile phase elution that renders satisfying separation results for the less retained group of compounds, followed by increasing the elution strength of the mobile phase in one step to such a degree that will separate the more retained group of compounds in an appropriate way. Step gradients can also be used for washing the column after every injection. In this case the elution strength is increased drastically after the last peak has eluted. Strongly retained impurities can then be removed from the column.
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A column is not merely a vessel for HPLC-media, it is an integrated part of the final product, and it cannot be packed by just anyone. Therefore, all Kromasil Original Columns are packed by us, here at Kromasil, to guarantee the best quality and usage of Kromasil.
Chiral stationary phases
Enantiomer separation is a challenging task to achieve. Kromasil Chiral media are available with four different chemistries that offer a broad range of selectivity for your chiral separation from analytical columns to full process scale bulk media needs.
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You need to quickly purify substances for further studies in the development of pharmaceuticals. Kromasil will help you achieve your goals with ease and efficiency.
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