Column performance, irreproducibility

The biggest problem in using NP-HPLC is its dramatic sensitivity to water. Even water traces (in the mobile phase or from the sample) may bind to the column, deteriorate its performance, and cause irreproducibility. In addition, particular care must be taken to ensure accurate pH, as in NP-HPLC, retention is very sensitive to the charge state of the analyte. Owing to these practical problems NP-HPLC is relatively rarely used. Its main application fields are separation of polyaromatic hydrocarbons, sterols, vitamins, chlorophylls, ceramides, and other Upid extracts. 

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