Investigation into reversed-phase chromatography peptide separation systems part V: Establishment of a screening strategy for development of methods for assessment of pharmaceutical peptides' purity

A Strategy for assessing peak purity of pharmaceutical peptides in reversed-phase chromatography methods using two-dimensional liquid chromatography coupled to mass spectrometry. Part II: Development of second-dimension gradient conditions

“…The resulting parameters, and the new prediction for the ϕ i needed for a target retention time of 15.83 min are shown at the bottom of Table 1. We see that the model still fits the data very well (average absolute difference between experimental and fit retention times is 0.5%) even though the resulting model parameters are very different from the first iteration (72.0 compared to 54.2 for S, and 1.40×10 9 compared to 8.40×10 6 for k w ). The new predicted value of ϕ i needed to obtain a 2 t r of 15.83 min is 24.3% ACN.…”

“…In a prior series of papers [6][7][8] we identified column and mobile phase (both volatile and non-volatile) combinations that exhibit large differences in selectivity for therapeutic peptides and related impurities. In Part I of this current series, we extended the prior work by assessing the separation of peptide isomers when using some of the most interesting column / mobile phase combinations identified in the prior work.…”

A strategy for assessing peak purity of pharmaceutical peptides in reversed-phase chromatography methods using two-dimensional liquid chromatography coupled to mass spectrometry. Part I: Selection of columns and mobile phases

“…Columns and mobile phases to be evaluated in this study were selected based on previous publications of column [4] and mobile phase characterisation [7]; see Tables 2 and 3. Weak (A) and strong mobile phases (B) were prepared as described in Table 3 and reference [8]. The gradient was standardised to 2.5 -100 -100 -2.5 -2.5 %B at 0 -60 -62 -65 -75 min and delivered at 0.3 mL/min.…”

“…In a series of previous publications, we have characterised 43 RPC columns [4][5][6] and 51 mobile phases [7] targeting peptide RPC method development. Subsequently, a method development strategy was published based on this characterisation work [8]. Both volatile and non-volatile mobile phase additives were investigated.…”

Controlling the elution order of insulin and its analogs in sub‐/supercritical fluid chromatography using methanesulfonic acid and 18‐crown‐6 as mobile phase additives

“…Mass spectrometry is often coupled to LC for characterization purposes but is obstructed by these buffers and requires substitution for TFA or formic acid. As a drawback, these often result in worse chromatographic performance regarding resolved peaks [9]. Alternatively, online or offline 2D‐LC can remove non‐volatile buffers before characterization through LC‐MS [10].…”