Optimization of HPLC conditions for the separation of complex crude mixtures produced in the synthesis of therapeutic peptide hormones

Abstract

SummaryThe solid phase peptide synthesis (SPPS) method is usually used for the synthesis of new peptides of pharmaceutical interest. However, the final drug product commonly results in complex mixtures, where the target peptide must be separated from the undesired impurities. The linear solvation energy relationships method (LSER) is used here in order to rapidly optimize the separation of the analytes present in the complex crude mixtures which are formed during the synthesis of peptide hormones with therapeutic interest, such as: carbetocin, eledoisin, leuprolide, goserelin and triptorehn. Resolution data for adjacent solute pairs may easily be predicted from these relationships. Separation under optimum conditions provides useful information about the purily of the target peptide and allows characterization of the mixture components by LC-ES-MS. In addition conditions for the preparative purification of the candidates could be extrapolated from these analytical data.

Capillary electrophoresis coupled to time of flight‐mass spectrometry of therapeutic peptide hormones

“…Therefore, development of separation, analysis and characterization methods remains one of the most challenging tasks to develop. Previously, methods based on CE and liquid chromatography (LC) with ultraviolet (UV) detection [12][13][14], and LC coupled to MS [15,16] were proposed for the separation and characterization of a series of peptides and related compounds, such as peptide impurities in crudes of reaction. In this work, we have established a method for separation and characterization of a series of peptide hormones of pharmaceutical interest and wide therapeutical use by CE-ES-MS using a sheath-flow interface.…”

Hydrophilic interaction chromatography for the analysis of biopharmaceutical drugs and therapeutic peptides: A review based on the separation characteristics of the hydrophilic interaction chromatography phases

“…We have to keep in mind that such a strong interaction between positively and negatively charged stationary phases and analytes often results in worse separation efficiency [379]. Afamelanotide (2014) 75921-69-6 1646.9 N-Ac-SYSXEHFRWGKPV-NH 2 X = L-norleucine −6.46 −5.91 −5.05 1.41 [372], [407], [408] Carbetocin (2015) 37025-55-1 988.2 N-(4-mercapto-1-oxobutyl)-Omethyl-L-tyr-L-ile-L-gln-L-asn-L-cys-L-pro-L-leu-Gly-NH 2 thioether cyclic (1-5) −1.06 −1.06 −1.06 −1.06 [409], [410], [411] Cetrorelix (2000) 120287-85-6 1431.0 N-Ac-(2-Naph)-AF-(3pyr)ASYXLRPA-NH 2 , X = N-5-(aminocarbonyl)-Dornithyl −2.03 −0.97 −0.62 1.38 [412], [413], [414] Degarelix ( 31 [428], [429], [430] Bold-and italic character stands for D-amino acid, 2-Naph-A, 3-(2-naphthalenyl)-alanine. Peptide therapeutics such as cyclosporins, linear gonadotropin-releasing hormones, cyclic hormones, and cyclic antibiotics were separated on six HILIC columns developed by Armstrong and co-workers [380].…”

Purification and identification of an antioxidant peptide from Pinctada fucata muscle

“…Using a 0.2-μm UF membrane, F2 was further separated by semi-preparative RP-HPLC on a Zorbax, SB C-18 column, and five purified peptides named FC1, FC2, FC3, FC4 and FC5 were separated as shown in Figure 3. A variety of peptides were eluted on RP-HPLC given their different polarities, and this technique was popularly applied in the isolation and purification of small peptide segments (Dan, Ganesan, Flood, Tsai, & Reif, 2000), especially those less than 5000 Da (Sanz-Nebot, Benavente, Toro, & Barbosa, 2001). The antioxidant activities are presented in Table 2.…”