Standard Test Method for Lipid Quantitation in Liposomal Formulations Using Ultra-High-Performance Liquid Chromatography (UHPLC) with Triple Quadrupole Mass Spectrometry (TQMS)

Importancia y uso:

5.1 Liposomes are vesicles of nanoscale dimensions, composed of lipid bilayers, which are used for various diagnostic and therapeutic applications (9). The growing interest in liposomal formulations in the delivery of various drugs, antisense oligonucleotides, cloned genes, or recombinant proteins by the biopharmaceutical industry, warrants QC and thorough characterization of the constituent lipids. Lipid structure, composition, and concentration are key attributes in determining the quality and efficacy of a liposomal drug product as they influence the stability of liposomes, drug loading, release kinetics, biodistribution, and pharmacokinetic properties (9). Cholesterol modulates the lipid membrane fluidity, elasticity, and permeability; hence, it plays a key role in controlled drug release and increased stability of the liposome (10).

5.2 This test method provides a rapid and reliable protocol for the determination of cholesterol, DSPE-PEG 2000, and HSPC in liposomal formulations using UHPLC-TQMS. Assessment of the stability of the analytes in terms of their degradation profiles is not included in this test method (11). This test method will benefit the biopharmaceutical industry in ascertaining quality assessment of liposomal formulations and monitoring batch-to-batch consistency for large-scale production, thereby facilitating safe and efficient drug development and regulatory review.

5.3 UHPLC-MS/MS measurements are analytically more sensitive and specific for lipid analysis compared to other contemporary techniques using universal detectors, such as a charged aerosol or an evaporative light-scattering detector. For liposomes, MS/MS has further advantages over ultraviolet detectors, as lipids lack chromophores for detection. In this test method, TQMS has been used as the MS/MS technique of choice because of its high selectivity, sensitivity, S/N, accuracy, and broad linear range of quantitation, thereby allowing reproducible quantitation of the analytes, especially at low concentrations.

5.4 According to the Current Good Manufacturing Practice regulations [21 CFR 211.194(a)(2)], users are required to verify the suitability of the test method under actual conditions of use. Validation should assess the suitability of the test method for the product matrix, recovery of the analytes from the product matrix, suitability of chromatographic conditions and column, appropriateness of the detector signal response, specificity, limit of detection and quantitation, accuracy, and precision. The user may need to optimize method parameters and cross validate if a different chromatography column, ionization method, or mass analyzer is used.

Subcomité:

E56.08

Referida por:

E3409-24

Volúmen:

14.02

Número ICS:

07.120 (Nanotechnologies)

Palabras clave:

cholesterol; DSPE-PEG 2000; HSPC; lipid quantitation; liposomal formulation; mass spectrometry;