Sekisui – Method Development for a Process Related Residual

BioProduction by Sekisui offers contract development and manufacturing services for the biopharmaceutical industry.  Their facility in Maidstone, UK has over 40 years of bioprocess development and manufacturing experience with expertise in the fermentation of enzymes and other proteins.

Project Overview

Sekisui required a method to test for clearance of the flocculant polydiallyldimethylammonium chloride (pDADMAC) throughout a production process. Previous to contacting BioPharmaSpec, Sekisui had investigated a mass spectrometric method, but it was not robust enough for reliable use. Other alternative methods, such as HPLC-SEC with ELSD detection, were discussed but BioPharmaSpec scientists were confident that a robust MS method could be developed.

BioPharmaSpec successfully developed a method by enhancing the relatively non-specific client-supplied LC-MS method to provide LC-MS/MS (LC-MRM) analysis. The newly developed method provided improved sensitivity and selectivity and also removed any interference from the LC solvents and/or product components.

Technical Consideration- Analysis of Residuals

pDADMAC improves the removal of contaminants by driving the charged flocculation of the negatively charged entities. Upon addition to cellular extracts, it rapidly flocculates nucleic acid and other cellular debris into larger particles via an ionic interaction mechanism. Due to the possible toxicity, flocculant polymers should be assessed for an acceptable residual level, and a clearance level must be reached to ensure drug safety.

pDADMAC is a large polymeric molecule composed of repeating units each containing a quaternary nitrogen. The large size, high positive charge density, propensity to form cationic adducts and solubility combine to give a structurally challenging species to work with.

A mass spectrometric method of pDADMAC analysis was developed using on-line LC/ES-MS. Initial mass spectrometric investigations identified key ions that were subsequently shown to be suitable for a quantitative analysis of pDADMAC in the sample matrix through a Multiple Reaction Monitoring approach.

Client Testimonial

“We use pDADMAC as a flocculation agent for one of our client’s protein products and needed to develop a robust method to identify and quantitate pDADMAC levels throughout the process. After unsuccessful attempts to develop a mass spec method in the past, we were delighted to find BioPharmaSpec and discover the specific technical expertise they had in this area. They developed a robust method that we use regularly to support this development project.” – Principal Scientist, R&D – Sekisui