Trace level detection of microbiological contaminants in pharmaceutical environment using Rapid Evaporative Ionisation Mass Spectrometry

Rapid evaporative ionization mass spectrometry (REIMS) yields highly specific phospholipid profiles of different bacteria and moulds which can be used to distinguish between different microorganisms. REIMS offers a new opportunity for the development of a lipid based, sample-preparation-free microbial identification system. This technique would be considerably faster than current methods and have the sensitivity and robustness compared to other techniques (e.g. MALDI).

This project is focusing on developing technique to detect and categorise different microorganisms in the presence of drug substances, excipients in a tablet formulation and in liquids at a suitable detection limit. Another focus of the project is to develop a technique which would be used for identifying isolates from environmental monitoring plates which would be an enabler for future real time release (as part of a wider control strategy). Pharmaceutical water analysis, absence/presence of specified organisms clauses which are required for a range of pharmaceutical dosage presentations are another two possible REIMS applications which this project is focusing. Comparing with existing methods and procedures in pharmaceutical companies, REIMS technology could potentially be a technique with great savings for the business (e.g. saving time and money, increasing sustainability, integrity and robustness, reducing delays for batch and stock release).

Below, I have put together project plan for the first 2 years and introduced timelines to show which part of the plan have been started or will be started in the upcoming months.

Big part of the project is environmental monitoring. In order to develop a technique which would be able to identify the plates, construction of pharmaceutical environmental isolates data base is required. Hundreds of samples (bacteria and moulds) are send to Imperial every
week from AstraZeneca, where samples are saved in to the sample data base for further use and identification by MALDI or 16S sequencing (about 80% of samples have to be sequenced to obtain true identification which is crucial for further technique development due to the not matching results from AZ used techniques for identification and Imperial MALDI).

1st -2nd Year Work Plan:

1. Construction of environmental REIMS identification database
a. Comparison between environmental and clinical samples
b. Construction and validation of REIMS identification database c. Comparison to AZ (AstraZeneca) in-house techniques
d. Samples analysis using REIMS, reference model building and validation
e. Investigation of samples storage conditions effect for analysis on REIMS

2. AZ water samples analysis on REIMS (purified water, water for injections, borehole water)
a. Develop technique to concentrate and enrich samples for analysis
b. Identify lower limit detection/CFU of bacteria required by regulations
c. Identify method for E.coli and Coliform detection such as short incubation, isotopic labelling, magnetic separation or biomarkers analysis.
d. Identify method for viable bacteria detection (reference model building for
prediction approach)

3. Fungi Identification
a. Construction and validation of REIMS identification database
b. Investigation of fungi colonies metabolic changes during 7 day growth on
REIMS
c. Samples analysis using REIMS, reference model building and validation d. Investigation of samples storage conditions effect for analysis on REIMS

4. Culturing free approach for drug substances or drug itself
a) Investigate solvents suitable for dissolving drug substances if required
b) Develop technique to concentrate samples for analysis (filtering approach)
c) Lower limit detection/CFU of bacteria required by regulations
d) Identify method for viable cell detection such as short incubation, isotopic labelling, magnetic separation or biomarkers analysis.

5. Instrument development
a. Investiga