Development of Sensitive and Robust Mass Spectrometric Methods for Detection of GM Material in Food Ingredients

This research project aims to develop a new method for the detection of GM proteins in food products.

<p>This project has seven objectives:

<p><b>1. To produce and digest recombinant GM proteins to obtain accurate masses of GM biomarker peptides</b>
GM proteins will be produced in pure milligram quantities. These proteins will then be digested with optimisation of substrate concentration, temperature and incubation period. A comprehensive analysis of peptides produced by digestion will be undertaken. A reproducible stable characteristic profile of peptides will be produced. Accurate masses and MS/MS data of these peptides will be produced on a variety of MS instrumentation. The chromatographic properties of peptides will be ascertained which will aid the peptide separation protocols in the third objective. A database of peptide masses and sequence tags from GM plants will be formed that can act as biomarkers in objectives 2, 4, 5 and 6.

<p><b>2. To optimise protein extraction from GM and non-GM soya, maize and tomato</b>
Simple, high recovery robust and reproducible protein extraction procedures will be developed. These will be compatible with digestion and separation procedures as well as MS identification approaches. The storage properties of the GM/non-GM material and respective protein extracts will be determined.

<p><b>3. To develop protein digestion and peptide separation conditions for GM proteins/peptides in soya, maize and tomato</b> Conditions for protein digestion of GM and non-GM proteins in complex mixtures will be evaluated. Digestion will then be performed optimising substrate concentration, temperature and incubation period. The reproducibility and stability of characteristic peptide profiles will also be assessed. Finally a protocol will be devised to optimise the recovery and elution conditions for GM peptides in endogenous mixtures using chromatography.

<p><b>4. To develop mass spectrometric methods for GM peptides</b>
Various, but complimentary, MS approaches will be used to determine accurate and reproducible masses of characteristic peptides derived from GM proteins.

<p><b>5. Assessment of the detection sensitivity in GM food products</b>
Previously developed methods will be used to detect GM proteins present in soya and maize containing 1, 0.5 and 0.1% GM product. GM control tomato material will be prepared and detection limits assessed using methods described. These will be repeated to assess the reproducibility, robustness and sensitivity of the methods developed.

<p><b>6. To carry out a feasibility study for the use of ion trap MS</b>
Once GM marker peptides have been identified using the previously described methods, the feasibility of using the ion trap system for routine detection of GM peptides will be performed. This is a more robust detection system.

<p><b>7. Data analysis and preparation of searchable database</b>
Data will be analysed and a database will be created for generic use with GM peptides.