In the initial few months of this project a five pronged approach was adopted. A major consideration was the potential use of instrumentation found in many wineries â€“ Fourier Transform Infra-Red (FTIR) spectroscopy typified by the FOSS Winescan. We also had access to two types of Near Infra-Red (NIR) instruments that were being used for other winegrowing projects. The final spectrometric approach was Raman Spectroscopy. The objective of using these instruments is to find reliable differences in the spectra that could be used to distinguish sound grapes from moldy grapes. Two additional technologies available at Fresno State are Flow Cytometry and hand-held Gas Chromatography â€“ zNose. The assay objectives are listed in the figure below.
Figure included in progress report.
Flow cytometry: a fluorescent compound that binds to chitin, a compound found in the cell wall and spores of molds (the lectin, wheat germ agglutinin) can be used in conjunction with flow cytometersâ€™s cell counting to detect molds. Mold fragments and spores were detected using this system. The method needs further development to quantify mold material present in grape loads.
Gas chromatography: all molds produce volatile organic compounds (VOCs) when growing (like a car exhaust). The zNose hand-held gas chromatograph is versatile and can be used to concentrate on identifying VOCs produced by common grape molds. Differences have been detected in the VOC patterns produced by the three molds growing on whole grapes. Identification of these VOCs will be the basis of an assay for mold detection.
Raman Spectroscopy was eliminated as a possible method when it was determined that the spectra from moldy and non-moldy grapes showed little variation.
NIR and FTIR Homogenized and sterilized grapes were inoculated with one of three molds or a combination of all three molds. Samples were taken according to a protocol, processed and run on the spectrometers . The NIR instruments distinguished moldy grapes from non-moldy grapes The FOSS FT 120 Winescan uses the whole spectrum and can distinguish moldy from nonmoldy grapes in a validation test.
The results from the FTIR are very encouraging, although there remains much testing to determine if this will be an effective system. The NIR, Flow Cytometry and zNose gas chromatography also offer very promising possibilities of quantifying grape rot. Raman Spectroscopy did not generate spectra that could be reliably used to quantify grape rot.
Further experimentation with FTIR and NIR spectroscopy, VOCs, and flow cytometry are recommended in a multipronged approach to establish a reliable method of quantifying grape rot.