Nicotiana benthamiana, Domin

Andersen, Trine Bundgaard, Rasmussen, Silas Anselm, Christensen, Søren Brøgger & Simonsen, Henrik Toft, 2019, Biosynthesis of tovarol and other sesquiterpenoids in Thapsia laciniata Rouy, Phytochemistry 157, pp. 168-174 : 170

2.4. Characterization of TlTPS18983 expressed in N. benthamiana

Volatiles from N. benthamiana leaves expressing Tl TPS18983 were captured by HS-SPME and analyzed by GC-MS. The analysis of these plants showed that expression of Tl TPS18983 generate at least 15 products. The identification of one of the major products, farnesene (5), was based on comparison of the mass spectrum of the compound with NIST and Wiley GC-MS spectrum libraries ( Fig. 4 , MS spectrum is provided in supplementary data Fig. 3 ). It was not possible to identify the remaining products. Using an injection port temperature of 160 ̊C instead of 250 ̊C did not alter the composition of the chromatogram, which supports that these compounds are not a result of the GC-MS analysis method. Extraction of the products was also attempted using organic solvents such as hexane and pentane mimicking procedures that were successful for other sqTPS products. However, for Tl TPS18983 we did not detect any compounds by GC-MS using organic solvent extraction.

Figures

Fig. 4. a) HS-SPME GC-MS analysis of TlTPS18983 expressed together AtHMGR, the control sample show the expression of AtHMGR alone. Farnesene (5) is seen as the major product (Rt 8.75 min), * show unidentified sesquiterpene like products. Based on this TlTPS18983 is a multiproduct terpene synthase with farnesene as the major product. MS spectra are presented in supplementary data.

Fig. 3. HS-SPME GC-MS analysis of TlTPS509 expressed together AtHMGR, the control shows the expression of AtHMGR alone. Guaiol (3) is seen as the major product (Rt 9.89 min), with bulnesol (4), Rt: 10.2, a-guaiene (Rt: 8.6), and a-bulnesene (Rt: 9.18) as minor products. * show unidentified sesquiterpene like products, which is either ketone formed during the biosynthesis or breakdown products of the alcohols formed during the GC-MS analysis. Based on this TlTPS509 is a guaiol synthase. MS spectra are presented in supplementary data.