Experimental Growth Conditions affect Direct and Indirect Defences in two Cotton Species.

Cotton has been used as a model plant to study direct and indirect plant defence against herbivorous insects. However, the plant growing conditions could have an important effect on the outcome of such plant defence studies. We examined how common experimental growth conditions influence constitutive and inducible defences in two species of cotton, Gossypium hirsutum and G. herbaceum. We induced plants by applying caterpillar regurgitant to mechanical wounds to compare the induction levels between plants of both species grown in greenhouse or phytotron conditions. For this we measured defence metabolites (gossypol and heliocides) and performance of Spodoptera frugiperda caterpillars on different leaves, the emission of plant volatiles, and their attractiveness to parasitic wasps. Induction increased the levels of defence metabolites, which in turn decreased the performance of S. frugiperda larvae. Constitutive and induced defence levels were the highest in plants grown in the phytotron (compared to greenhouse plants), G. hirsutum and young leaves. Defence induction was more pronounced in plants grown in the phytotron and in young leaves. Also, the differences between growing conditions were more evident for metabolites in the youngest leaves, indicating an interaction with plant ontogeny. The composition of emitted volatiles was different between plants from the two growth conditions, with greenhouse-grown plants showing more variation than phytotron-grown plants. Also, G. hirsutum released higher amounts of volatiles and attracted more parasitic wasps than G. herbaceum. Overall, these results highlight the importance of experimental abiotic factors in plant defence induction and ontogeny of defences. We therefore suggest careful consideration in selecting the appropriate experimental growing conditions for studies on plant defences.

Improved separation by at-column dilution in preparative hydrophilic interaction chromatography.

We evaluated at-column dilution (ACD) as a possible approach to reduce peak distortion caused by the injection of strong solvents in preparative hydrophilic interaction chromatography (HILIC). Three model compounds, namely uridine, cytidine and guanosine were selected and injected either in conventional or ACD conditions, using various proportions of water and acetonitrile in the sample diluent. Plate number and peak capacity were systematically investigated under both isocratic and gradient elution conditions, respectively. In isocratic conditions, ACD was found to provide higher plate numbers with sample diluent containing more than 40% water, but lower plate numbers with injections of less than 40% water, in comparison to a conventional injection system at the preparative scale. These contrasting results were attributed to the fact that i) efficiency was fundamentally reduced in ACD, due to the low make-up flow rate that was used to flush the injection loop, but ii) the trend was reversed for highly aqueous injection thanks to the capacity of ACD to maintain similar efficiency regardless of the injection solvent. On the other hand, peak capacity was comparable between ACD and conventional systems in gradient elution for injections containing low amounts of water. However, ACD became increasingly advantageous as the proportion of water in the injection solvent and/or the injection volume increased. This was confirmed by the separation of a plant root aqueous extract. Altogether, our findings suggest that ACD is beneficial both in isocratic and gradient modes when strongly aqueous solutions are injected, offering an attractive approach to increase loadability, while preserving peak shapes in preparative HILIC.