Automated dynamic sampling system for the on-line monitoring of biogenic emissions from living organisms

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Abstract

An automated system for continuous on-line monitoring of biogenic emissions is presented. The system is designed in such a way that volatiles, emitted as reaction to biotic or abiotic stress, can be unequivocally elucidated. Two identical sampling units, named target and reference bulb, are therefore incorporated into the system and consecutively analyzed in monitoring experiments. A number of precautions were considered during these experiments to avoid the application of unwanted stress onto both organisms. Firstly, the system is constructed in such a way that both bulbs are continuously flushed, i.e. before, during and after analysis, with high purity air to avoid any accumulation of emitted volatiles. Moreover, the air is pre-humidified by bubbling it through water to sustain the biological samples for longer periods in the in vitro environment. Sorptive enrichment on polydimethylsiloxane (PDMS) was used to trap the headspace volatiles. The hydrophobic nature of this material permitted easy removal of trapped moisture by direct flushing of the sampling cartridge with dry air before desorption. The system was used to monitor the emissions from in vitro mechanically wounded ivy (Hedera helix) and of in vitro grown tomato plants (Lycopersicon esculentum Mill.) upon cotton leafworm (Spodoptera littoralis) feeding. Differences in light and dark floral emissions of jasmine (Jasminum polyanthum) were also studied.

Introduction

An extremely exciting field of research comprises the search for the overall biological significance of components released by biological materials, i.e. biogenic emissions. Owing to the large diversity in emission sources (animals, plants) as well as in compound types (permanent gases, volatile organics), the entire field of interest is very wide and covers many different scientific disciplines. Insect ecology, phytochemistry, plant physiology, entomology, food and perfume chemistry are some of the areas where investigation of volatile emissions is carried out regularly.

The potential impact generated results may exhibit is far-reaching and not restricted solely to these areas. It is quite well known, for instance, that the communication within troops of social insects (bees, ants, wasps) primarily passes via volatile secretions (pheromones) [1], an effect that has been employed already in specific insect lures [2]. Moreover, also the field of plant volatile research has experienced great progress. It has been demonstrated, for example, that certain plant volatiles inhibit fungal proliferation [3], perhaps indicating future directions for genetic improvement. Additionally, particular volatile organics serve as critical volatile markers in such insect behaviors as host finding, feeding and ovipositioning [4], [5], and can be eventually exploited to craft effective methods for control of insect pests, as an alternative to the toxic agrochemicals applied today [6]. A comprehensive overview of the various classes of plant volatiles, their occurrence in crops and non-crops and their overall significance in environmental interactions was provided by Charron et al. [7].

As can be expected from the literature dealing with these topics, presented results primarily focus on the final outcome of the above mentioned effects and although the analytical part is really critical with respect to the reliability of the final results, it is often merely of secondary importance. The prime portion of biogenic emissions are analyzed using dynamic adsorptive enrichment, a technique which was developed nearly 40 years ago for the analysis of apolar to medium polar volatiles in air. The success of this approach is primarily due to the significantly higher sensitivity that can be attained compared to the alternative (static) methods such as solid-phase microextraction (SPME) [8] and headspace sorptive extraction (HSSE) [9]. Unfortunately, dynamic enrichment is not free of hazards, especially when applied without full knowledge of its potential drawbacks. However, all of these problems are a direct consequence of the use of adsorbents and originate from the strong interactions that occur between trapped analytes and the adsorbent surface. Therefore, sufficient retention and high enrichment factors hardly are an issue, though more harsh desorption conditions are required to remove the analytes.

In general, liquid extraction of the adsorbent bed with a suitable organic solvent is sufficiently invasive to allow complete desorption. Unfortunately, a great part of the enrichment effect acquired through sampling has to be sacrificed since only a small portion of the final liquid extract is available for subsequent analysis. Consequently, relatively long sampling times and/or high sampling flow-rates are necessary to deliver sufficient sensitivity in a reasonable amount of time.

From a sensitivity standpoint, it is much more attractive to employ the alternative method, i.e. thermal desorption. Here, the adsorbent is inserted in a small oven placed on top of the gas chromatograph (GC), which is subsequently heated to elevated temperatures. When properly designed, this set-up allows complete transfer of the sample from trap to analytical column. However, the strength of the analyte–surface interactions calls for high desorption temperatures, which can lead to the formation of artifacts and incomplete desorption, especially when polar and/or higher molecular mass compounds (MW) are involved [10], [11], [12].

Recently, sampling tubes packed with 100% polydimethylsiloxane (PDMS) particles were introduced for the enrichment of volatile emissions [13], [14]. With this material, preconcentration occurs by sorption or dissolving of the target analytes in the bulk of the liquid phase instead of adsorption onto a porous surface. Since this process is less energetic compared to adsorption, trapped volatiles are easily desorbed from the trap using thermal desorption, even at relatively mild temperatures. In addition, the absence of active sites avoids displacement effects and catalytic modifications of trapped components from occurring [12]. The performance of the dynamic configuration with PDMS has been evaluated for the analysis of plant volatiles, where it performed superior to Tenax, which was characterized by a significant interference of degradation products arising from decomposition of the polymeric skeleton and incomplete removal of higher MW analytes [15].

Just recently, we described a completely automated and stand-alone system for the analysis of gaseous emissions, e.g., ethylene, released by plants [16]. However, in order to be able to analyze regular volatile emissions, an adapted version of this system had to be developed. These adaptations were carried out successfully and the system was immediately used for determining the response of Arabidopsis thaliana to oxidative damage [17]. The results presented in that particular paper mainly dealt with the various problems and strategies encountered when identification of unknown volatiles has to be performed, without paying too much attention to the practical operation and additional possibilities of the device. These particular matters will be discussed in the present paper.

Section snippets

Biological materials

Ivy (Hedera helix) and jasmine plants (Jasminum polyanthum) were purchased from a local florist and placed in an isolated room under four TL-lamps (Osram, 32 W/lamp) equipped with a time controller to obtain a 16 h photoperiod (day, 4.00 a.m. to 8.00 p.m.). The temperature inside the room fluctuated around 25°C during the light period and 18°C during the dark period. Plants were kept in this chamber until analyzed.

Three-week-old tomato plants (Lycopersicon esculentum Mill.) of the cultivar

Results and discussion

The relative complexity of the final set-up (see Fig. 1) is a direct consequence of the necessity for appropriate accommodation of the organisms during analysis. When dealing with plant material, this primarily implied a regular supply of water and nutrients, control of temperature, relative humidity (RH), light strength and photoperiod. The two latter factors were easily implemented by means of a set of lamps, which were equipped with a time-controlled power switch. Practical implementation of

Conclusion

The instrumental set-up for automated on-line monitoring of biogenic emissions presented in this paper has proven to be very flexible and versatile. The use of a reference unit can compensate for external parameters such as temperature, relative humidity and light strength. System operation and chromatographic performance were strongly influenced by the unavoidable presence of water, which demanded the use of post-sampling purging with dry air. The use of sorption-based enrichment and the

Acknowledgements

We thank Ghent University for supporting this work through grant GOA 12051898. JV gratefully acknowledges the Flemish Institute for the Promotion of Scientific and Technological Research in the Industry (IWT), Flanders, Belgium for a study grant. Ir. K. Maes and Prof. Dr. Ir. L. Tirry from the Faculty of Agriculture of Ghent University are thanked for supplying the tomato plants and the caterpillars.

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