AVT - Biochemical Engineering, RWTH Aachen University, Forckenbeckstr. 51, D-52074, Aachen, Germany.
AVT - Chemical Process Engineering, RWTH Aachen University, Forckenbeckstr. 51, D-52074, Aachen, Germany.
BMC Plant Biol. 2018 Jun 1;18(1):101. doi: 10.1186/s12870-018-1305-6.
Ethylene is an important plant hormone that controls many physiological processes in plants. Conventional methods for detecting ethylene include gas chromatographs or optical mid-infrared sensors, which are expensive and, in the case of gas chromatographs, are hardly suitable for automated parallelized online measurement. Electrochemical ethylene sensors are cheap but often suffer from poor resolution, baseline drifting, and target gas oxidation. Thus, measuring ethylene at extremely low levels is challenging.
This report demonstrates the integration of electrochemical ethylene sensors into a respiration activity monitoring system (RAMOS) that measures, in addition to the oxygen transfer rate, the ethylene transfer rate in eight parallel shake flasks. A calibration method is presented that is not prone to baseline drifting and considers target gas oxidation at the sensor. In this way, changes in ethylene transfer rate as low as 4 nmol/L/h can be resolved. In confirmatory experiments, the overall accuracy of the method was similar to that of gas chromatography-mass spectrometry (GC/MS) measurements. The RAMOS-based ethylene determination method was exemplified with parsley suspension-cultured cells that were primed for enhanced defense by pretreatment with salicylic acid, methyl jasmonate or 4-chlorosalicylic acid and challenged with the microbial pattern Pep13. Ethylene release into the headspace of the shake flask was observed upon treatment with salicylic acid and methyl jasmonate was further enhanced, in case of salicylic acid and 4-chlorosalicylic acid, upon Pep13 challenge.
A conventional RAMOS device was modified for simultaneous measurement of the ethylene transfer rate in eight parallel shake flasks at nmol/L/h resolution. For the first time electrochemical sensors are used to provide a medium-throughput method for monitoring ethylene release by plants. Currently, this can only be achieved by costly laser-based detection systems and automated gas chromatographs. The new method is particularly suitable for plant cell suspension cultures. However, the method may also be applicable to intact plants, detached leaves or other plant tissues. In addition, the general principle of the technology is likely extendable to other volatiles or gases as well, such as nitric oxide or hydrogen peroxide.
乙烯是一种重要的植物激素，控制着植物的许多生理过程。传统的乙烯检测方法包括气相色谱仪或光学中红外传感器，这些方法既昂贵，又不适合自动化的平行在线测量（在气相色谱仪的情况下）。电化学乙烯传感器价格便宜，但通常分辨率差，基线漂移，以及目标气体氧化。因此，极低水平的乙烯测量具有挑战性。
本报告展示了将电化学乙烯传感器集成到呼吸活动监测系统（RAMOS）中，该系统除了测量氧气传递率外，还可以在八个平行摇瓶中测量乙烯传递率。提出了一种不易发生基线漂移且考虑传感器中目标气体氧化的校准方法。通过这种方式，可以分辨低至 4 nmol/L/h 的乙烯传递率变化。在验证实验中，该方法的整体准确性与气相色谱-质谱（GC/MS）测量相似。基于 RAMOS 的乙烯测定方法以欧芹悬浮培养细胞为例，这些细胞经过水杨酸、茉莉酸甲酯或 4-氯水杨酸预处理以增强防御能力，并用微生物模式 Pep13 进行挑战。在用水杨酸处理时观察到摇瓶顶空释放乙烯，在水杨酸和 4-氯水杨酸的情况下，茉莉酸甲酯进一步增强，在 Pep13 挑战时进一步增强。
对传统的 RAMOS 设备进行了修改，以在 nmol/L/h 的分辨率下同时测量八个平行摇瓶中的乙烯传递率。电化学传感器首次用于提供一种高通量方法来监测植物的乙烯释放。目前，这只能通过昂贵的基于激光的检测系统和自动化气相色谱仪来实现。新方法特别适用于植物细胞悬浮培养物。然而，该方法也可能适用于完整植物、离体叶片或其他植物组织。此外，该技术的一般原理很可能也可扩展到其他挥发性或气体，如一氧化氮或过氧化氢。
