# Proteomic Evaluation of Insecticidal Action of Phosphine on Green Peach Aphids, _Myzus persicae_
## 4 Discussion  
PH\({}_{3}\) toxicity has been widely studied using its target invertebrates, including stored products insect pests, to understand how it disrupts normal physiological functions, leading to death [10; 11; 12]. A recent paper by [PERSON] et al. [17] reviewed the two major routes of PH\({}_{3}\) toxicity; an energy imbalance via the breakdown of the electron transport system in mitochondria, and metabolic disturbance. Other toxic mechanisms by which PH\({}_{3}\) acts have been identified in rats and other mammals, such as a neurological disorder via potent inhibition on AChE activity, and severe inhibitory effects on metalloprotein, such as catalases and COX [18]. Prior to an amassing a deeper understanding of PH\({}_{3}\) toxicity in _M. persicae_, many studies showed that PH\({}_{3}\) interfered with COX activity, which is known as complex III in the electron transport chain [18; 19]. Cytochrome c cox is a hemeprotein, within which PH\({}_{3}\) can easily form coordination bonds with the ferric ions, thereby reducing COX activity to produce less ATP than normal [18]. In our study, _M. persicae_ was exposed to low PH\({}_{3}\) concentrations as LCT\({}_{10}\) value (LC\({}_{10}\) value), which did not kill _M. persicae_. As shown in Figure 2, in the PH\({}_{3}\)-treated adults _M. persicae_ COX activity decreased with increasing concentrations of PH\({}_{3}\). However, this does not indicate that a direct inhibition of PH\({}_{3}\) on the target site of COX occurred or there was an indirect inhibition of COX activity by PH\({}_{3}\). LC-MS/MS analysis showed that COX expression decreased after PH\({}_{3}\) treatment (Figure 4A). Therefore, a reduction in COX activity after PH\({}_{3}\) treatment was related to the reduced expression of the corresponding protein in _M. persicae_. This result was confirmed by [PERSON] and [PERSON] [18], who showed that no spectral changes with COX were found after the PH\({}_{3}\) treatment, indicating that there was no direct or weak contact between the COX protein and PH\({}_{3}\) molecules. On the contrary, other hemeproteins, hemoglobin, and myoglobin showed slow deoxygenation with spectral changes after PH\({}_{3}\) treatment [18]. Interestingly, changes in COX activity were not dramatic, whereas cytochrome b-c1 complex subunit Rieske and ATP synthase subunit g in the PH\({}_{3}\)-treated _M. persicae_ decreased by around 0.5-fold in terms of protein expression when compared with the control. Similarly, a reduction in ATP synthase was confirmed by [PERSON] et al. [20], which showed a 0.15-fold change in PH\({}_{3}\)-treated larvae of the peach fruit moth, _Carposina [PERSON]_ [PERSON], using 2-dimesional electrophoresis with the MALDI TOF MS analysis. In this study, the most important finding is the dramatic change in NADH dehydrogenase (ubiquinone) flavoprotein 1 (\(\times\)3.35-fold), which is complex I in the electron transport chain. It may be the first evidence that this protein is involved in PH\({}_{3}\) toxicity in _M. persicae_. The ETC in the PH\({}_{3}\)-treatedM. persicae_ was largely modified and the imbalance in ATP production was probably related to the metabolic disorder, leading to death of _M. persicae_. However, regarding the up-regulation of NADH dehydrogenase (ubiquone) flavoprotein 1 (complex I), one possible cause of PH\({}_{3}\) toxicity may be the overproduction of reactive oxygen species (ROS). As ROS are primarily generated in the mitochondrial ETC, complexes I and III are major sites for the production of small quantities of ROS [20; 21]. Normally, the generated ROS is converted to hydrogen peroxide by superoxide dismutase, and then hydrogen peroxide can be transformed to water by peroxididases, which PH\({}_{3}\) inhibits in _Sitophilus granarius_[22]. Therefore, more ROS formation will probably occur in response to the up-regulation of complex I after PH\({}_{3}\) fumigation, and this ROS generation can lead to the death of _M. persicae_ during PH\({}_{3}\) fumigation.