Investigation Of Metabolic Process Of ABA And 6-BA Based On Micro-valve Spray Chip-MS Platform

Microfluidic chip coupled with mass spectrometry has wide applications in proteomics,cell analysis,and drug evaluation,because of its advantages in high throughput,strong versatility and high sensitivity etc.In this work,automated switching injection was achieved based on microfluidic microvalve technology,considering long time-consuming and poor reproducibility of manual sampling methods.Moreover,a high-throughput microvalve spray chip coupled with mass spectrometry was constructed and applied in following researches.1.In this part,a microvalve spray chip was prepared based on polydimethylsiloxane(PDMS).With integrated microvalves,microchannels,and microemitter on the microchip,automatic switching sampling of high-throughput samples was achieved.The microchip and mass spectrometer were online coupled with the monolithic electrospray emitter.We fully evaluated the microvalve,and the results indicated its good performance-Flow directions and flow-stop state of sample fluids can be fully controlled with a flow rate less than 3 μL/min.In addition,abscisic acid(ABA),rhodamine B(RhB),6-benzyladenine(6-BA)and chlorantraniliprole(CAP)were detected for 5 cycles in 1 h with the proposed platform.Taking RhB as an example,the ion current intensity is about 1.7×107 and the RSD is 2.67%.The results showed that the proposed microfluidic platform achieved decent ionization efficiency,reproducibility and stability.2.In this section,a bean sprout culture chip was prepared and connected with the aforementioned microfluidic platform for on-line sampling and real-time monitoring of plant hormone metabolism in the whole growth cycle of bean sprouts.We achieved online culture of on-chip bean sprouts and monitored the metabolic processes of 6-BA and ABA in bean sprouts under different soaking conditions within a 60 h growth cycle.The corresponding results showed that the amounts of 6-BA was between 0.01 ppt-0.1 ppm,and its counterpart of ABA was between 0.001 ppt-0.01 ppb,being lower than the control group(0.001 ppt-0.1 ppb),when the soaking condition was 30 ppm 6-BA solution.The amounts of 6-BA was between 0.01 ppb-1 ppm,and the ABA amounts was between 1 ppt-0.01 ppm,being higher than the control group,when the soaking condition was 45 ppm 6-BA solution In a sum,the introduction of exogenous hormone 6-BA with different concentrations can in a certain extent either inhibit or stimulate the synthesis of endogenous hormone ABA.Compared with traditional hormone detections,the proposed analysis platform avoided the complex sample pretreatment process.3.Mass axis drifting always exists in mass spectrometry.In order to inhibit this problem,a high-precision mass-to-charge ratio analysis method based on the principle of pulsed reference mass calibration was developed in a chip-MS platform.During the detection of the samples,pulse-generated standard reference droplets was driven to pass through the electrospray region,and standard reference ions were thus introduced.With this method,we detected 12 organic small molecules,including pyridalyl,reserpine etc.The results showed that the relative deviation of m/z was between 0.0-8.6 ppm,when the analyte(m/z 226-912)concentration was 1 ppm,which was better than the absence of the corresponding mass calibration(13.7-74.7 ppm).Therefore,the results showed that the proposed mass calibration method was feasible.