Development of a Transient Absorption Spectrometer for the Study of the Proton-motive Force (PMF) in Plant Thylakoid Membranes.

摘要

A transient-absorption spectrometer to study the electric field component of the proton-motive force (PMF) has been developed. Excitation by an actinic light source (655nm) causes absorption changes of pigments in the sample at 520nm on the order of 0.1%. Using a probe light source at 520nm, the change in transmission before and after actinic excitation can be detected and quantified. Because the change in transmission is small, a high signal to noise ratio (SNR) is required. To reduce the effect of optical shot noise and front end electrical noise a high intensity source at 520nm is utilized. However, continuous high intensity light at 520nm over time can cause actinic excitation in the sample, so the 520nm probe is pulsed at high optical intensity and low duty cycle to reduce the net incident flux.;The developed system includes 1) a LED driver to drive 1A current pulses through three high power probe LEDs and four high power actinic LEDs, 2) an analog front end which uses a 10mm by 10mm photodiode and a trans-impedance amplifier to detect the pulses, as well as several signal processing stages, 3) a microcontroller to generate the probe timing waveforms, digitize the detected signal, and send the data to a PC, and 4) a LabView computer interface to set input parameters, display the collected data, and save the data to a text file.;Experimental verification of system peformance was performed by testing the system with and without a plant sample. The results show that the system works as expected and is capable of measuring the data required to determine the electric field component of the PMF.