Study On Fluorescence Kinetic Technique For Measuring Phytoplankton Photosynthetic Rate

As the primary producer of food chain in water, the phytoplankton is a indicator of water health condition and fundamentally affects global biogeochemical cycles and climate changes. Accurate measurement of photosynthetic rate of phytoplankton is significant for monitoring aquatic ecological environment, preventing algal bloom and red tide, evaluating fishery resources and predicting global climate change. The traditional measurement methods of photosynthetic rate such as Black-White Bottle,14C and 180 requires "field sampling-incubation cultivating-offline analyzing" have the defects of long measuring time, low efficiency, thus are difficult to meet the needs of field fast measurement. As a new rapid and efficient measurement technique, the chlorophyll fluorescence method is mainly used in the measurement of chlorophyll concentration and photosynthetic parameters. There is few analysis of photosynthetic rate, and the measurement system is complex with low signal-to-noise ratio.According to the demand of rapid measurement of phytoplankton photosynthetic rate and the disadvantages of chlorophyll fluorescence method, this paper studied a phytoplankton photosynthetic rate measurement technology based on fluorescence kinetics.First, a fluorescence kinetic curve measuring system for phytoplankton was established. Based on the analysis of excitation light interference suppression,photosynthetic electron transport chain blocking positions, as well as high speed and sensitive signal detection technique, the design of the measurement system concerned on three aspects: excitation and emission light path, excitation intensity modulation,and fluorescence signal detection. The excitation intensity adaptive control technology was studied, and a variable optical pulse control circuit is designed,realizing the adaptive control of measuring light intensity for different measurement process, thus solves the precise regulation of photosynthetic electron transport chain block sites; The fast acquisition of fast phase and high sensitivity of relaxation fluorescence detection technology was studied, and low noise photoelectric signal conversion circuit and multi-time-alternation sampling circuit were designed,realizing the accurate sampling of fast phase fluorescence kinetics in microsecond level. A detection circuit with high sensitive based on synchronous integral technique was designed to obtain the weak relaxation phase fluorescence. Finally, a high sensitivity and high speed measurement system for fluorescence kinetic was established. Experiments show that the sampling rate for 200?s fast phase fluorescence kinetics is 1.50MHz with 16 bits A/D resolution, and the signal-to-noise ratio of phytoplankton with 0.125 ?g/L chlorophyll concentration is above 11.2dB.Second, the phytoplankton photosynthetic rate inversion method from fluorescence kinetic curve was developed. Based on the fast phase and relaxation phase fluorescence kinetic curve, the inversion methods including algorithm of sliding window slope for maximum fluorescence yield, linear least squares algorithm for photochemical efficiency (Fv/Fm) and functional absorption cross section (PSI) and discrete iterative algorithm for PQ average reduction time constant (?QA ) were studied,realizing the inversion of phytoplankton photosynthetic parameters. Based on the energy transfer process of photosynthesis, the photosynthetic rate analysis method was established using QA and PQ as analyzing nodes. The Megard model was then used to analyze the photosynthetic rate light response curves, and the biological characterization parameters of photosynthetic rate such as maximum photosynthetic rate (PMax), initial slope (?), maximum intensity (IMax) were obtained.Finally, a comparative experiment of phytoplankton photosynthetic rate of based on fluorescence kinetics was carried out. By measuring the chlorophyll fluorescence kinetics curves of Chlorella pyrenoidosa under conditions of different light, nutrient and Cu2+ concentration, the photochemical quantum efficiency, functional absorption cross section and PQ reduction time constant average photosynthetic parameter were obtained. the correlation coefficient of photochemical quantum efficiencies (Fv/Fm)that measured by fluorescence kinetic and Water-PAM is more than 0.95, and the variation tendency of other parameters were also consistent with theoretical analysis.The photosynthetic electron transport flux of Chlorella pyrenoidosa that measured by fluorescence kinetic under DCMU and salt stress as well as under different light,nutrient comditions possessed good linear correlation with the photosynthetic oxygen release measured by Chlorolab2, and the minimum correlation coefficients were 0.876, 0.889, 0.968 and 0.948.