The Study On Preparation Of The Algae Standard For Fluorescence Determination Of Water Chlorophyll

The content of chlorophyll a,one of the eutrophication index of water,is a regular monitoring item in the environmental monitoring of water.In the process of the on the sport determination of chlorophyll a by fluorescence direction of algae,the following problems exist: the uneven distribution and easy sedimentation of algae cells,unstable suspension system,the decline of measurement result accuracy,etc.To overcome the above problems,the good samples of algae must be acquired.Therefore,a large numberof the studies on the preparation of good samples of algae are in progress.In the present paper,the research results in China and the world on determination of chlorophyll a by fluorescence direction of algae,microrganism suspension,stability and preservation are summarized.Based on the laboratory methods of the determination of chlorophyll a by fluorescence detection,the following results are obtained:1)Based on its own advantages and experimental advantages of Chlorella pyrenoidosa,it was choose as the target algae and the regression equation between its optical density value and its fluorescence intensity was y=74.831x+16.94(R2=0.9945).The best D(663 nm)range is 0.219~0.777,the corresponding fluorescent intensity is31.606~74.722.2)Due to the sedimentation of algae cells,the fluorescence value of the algae is greatly influenced by standing duration of the algae samples.When the standing duration of the algae sample is 1 min,the attenuation rate of fluorescence is ?F/F0=-9.96% and the accurate algae density cannot be determined.3)When agar powder of 0.8~1.0 g·L-1 is added to the culture medium of Chlorella pyrenoidosa,the suspension,even distribution and stability of the algae solution is improved dramatically.4)When the chlorophll a in water is determined by fluorescent detection,if the following suspending agent is added,the algae suspension is improved: agar 0.06%,xanthan-gum 0.20%,sodium alginate 0.4%.When T=1 h,the variation rate of fluorescent intensity is-1.16%;in the determination of the actual water sample,when T=1 h,the variation rate of fluorescent intensity is less than±3.5%,the accuracy is relative high.5)For the Chlorella pyrenoidosa with a good growth,when centrifuge rotate is 4 500 r.min-1and the centrifuge time is 10 min,it is concentraed 100 times.The preservationagent is 15% skim milk powder,mixed in the volum proportion 1:1.The preparatory freeze: 4? freeze 12 h + liquid nitrogen(-196?)12h.The time for the preservation of the freeze-dried Chlorella pyrenoidosa is-20 ?.The survival rate of the freeze-dried Chlorella pyrenoidosa is: higher than 40% if it is preserved for 40 d,higher than 30% if it is preserved for 100 d.