本文關(guān)鍵詞： 葉綠素a測(cè)定 熒光檢測(cè)法 藻體標(biāo)準(zhǔn)品 懸浮穩(wěn)定劑 真空冷凍干燥法　出處：《河北科技大學(xué)》2016年碩士論文　論文類型：學(xué)位論文

【摘要】：葉綠素a含量是水體富營(yíng)養(yǎng)化指標(biāo),熒光法因測(cè)定時(shí)間短、可以實(shí)現(xiàn)現(xiàn)場(chǎng)在線測(cè)定等優(yōu)勢(shì)在水體葉綠a含量測(cè)定中得到廣泛應(yīng)用,但在測(cè)定過(guò)程,存在著藻細(xì)胞在水體分散不均勻、藻體懸浮體系不穩(wěn)定、易沉降,無(wú)藻體標(biāo)準(zhǔn)品等問(wèn)題,影響測(cè)量結(jié)果的準(zhǔn)確度。論文研究了利用水體懸浮劑制備藻體標(biāo)準(zhǔn)品用于葉綠素?zé)晒猬F(xiàn)場(chǎng)測(cè)定的制備技術(shù),技術(shù)研究對(duì)水體葉綠素?zé)晒夥y(cè)定的方法標(biāo)準(zhǔn)化有重要意義。文章簡(jiǎn)述了熒光檢測(cè)法對(duì)葉綠素a測(cè)定的國(guó)內(nèi)外研究進(jìn)展,微生物懸浮穩(wěn)定技術(shù)、保存技術(shù)的研究進(jìn)展。以葉綠素a的實(shí)驗(yàn)室熒光法測(cè)定方法為基礎(chǔ),得到如下結(jié)論:1)基于蛋白核小球藻的自身優(yōu)勢(shì)及實(shí)驗(yàn)優(yōu)勢(shì),確定蛋白核小球藻作為目標(biāo)藻體,建立了蛋白核小球藻的吸光度值與熒光強(qiáng)度值的標(biāo)準(zhǔn)曲線y=74.831x+16.94(R2=0.9945),最優(yōu)D(663 nm)范圍為0.219~0.777,對(duì)應(yīng)的熒光強(qiáng)度值為31.606~74.722。2)藻細(xì)胞沉降,導(dǎo)致其熒光值受靜置時(shí)間影響較大,靜置1 min時(shí),熒光值的衰減率達(dá)△F/F0=-9.96%,無(wú)法獲得準(zhǔn)確的藻濃度。3)藻體培養(yǎng)過(guò)程中向培養(yǎng)基中添加瓊脂粉0.8~1.0 g·L-1培養(yǎng)蛋白核小球藻,藻液的懸浮性、均勻性、穩(wěn)定性都明顯增強(qiáng)。4)在熒光法測(cè)定水樣葉綠素過(guò)程中,提出一種藻體懸浮穩(wěn)定技術(shù),在藻液熒光測(cè)定環(huán)節(jié)添加復(fù)合懸浮劑,即瓊脂0.06%、黃原膠0.20%、卡拉膠0.40%,T=1h時(shí)熒光值的變化率為-1.16%,并對(duì)實(shí)際水樣測(cè)定,T=1 h時(shí)熒光值的變化率小于±3.5%、精密度較高小于1.5%。5)對(duì)長(zhǎng)勢(shì)較好的蛋白核小球藻在最佳離心條件下,即離心轉(zhuǎn)速為4 500 r.min-1,離心時(shí)間為為10min濃縮100倍;保護(hù)劑:15%脫脂奶粉,體積比1:1混合;預(yù)凍方式:4℃預(yù)凍12 h+液氮(-196℃)冷凍12 h;凍干保存溫度:-20℃。保存時(shí)間:40 d(存活率高于40%);100 d(存活率高于30%)。
[Abstract]:Chlorophyll a content is a eutrophication index of water body. Because of the short measuring time, fluorescence method can be used widely in the determination of leaf green a content in water body, but it can be used in the determination process. There are some problems such as uneven dispersion of algal cells in the water body, unstable suspension system of algae body, easy settling, and no algal body standard, etc. In this paper, the preparation technology of algae standard for chlorophyll fluorescence in situ was studied. The technical research is of great significance to the standardization of the methods for the determination of chlorophyll a in water body. This paper briefly introduces the domestic and foreign research progress of fluorescence detection for the determination of chlorophyll a and the technique of microorganism suspension stabilization. Based on the fluorescence assay of chlorophyll a in laboratory, the following conclusions are obtained: 1) based on the advantages of Chlorella protein and its experimental advantages, Chlorella Proteinuca is identified as the target alga. The standard curve of absorbance value and fluorescence intensity value of Chlorella pyrenoidosa was established. The optimum range of DX 663nm was 0.2196nm and the corresponding fluorescence intensity value was 31.606n 74.722.2) the fluorescence value of Chlorella globosa was significantly influenced by standing time, and the fluorescence value was statically affected by 1 min. The attenuation rate of fluorescence value was as high as F / F _ 0 ~ (-9.96). It was impossible to obtain accurate algal concentration. 3) adding Agar powder (0.81.0 g 路L ~ (-1)) to the culture medium of Chlorella vulgaris, the suspension and homogeneity of algal fluid, were obtained by adding Agar powder (0.81.0 g 路L ~ (-1)) to the culture medium. In the process of determination of chlorophyll in water samples by fluorescence method, a suspension stabilization technique for algae was put forward, in which compound suspensions were added in algal liquid fluorescence determination. That is, Agar 0.06, Xanthan 0.20, carrageenan 0.40T + 1 h fluorescence change rate is -1.16, and the change rate of fluorescence value is less than 鹵3.5 at 1 h and higher precision is less than 1.5. 5) under the best centrifugation conditions. The centrifugal speed is 4 500 r.min-1, the centrifugation time is 10 min, the concentration is 100 times, the protective agent is 1: 15% skim milk powder, the volume ratio is 1: 1; The pre-freezing method was pre-frozen at 4 鈩，

本文編號(hào)：1516896