phyto‑pam 全球第一款可自动对浮游植物分类的荧光仪
schreiber教授因发明pam系列调制叶绿素荧光仪而获得首届国际光合作用协会（ispr）创新奖
1983年，walz公司首席科学家、德国乌兹堡大学的ulrich schreiber教授设计制造了全世界第一台调制荧光仪——pam-101/102/103，并在植物生理、生态、农学、林学、水生生物学等领域得到广泛应用，出版了大量高水平研究文献。但该仪器由于采用光电二极管为检测器，因此只能检测高等植物、室内培养的微藻等叶绿素含量较高（> 10 mg l-1）的样品。
在pam-101/102/103出现的同时，schreiber教授就有了设计一台多波长调制荧光仪的构想。1988年，schreiber教授和他的博士后kolbowski博士第一次设计出了16波长（led）荧光分光光度计。
|  |
1995年，schreiber教授和kolbowski博士一起设计出了全世界第一台可对浮游植物自动分类的调制叶绿素荧光仪phyto-pam。phyto-pam采用调制技术，利用4种不要波长的led作为光源，利用光电倍增管作为检测器，可以对水样中的蓝藻、绿藻、硅藻/甲藻自动分类，并分别测量它们的叶绿素含量和光合活性。
phyto-pam由于采用光电倍增管作为检测器，因此检测限达到 0.1 μg l-1 chl。根据研究对象和研究目的不同，可有3套系统供您选择。
phyto-pam是水域生态学、海洋与湖沼学、水质监测等领域的有效工具。
系统描述：
脉冲-振幅-调制（pulse-amplitude-modulation, pam）技术的测量原理是基于对调制测量光激发的荧光信号的选择性放大。在phyto-pam浮游植物荧光仪中，微秒级的测量光脉冲是由4种不同颜色的发光二极管（led）阵列发出的：蓝色（470 nm）、绿色（520 nm）、浅红色（645 nm）和深红色（665 nm）。不同颜色的测量光脉冲在高频率下交替应用，就可以获得4种波长的光激发出的半同步的荧光信号。结合不同藻类的参考光谱（reference spectrum）就可区分不同藻类，并分别测量它们的光合活性和叶绿素含量。
phyto-pam可以对蓝藻、绿藻和硅/甲藻进行分类。由于硅藻和甲藻的色素组成差别不大，目前技术上还很难对它们进行区分。要想对它们区分，除了考虑色素组成外，更重要的是考虑捕光色素-蛋白复合体的结构特别是横向截面积，这必须结合“泵”和“探针”法测量荧光。目前walz公司正结合“泵”和“探针”法开发对微藻分类更多、更精确的仪器。phyto-pam还可以测量这些藻类的叶绿素浓度（检测限为0.1 μg l-1 chl）。phyto-pam更加强大的功能是可以探测自然水样中蓝藻、绿藻和硅/甲藻的光合活性和光适应状态。
phyto-pam采用微型光电倍增管作为检测器，可以检测及其微弱的灵敏变化，同时还具备强光自动关闭的保护功能，因此仪器操作和维护更加容易。
特点：
1) 全世界第一台可对浮游植物自动分类的调制叶绿素荧光仪
2) 4波长光源：470、520、645和665 nm
3) 对蓝藻、绿藻和硅/甲藻进行分类
4) 可选配室内系统（i）、野外系统（ii）和测附着藻类/大型藻类的系统（iii）
5) 灵敏度高，检测限为0.1 μg l-1 chl
6) 专业phytowin操作软件，数据收集、分析和存贮功能强大
7) 用户可利用培养的微藻做参考光谱，非“黑匣子”
8) 可在野外测量后根据水体藻类组成利用优势种（一种或多种）的参考光谱校对实验结果
功能：
1） 可对蓝藻、绿藻和硅/甲藻自动分类（定性）
2） 可自动测量水样中蓝藻、绿藻和硅/甲藻的叶绿素含量（定量）和总叶绿素含量
3） 可同时测量水样中蓝藻、绿藻和硅/甲藻的光合作用和总光合活性
4） 可测量光合作用的量子产量和相对电子传递速率
5） 可自动记录量子产量和相对电子传递速率的快速光响应曲线
6） 用户可做自己的参考光谱
7） 可连接记录仪或示波器记录原始荧光诱导动力学曲线
应用领域：
多用于水生生物学、水域生态学、海洋学、湖沼学、水质监测和预警、微藻生理学、微藻抗逆性、环境科学、生态毒理学、极地藻类（冰藻）研究等领域，对于了解自然水体中藻类种群的动态变化、水华/赤潮预警、野外水体中光合作用的时空变化、校正初级生产力的计算等有较大帮助。
系统组成：
phyto-pam的主机连接不同的检测器可以组成3套不同的测量系统：
系统i：实验室版本，利用光学单元ed-101us/mp和标准10×10 mm样品杯检测荧光
系统i的所有光电元件均需安装在铁架台上，适合实验室用。但由于主机phyto-c内置大容量电池，因此它也可以在野外或在船上使用。系统i的一个突出优点是光学单元ed-101us/mp的开放式设计，它允许安装不同的滤光片或不同颜色的光化光led阵列。与系统ii的phto-ed相比，10×10 mm样品杯中的光场分布更加均匀。同时，系统i还可以连接温度控制器us-t和微型磁力搅拌器phyto-ms。这些特点决定了系统i更加适合浮游植物光合作用的基础研究。
系统ii：野外便携式版本，利用phyto-ed和直径15 mm的样品杯检测荧光
在系统ii中，所有光电元件都整合在便携式的激发-检测单元phyto-ed中。phyto-ed密封防水。系统ii最适合在野外或在船上工作，当然室内也完全可以使用。
系统iii：光纤型版本，利用phyto-edf检测附着藻类或大型藻类的荧光
系统iii的光纤型激发-检测单元phyto-edf可以检测所有生长在表面的光合生物的光合作用。比较适合的测试材料包括附着藻类、底栖藻类、藻垫（microbial mats）和大型藻类等。由于采用光纤传导信号而且测量面积小，因此灵敏度比系统i和ii要低。但是由于附着藻类等材料的叶绿素含量远远高于水体中的浮游植物，因此系统iii的灵敏度完全可满足实验要求。
● 基础配置
○ 可选配置 | 系统i
（实验室版）
 | 系统ii
（野外版）
 | 系统iii
（光纤版）
 |
| 主机phyto-c | ● | ● | ● |
| 测量光led阵列phyto-ml | ● | | |
| 光化光led阵列phyto-al | ● | | |
| 光电倍增管pm-101p | ● | | |
| 光学单元ed-101us/mp | ● | | |
| 工作台st-101 | ● | | |
| 激发-检测单元phyto-ed | | ● | |
| 光纤型激发-检测单元phyto-edf | | | ● |
| 微型磁力搅拌器phyto-ms | ○ | | |
| 球状微型光量子探头us-sqs | ○ | ○ | ○ |
| 温度控制器us-t | ○ | | |
| 搅拌器water-s | | ○ | |
技术参数：
测量光：波长470、520、645和665 nm的测量光led。
光化光：波长655 nm的led；光化光强度0～2000 μmol m-2 s-1 par（系统i和ii）或0～1300 μmol m-2 s-1 par（系统iii）。
饱和脉冲：波长655 nm的led；饱和脉冲强度4000 μmol m-2 s-1 par（系统i和ii）或2600 μmol m-2 s-1 par（系统iii）。
信号检测：光电倍增管，带短波截止滤光片（λ>710 nm）；选择性锁相放大器。
测量参数：ft, f(或fo), fm(或 fm’), δf, y(δf/ fm’或fv/fm), etr和chl浓度等。
环境温度：-5～＋45 ℃，已在极地成功应用。 部分文献[1] howeth jg, leibold ma. planktonic dispersal dampens temporal trophic cascades in pond metacommunities. ecology letters 2008;11 (3):245-57.
[2] ingleton t, kobayashi t, sanderson b, patra r, macinnis-ng cmo, hindmarsh b, bowling lc. investigations of the temporal variation of cyanobacterial and other phytoplanktonic cells at the offtake of a large reservoir, and their survival following passage through it. hydrobiologia 2008;603 (1):221-40.
[3] schmitt-jansen m, altenburger r. community-level microalgal toxicity assessment by multiwavelength-excitation pam fluorometry aquatic toxicology 2008;86 (1):49-58.
[4] wang g, chen k, chen l, hu c, zhang d, liu y. the involvement of the antioxidant system in protection of desert cyanobacterium nostoc sp. against uv-b radiation and the effects of exogenous antioxidants ecotoxicology and environmental safety 2008;69 (1):150-7.
[5] zhang m, kong fx, wu x, xing p. different photochemical responses of phytoplankters from the large shallow taihu lake of subtropical china in relation to light and mixing. hydrobiologia 2008;603 (1):267-78.
[6] 康丽娟, 潘晓洁, 常锋毅, 李敦, 沈银武, 刘永定. hco3-碱度增加对铜绿微囊藻光合活性和超微结构的影响. 武汉植物学研究 2008;26 (1):70-5.
[7] alsterberg c, sundbäck k, larson f. direct and indirect effects of an antifouling biocide on benthic microalgae and meiofauna journal of experimental marine biology and ecology 2007;351 (1-2):56-72.
[8] dimier c, corato f, saviello g, brunet c. photophysiological properties of the marine picoeukaryote picochlorum rcc237 (trebouxiophyceae, chlorophyta). journal of phycology 2007;43 (2):275-83.
[9] dimier c, corato f, tramontano f, brunet c. photoprotection and xanthophyll-cycle activity in three marine diatoms. journal of phycology 2007;43 (5):937-47.
[10] domis lnds, mooij wm, huisman j. climate-induced shifts in an experimental phytoplankton community: a mechanistic approach. hydrobiologia 2007;584:403-13.
[11] kim mk, park jw, park cs, kim sj, jeune kh, chang mu, acreman j. enhanced production of scenedesmus spp. (green microalgae) using a new medium containing fermented swine wastewater. bioresource technology 2007;98 (11):2220-8.
[12] schmitt-jansen m, altenburger r. the use of pulse-amplitude modulated (pam) fluorescence-based methods to evaluate effects of herbicides in microalgal systems of different complexity toxicological and environmental chemistry 2007;89 (4):665-81.
[13] tang d, shi s, li d, hu c, liu y. physiological and biochemical responses of scytonema javanicum (cyanobacterium) to salt stress journal of arid environments 2007;71 (3):312-20.
[14] xing w, huang w-m, li d-h, liu y-d. effects of iron on growth, pigment content, photosystem ii efficiency, and siderophores production of microcystis aeruginosa and microcystis wesenbergii current microbiology 2007;55:94-8.
[15] zhang m, kong f, xing p, tan x. effects of interspecific interactions between microcystis aeruginosa and chlorella pyrenoidosa on their growth and physiology. international review of hydrobiology 2007;92 (3):281-90.
[16] 张曼, 曾波. phytopam浮游植物分析仪用于微藻光合作用研究中几种参数设定的优化. 植物生理学通讯 2007;43 (1):148-52.
[17] ban a, aikawa s, hattori h, sasaki h, sampei m, kudoh s, fukuchi m, satoh k, kashino y. comparative analysis of photosynthetic properties in ice algae and phytoplankton inhabiting franklin bay, the canadian arctic, with those in mesophilic diatoms during cases 03-04. polar biosciences 2006;19:11-28.
[18] bontes bm, pel r, ibelings bw, boschker hts, middelburg jj, donk ev. the effects of biomanipulation on the biogeochemistry, carbon isotopic composition and pelagic food web relations of a shallow lake. biogeosciences 2006;3:69-83.
[19] liang y, beardall j, heraud p. changes in growth, chlorophyll fluorescence and fatty acid composition with culture age in batch cultures of phaeodactylum tricornutum and chaetoceros muelleri (bacillariophyceae). botanica marina 2006;49 (2):165-73.
[20] lürling m, geest gv, scheffer m. importance of nutrient competition and allelopathic effects in suppression of the green alga scenedesmus obliquus by the macrophytes chara, elodea and myriophyllum hydrobiologia 2006;556 (1):209-20.
[21] mulderij g, smolders ajp, van donk e. allelopathic effect of the aquatic macrophyte, stratiotes aloides, on natural phytoplankton. freshwater biology 2006;51 (3):554-61.
[22] quigg a, kevekordes k, raven ja, beardall j. limitations on microalgal growth at very low photon fluence rates: the role of energy slippage photosynthesis research 2006;88 (3):299-310.
[23] roessink i, belgers jdm, crum sjh, van den brink pj, brock tcm. impact of triphenyltin acetate in microcosms simulating floodplain lakes. ii. comparison of species sensitivity distributions between laboratory and semi-field. ecotoxicology and environmental safety 2006:in press.
[24] bontes bm, pel r, ibelings bw, boschker hts, middelburg jj, donk ev. the effects of biomanipulation on the biogeochemistry, carbon isotopic composition and pelagic food web relations of a shallow turf lake. biogeosciences discussions 2005;2:997-1031.
[25] casotti r, mazza s, brunet c, vantrepotte v, ianora a, miralto a. growth inhibition and toxicity of the diatom aldehyde 2-trans, 4-trans-decadienal on thalassiosira weissflogii (bacillariophyceae). journal of phycology 2005;41 (1):7-20.
[26] fietz s, bleiß w, hepperle d, koppitz h, krienitz l, nicklisch a. first record of nannochloropsis limnetica (eustigmatophyceae) in the autotrophic picoplankton from lake baikal. journal of phycology 2005;41 (4):780-90.
[27] heraud p, roberts s, shelly k, beardall j. interations between uv-b exposure and phosphorus nutrition. ii. effects on rates of damage and repair. journal of phycology 2005;41 (6):1212-8.
[28] jakob t, schreiber u, kirchesch v, langner u, wilhelm c. estimation of chlorophyll content and daily primary production of the major algal groups by means of multiwavelength-excitation pam chlorophyll fluorometry: performance and methodological limits. photosynthesis research 2005;83:343–61.
[29] shelly k, roberts s, heraud p, beardall j. interactions between uv-b exposure and phosphorus nutrition. i. effects on growth, phosphate uptake, and chlorophyll fluorescence. journal of phycology 2005;41 (6):1204-11.
[30] van der grinten e, janssen aphm, mutsert kd, barranguet c, admiraal w. temperature- and light-dependent performance of the cyanobacterium leptolyngbya foveolarum and the diatom nitzschia perminuta in mixed biofilms. hydrobiologia 2005;548 (1):267-78.
[31] wang g, chen l, li g, li d, hu c, chen h, liu y, song l. improving photosynthesis of microalgae by changing the ratio of light-harvesting pigments. chinese science bulletin 2005;50 (15):1622-6.
[32] hu z-q, liu y-d, li d-h. physiological and biochemical analyses of microcystin-rr toxicity to the cyanobacterium synechococcus elongatus. environmental toxicology 2004;19 (6):571-7.
[33] van der grinten e, janssen m, simis sgh, barranguet c, admiraal w. phosphate regime structures species composition in cultured phototrophic biofilms. freshwater biology 2004;49:369-81.
[34] van der grinten e, simis s, barranguet c, admiraal w. dominance of diatoms over cyanobacterial species in nitrogen-limited biofilms archiv fuer hydrobiologie 2004;161 (1):98-111.
[35] verspagen jmh, snelder eofm, visser pm, huisman j, mur lr, ibelings bw. recruitment of benthic microcystis (cyanophyceae) to the water column: internal buoyancy changes or resuspension? journal of phycology 2004;40 (2):260-70.
[36] 李阔宇, 宋立荣, 万能. 底泥中微囊藻复苏和生长特性的研究. 水生生物学报 2004;28 (2):113-8.
[37] lurling m. daphnia growth on microcystin-producing and microcystin-free microcystis aeruginosa
in different mixtures with the green alga scenedesmus obliquus. limnology and oceanography 2003;48 (6):2214-20.
[38] lürling m, verschoor am. fo-spectra of chlorophyll fluorescence for the determination of zooplankton grazing. hydrobiologia 2003;491:145-57.
[39] mulderij g, van donk e, roelofs2 gm. differential sensitivity of green algae to allelopathic substances from chara. hydrobiologia 2003;491:261-71.
[40] verschoor am, takken j, massieux b, vijverberg j. the limnotrons: a facility for experimental community and food web research. hydrobiologia 2003;491:357-77.
[41] young eb, beardall j. photosynthetic function in dunaliella tertiolecta (chlorophyta) during a nitrogen starvation and recovery cycle. journal of phycology 2003;39 (5):897-905.
[42] körner s, nicklisch a. allelopathic growth inhibition of se-lected phyplankton species by submerged macrophytes. journal of phycology 2002;38:862-71.
[43] schreiber u, gademann r, bird p, ralph pj, larkum awd, kühl m. apparent light requirement for activation of photosynthesis upon rehydration of desiccated beachrock microbial mats. journal of phycology 2002;38:125-34.
[44] nicklisch a, köhler j. estimatin of primary production with phyto-pam-fluorometry. ann. report inst. freshw. ecol. inland fish. berlin 2001;13:47-60.
[45] varotto c, pesaresi p, maiwald d, kurth j, salamini f, leister d. identification of photosynthetic mutants of arabidopsis by automatic screening for altered effective quantum yield of photosystem 2. photosynthetica 2000;38 (4):497-504.
[46] schreiber u. chlorophyll fluorescence: new instruments for special applications. in: garab g, editor. photosynthesis: mechanisms and effects. dordrecht: kluwer academic publishers, 1998.
[47] kolbowski j, schreiber u. computer-controlled phytoplankton analyzer based on 4-wavelengths pam chlorophyll fluorometer. in: mathis p, editor. photosynthesis: from light to biosphere. dordrecht: kluwer academic publishers, 1995. pp. 825-8.
