Performance Check of Vegetation Fluorescence Imaging Lidar through In Vivo and Remote Estimation of Chlorophyll Concentration Inside Plant Leaves

Laser-induced fluorescence imaging lidar was developed for in vivo plant/vegetation monitoring. Fluorescences of poplar tree leaves growing naturally at a distance of 60 m from the lidar were successfully detected as two spectral images at wavelengths of 685 nm and 740 nm. By comparing chlorophyll concentration quantified with high performance liquid chromatography, it was confirmed that the intensity ratio of the two wavelengths was converted into the chlorophyll concentration inside the leaves. The intensity ratio of the images reflected the status of the poplar tree in the process of senescence so well that it was possible to assess the living status of the tree as a numerical value. The performance characteristics and the potential of the laser-induced fluorescence imaging lidar for monitoring the physiological activities of plants and vegetation are described.     

基于激光诱导叶绿素荧光寿命成像技术的植物荧光特性研究

针对植物荧光遥感探测中信号易受干扰的问题, 提出了一种用于评估植物生长状况及环境监测的荧光寿命成像技术. 采用凹透镜对355 nm波长的激光扩束, 再照射植物激发叶绿素荧光, 由增强型电荷耦合器件接收荧光信号. 采用时间分辨测量法, 连续用相同激光脉冲照射植物以激发相同的荧光信号, 同时不断改变激光脉冲触发探测器启动的延时时间, 从而能够得到完整的离散荧光信号分布图像. 对植物特定位置点产生的离散荧光信号进行拟合, 再运用一种改进型的迭代解卷积法可反演高精度的荧光寿命; 进而反演图像各点的荧光寿命以生成植物的荧光寿命分布图. 该方法所绘制的荧光寿命图比荧光强度图能更准确地反映植物内部的叶绿素含量, 并对活体植物叶绿素荧光寿命的物理特性进行了初步研究, 证明叶绿素荧光寿命与植物生理状态存在一定关联; 并且叶绿素荧光寿命与活体植物所处环境存在着复杂的关系. 未来将与生物物理学家们合作, 继续探寻叶绿素荧光寿命与植物生存环境的关系.     

Water Deficit and Salt Stress Diagnosis Through LED Induced Chlorophyll Fluorescence Analysis in Jatropha curcas L.

LED induced chlorophyll fluorescence analysis is employed to investigate the effect of water deficit and salt stress upon the growth process of Jatropha curcas L.. Red(Fr) and far-red(FFr) chlorophyll fluorescence around 685 nm and 735 nm, respectively, were observed and examined as a function of the stress intensity(salt concentration and water deficit). The fluorescence ratio Fr/FFr which is a valuable nondestructive and nonintrusive indicator of the chlorophyll content of leaves was exploited to monitor the jatropha plants under stress. The data indicated that salinity plays a minor role in the chlorophyll concentration of leaves for NaCl concentrations in the 25 to 200 mM range. The fluorescence ratio also permitted the detection of damage caused by water deficit in the early stages of the plants growing process. A significant variation of the Fr/FFr ratio was observed in the first 10 days of the experiment, and before signs of visual stress became apparent. The results suggest that the Fr/FFr ratio is an early-warning indicator of water deficit stress     

Spectral changes in the fluorescence of chlorophyll during photosynthesis induction

With the help of a light-emitting diode with a radiation maximum at 407 nm and an S-2000 UV-VIS spectrometer connected with a computer, the spectral changes in the fluorescence of chlorophyll from acacia leaves (Acacia sp.) preliminarily subjected to a dark adaptation are studied. It is found that, in the slow induction phase, the Kautsky effect manifests itself in the “compression” of the intensity of the chlorophyll fluorescence spectrum, with this spectrum exponentially approaching the steady-state shape with a time constant of 10–20 s. Once the steady-state fluorescence spectrum of chlorophyll was established, the amount of energy delivered to the upper singlet level is one and a half times greater than at 735 nm. The ratio between the energies spent for photochemical processes of photosynthesis and for fluorescence depends on the wavelength and the instant of time of the induction period. In the steady-state state of the chlorophyll fluorescence, the values of this ratio at 685 and 735 nm are equal to 5.9 and 3.4, respectively.     

荧光激光雷达技术探测水面油污染系统仿真研究

为实现对水面油污染的探测, 根据荧光激光雷达系统的发展趋势, 采用激光诱导荧光技术, 建立了适用于水面油污染探测荧光激光雷达的系统模型. 提出采用单激光器结合增强电荷耦合器件的小型荧光激光雷达探测系统, 通过分析激光器单脉冲能量与探测浓度之间的关系, 结合荧光激光雷达系统参数, 对系统模型的探测能力与信噪比等进行了数值仿真. 结果表明, 系统选用单脉冲能量50 μJ的355 nm Nd:YAG激光器作为激发光源, 白天在7 m的距离处探测信噪比可以达到10, 满足实验室搭建模拟系统的要求. 针对实际探测水面油污染, 提出采用增大激光器功率的方法, 并通过模拟计算验证了采用50 mJ的单脉冲能量激光器在230 m的探测距离处可得到与实验室相同的结果, 对实际系统的搭建具有指导意义.     

饱和激发下叶绿素荧光非线性变化及影响分析

叶绿素浓度是海洋初级生产力的重要指标之一,激光诱导荧光技术可以实现海水叶绿素浓度的快速测量。测量叶绿素浓度的传统激光诱导荧光原理,是利用叶绿素荧光与水体Raman散射的强度比值（I_F/R）进行反演,即叶绿素浓度n_chl＝CI_F/R,其中C为系统常量。这是依据叶绿素荧光685 nm、水体Raman散射强度都与激发光强呈线性关系。然而,该理论并没有考虑诱导荧光饱和现象的存在。当诱导激光强度达到一定程度后,685 nm荧光强度随激发光强非线性变化。另外,值得注意的是,水体Raman散射并不存在信号饱和现象。为了探讨饱和激发造成荧光非线性变化的影响,在激光诱导荧光技术测量叶绿素浓度的实验中,设计两种测量方案,即：不同激光功率诱导单一浓度样本的荧光测量,和固定激光功率时不同浓度样本的荧光测量。实验中利用Nd∶YAG三倍频激光355 nm激发获得叶绿素溶液的404 nm处 Raman散射和685 nm荧光。实验结果分为2部分进行讨论：（1）为了分析饱和激发造成荧光变化的非线性特性,通过调节激发光功率测量溶液的受激发射光谱,发现水体Raman散射强度与激发光强呈线性关系,而685 nm荧光强度出现饱和激发下的非线性变化。而且,随叶绿素浓度的增加,685 nm荧光的非线性趋势更为明显,Raman散射强度与激发光强的线性关系中斜率变小。数据分析表明,685 nm荧光数据拟合的4阶多项式和Raman散射效率值,可以定性地表征685 nm荧光的饱和程度。（2）考虑实际海洋激光雷达探测叶绿素浓度应用中存在饱和激发荧光非线性现象,为了分析荧光非线性对传统叶绿素浓度反演理论适用性的影响,在固定激发光强情况下对不同浓度叶绿素溶液的发射光谱进行测量。将激发光功率调节至52.00,80.70,132.10和197.30 mW·cm-2,获取相应激发光强下685 nm荧光与水体Raman散射的强度比值和叶绿素浓度之间的关系。实验表明,激发光强不变的情况下,685 nm荧光与水体Raman散射的强度比值,与叶绿素浓度仍满足线性关系。但是,在较高光强激发时,饱和激发造成的叶绿素荧光非线性变化,导致利用传统激光诱导荧光理论反演的叶绿素浓度值偏小。因此,需要对饱和激发下荧光非线性的影响进行修正,其关系为I_F/R=n_chl/C+C_F,修正值C_F不可忽略。另外,值得一提的是,修正关系中系统常量C随激发光强增加而增大。研究表明,饱和激发造成的荧光非线性,会对激光诱导荧光技术测量叶绿素浓度产生影响,但由于造成荧光非线性因素的复杂性,仅通过荧光数据拟合获得的多项式,无法定量说明其影响权重。然而,当激发光强不变时,可以实验测量获得基于激光诱导荧光原理的修正关系,从而准确反演叶绿素浓度。     

Analysis of Chlorophyll Fluorescence Spectra in Some Tropical Plants

Laser-induced chlorophyll fluorescence (LICF) emission spectra of leaves of some tropical plants were measured using a compact fiber-optic fluorosensor with a continuous-wave blue diode laser as exciting source and an integrated digital spectrometer. Different chlorophyll-fluorescence signatures of light-green, fully-green, and yellow leaves were monitored at room temperature. Deconvolution procedure was used to determine fluorescence band position and width. Calibration of the fluorescence ratio F690/F730 relative to the 404 nm excitation is done from the curve-fitted parameter. An erratum to this article can be found at     

水稻叶绿素浓度对荧光光谱的影响

为了加强对水稻生长状况的监测,指导水稻的田间施肥,提高施肥利用效率,以增加农作物产量、提高粮食品质,实验室搭建了基于激光诱导荧光技术的荧光探测系统,以研究水稻叶片的叶绿素含量与荧光峰值比之间的相关性。文中测量样本为水稻分蘖期和拔节期的倒二叶,栽培地区位于中国江汉平原。文中先采用凯氏定氮法和相应公式结合测得水稻叶片的叶绿素含量(mg·g-1),再用搭建的荧光探测系统采集了水稻叶片不同叶绿素含量的荧光光谱(激发波长为355 nm)。获得了水稻叶片在不同叶绿素含量下的荧光光谱数据库,定量分析了荧光峰值比F₇₄₀/F₆₈₅(荧光谱峰740 nm、685 nm处的荧光强度比)与叶绿素含量的相关性,发现叶绿素含量的变化对荧光光谱特性影响明显。由实验数据分析可知荧光参数中的峰值比(F₇₄₀/F₆₈₅)与叶绿素含量呈现很好的线性正相关,相关系数(R2)在水稻的分蘖期和拔节期分别达到了0.901 3和0.912 5。实验分析结果表明诱导荧光光谱技术具有方便、快捷、无损等优点,在农作物生长状况的遥感定量监测等方面也具有一定发展潜力。     

Stress-Induced Alteration of Chlorophyll Fluorescence Polarization and Spectrum in Leaves of <Emphasis Type="Italic">Alocasia macrorrhiza</Emphasis> L. Schott

The value of intrinsic chlorophyll fluorescence polarization, and the intensity in emission spectrum were investigated in leaf segments of Alocasia macrorrhiza under several stress conditions including different temperatures (25–50°C), various concentrations of NaCl (0–250 mM), methyl viologen (MV, 0–25 μM), SDS (0–1.0%) and NaHSO₃ (0–80 μM). Fluorescence emission spectrum of leaves at wavelength regions of 500–800 nm was monitored by excitation at 436 nm. The value of fluorescence polarization (P value), as result of energy transfer and mutual orientation between chlorophyll molecules, was determined by excitation at 436 nm and emission at 685 nm. The results showed that elevated temperature and concentrations of salt (NaCl), photooxidant (MV), surfactant (SDS) and simulated SO₂ (NaHSO₃) treatments all induced a reduction of fluorescence polarization to various degrees. However, alteration of the fluorescence spectrum and emission intensity of F₆₈₅ and F₇₃₁ depended on the individual treatment. Increase in temperature and concentration of NaHSO₃ enhanced fluorescence intensity mainly at F₆₈₅, while an increase in MV concentration led to a decrease at both F₆₈₅ and F₇₃₁. On the contrary, NaCl and SDS did not cause remarkable change in fluorescence spectrum. Among different treatments, the negative correlation between polarization and fluorescence intensity was found with NaHSO₃ treatments only. We concluded that P value being measured with intrinsic chlorophyll fluorescence as probe in leaves is a susceptible indicator responding to changes in environmental conditions. The alteration of P value and fluorescence intensity might not always be shown a functional relation pattern. The possible reasons of differed response to various treatments were discussed.     

Laser-Induced Chlorophyll Fluorescence: A Technique for Detection of Dimethoate Effect on Chlorophyll Content and Photosynthetic Activity of Wheat Plant

Laser-induced chlorophyll fluorescence (LICF) spectra and fluorescence induction kinetics (FIK) curves of wheat plant leaves treated with different concentrations (50, 100 and 200 ppm) of dimethoate are recorded. LICF spectra are recorded in the region of 650-780 nm using violet diode laser (405 nm) and FIK curve at 685 and 730 nm with red diode laser (635 nm) for excitation. The fluorescence intensity ratios (FIR) are determined from LICF spectra and vitality index (R(fd)) from FIK curves. These parameters along with photosynthetic pigment contents and growth parameters are used to analyze the effect of dimethoate on wheat plants. The result indicates that lower concentration of 50 ppm shows stimulatory response while higher concentrations of dimethoate are hazardous for growth, photosynthetic pigments and activity of wheat plants.     

双波长荧光雷达探测大气生物气溶胶的性能分析

生物气溶胶在大气中扩散极易传播和发生各种流行疾病,也是生物武器投放的主要形式,实现生物气溶胶实时、远距离的探测显得尤为重要。构建了一台双波长荧光雷达用于大气中生物气溶胶的预警和识别。该雷达系统采用Nd∶YAG固体激光器作为激励光源,基频1 064 nm、四倍频266 nm作为工作波长。基于激光诱导荧光雷达探测原理,对红外波段的弹性散射信号和紫外波段诱导的荧光信号进行数值分析。结果显示,在探测误差小于10%的情况下,距离为1.0 km时,单激光脉冲测量得到白天和夜晚细菌孢子的最小探测浓度分别为15 100个颗粒·L-1和8 386个颗粒·L-1;当脉冲数累加到10 000时,白天和夜晚的细菌孢子最小探测浓度显著改善,分别为144个颗粒·L-1和77个颗粒·L-1。分析结果还表明,通过红外波段确定细菌孢子云团位置后,为了提高系统对细菌孢子的探测性能,可增加紫外激光脉冲数量,延长荧光信号采集时间。     

苹果树冠层不同光照区域叶绿素荧光性状计算方法

植物冠层内光照分布决定了植物物理过程与生态环境之间的交互关系,是植物冠层叶绿素荧光动力学研究的重要基础。光谱技术在构建植株冠层组分含量的预测模型中发挥了重要的作用,而针对自由纺锤形苹果树冠层不同光照区域的叶绿素荧光性状研究文献报道较少。因此,以自由纺锤形苹果树为研究对象,将苹果树冠层空间划分为5层,每层划分为边长50 cm的立方体网格,并测定各个网格空间的光照强度,确定其光照分布情况。以此为基础,获取不同光照区域的光谱数据及对应叶绿素荧光性状参数值,通过光谱一阶微分去除系统误差干扰,确定红边区域(660～760 nm)的一阶微分光谱曲线,利用BP神经网络建立红光范围(680～760 nm)内一阶微分光谱最大值与叶绿素荧光性状参数关系模型,创新性地提出了苹果树冠层不同光照区域的叶绿素荧光性状计算方法。并采用决定系数、均方根误差、平均绝对百分比误差等对模型进行有效性评价,试验结果表明：该叶绿素荧光计算方法的计算精度均在80%以上,研究成果可为苹果树的整形修剪,获取充足光照分布提供理论依据。     

激光诱导荧光雷达探测系统仿真研究

激光诱导荧光技术是一种应用广泛的新型遥测技术,为了指导实际激光诱导荧光探测系统的搭建,介绍了荧光检测原理,设计了激光诱导荧光雷达探测系统。以测量水面石油类污染物为例,通过分析激光诱导荧光雷达探测系统中发射系统及接收系统主要参数对系统信噪比的影响,可根据测量条件选择合适的系统参数。在实验室条件下,选用单脉冲能量为50 μJ的Nd:YAG激光器,接收视场角0.1 mrad,滤光片带宽60 nm的接收系统,可以检验浓度为10个/L的粒子。     

基于生物光学算法的海洋赤潮监测

机载海洋激光雷达具有高效搜索的能力,已成为海洋环境监测的重要手段。以机载蓝绿激光雷达监测海洋赤潮为研究应用背景,通过检测海洋中叶绿素的质量浓度来实现对海洋赤潮消长信息的获取。在分析海水固有光学性质的基础上,基于生物光学算法,构建了机载蓝绿激光雷达后向散射信号检测的的叶绿素质量浓度监测模型。仿真计算证明该模型能够有效探测出叶绿素质量浓度的大小,为机载海洋赤潮监测增加了一种方法。     

叶绿体不同浓度下光诱导延迟荧光光谱研究

针对《光子学报》文献［1］中实验结果与已有报道相关光谱特征的显著差异,利用多种光谱学手段对不同浓度下叶绿体延迟荧光行进了研究.实验结果表明：PSⅠ反应中心叶绿素P700对PSⅡ所发685 nm成份的吸收随叶绿体浓度的增加而增强,从而导致PSⅠ发出的730 nm成份增强,而使得延迟荧光光谱中730 nm成为主峰,甚至685 nm成份的消失.该研究结果表明：叶绿体延迟荧光光谱中730 nm成份,是由PSⅠ作用中心叶绿素P700对PSⅡ中所发685 nm成份延迟荧光的重吸收,产生激发态所发出的荧光.该结论有助于延迟荧光光谱中各成份产生机理的进一步研究.     

琅琊山景区不同叶绿素条件下树种叶片光谱差异分析

叶绿素含量高低反映植物健康状况,研究景区树种叶片叶绿素绝对值(SPAD)不同的光谱变化规律能为叶绿素高光谱监测波段识别与景区树种管理提供理论支撑。从琅琊山景区灌木和乔木类选取9个常见树种,探讨相同树种叶片SPAD值变化时的光谱差异,同时,横向对比相同SPAD值不同树种叶片的光谱特征,并深入分析不同树种叶片SPAD值与单波段原始光谱、光谱倒数、一阶微分、二阶微分及波段组合差值指数、归一化指数、比值指数、一阶微分归一化指数、一阶微分比值指数之间的关系。结果表明：9个所测树种叶片随着叶绿素SPAD值的升高,光谱变化规律各不相同,在可见光波段区分明显,总体上,光谱反射率最高的样本组SPAD值较低;叶绿素SPAD值相同时,在可见光波段,桂花较其余树种反射率整体较高; 在780~1 350 nm波段,广玉兰叶片反射率始终排前三,其余波段变化规律不明显;原始光谱反射率的二阶微分与海桐叶片SPAD值相关系数最大,一阶微分与其余8种相关性最高;与灌木、落叶乔木叶片SPAD值相关系数最大的光谱指数分别为差值指数、一阶微分归一化指数,与常绿乔木、不分树种相关系数最大的为一阶微分比值指数。     

测定叶绿素a浓度,漫散射衰减系数和离水辐亮度的海洋激光雷达

研制了一台多参数的主被动船载蓝绿光海洋激光雷达系统（ＢＬＯＬ），它可用于我国东海海域的海色遥感印证。该海洋激光雷达系统能被动地测量离水辐亮度Ｌｗ，主动测量漫散射衰减系数ｋ和叶绿素ａ浓度Ｃ。雷达系统安装在青岛海洋大学的调查船上（约２５００吨位），曾在２１个站位进行了实验。实验的区域从上海东开始到冲绳岛附近。用该海洋激光雷达系统测量的结果与用传统的方法测量的结果符合的很好。     

激光诱导叶绿素荧光寿命的测量及其特性分析

提出了一种用于评估植物生长状况及环境监测的激光诱导叶绿素荧光寿命测量方法. 采用波长355 nm的激光作为光源激发叶绿素荧光, 由光电倍增管接收其荧光信号, 由于被测叶绿素荧光衰减函数与激光脉冲、仪器响应函数卷积在一起, 根据它们的特性, 运用时间分辨测量法分别测得叶绿素荧光及其背景信号, 并结合一种新型解卷积算法可分离出真实的叶绿素荧光衰减函数, 从而获取叶绿素的荧光寿命. 测试结果表明: 该方法能够实现叶绿素荧光寿命的高精度实时监测, 对不同叶绿素含量的溶液荧光寿命进行了测试, 证明叶绿素含量与其荧光寿命具有相关性, 并且拟合了叶绿素含量与荧光寿命的标定曲线. 关键词： 荧光寿命 激光诱导荧光 时间分辨测量法 叶绿素含量     

叶绿体延迟荧光中730nm成分产生机理的光谱学研究

叶绿体685 nm延迟荧光成分被认为源于PSⅡ作用中心的电荷复合。利用多种光谱学测量手段研究了叶绿体延迟荧光光谱中730 nm峰的产生机制。不同浓度下叶绿体延迟荧光光谱实验结果表明：初始随浓度的增加,延迟荧光光谱中685和730 nm成分强度均增强;当浓度增加到7.8 μg·mL-1时,685 nm成分强度达最大,730 nm成分强度继续上升;当浓度增加到31.2 μg·mL-1时,延迟荧光光谱中730 nm成分强度达最大,而685 nm成分已明显下降。吸收光谱实验结果表明：A₆₈₅/A₇₃₀在叶绿体浓度增加的过程中几乎不变。叶绿体730 nm荧光成分的激发光谱实验结果表明：685 nm光对730 nm荧光有较高的激发效率。上述实验结果表明叶绿体延迟荧光光谱中730 nm峰是由PSⅡ所发685 nm成分激发PSⅠ所产生的荧光。同一浓度下叶绿体延迟荧光光谱波形随延迟时间(1～9 s)的不变性进一步证明了这一结论。     

Statistical modelling of laser induced fluorescence in plant cover

A system of mutually coupled radiative transfer equations is proposed as a formal basis for the numerical analysis of a broad range of spectroscopic effects associated with the propagation of laser radiation in the environment. In particular, this concerns the reabsorption of laser induced fluorescence which inevitably occurs in dense disperse media containing two or more fluorophors. One practical application is for the pressing problem of lidar monitoring the state of plant cover, in particular the concentration of chlorophyll, which regulates the vital activity of plants. A new concept for an optical model of plant cover has been developed in which leaves are not treated as separate scattering elements, but as local volumes of a multiphase medium with a complex polydisperse structure. A modified Monte-Carlo algorithm is created for imitating the fluorescence and reabsorption processes. Test calculations confirm the adequacy of this approach. Translated from Zhurnal Prikladnoi Spektroskopii, Vol. 76, No. 3, pp. 411–418, May–June, 2009.