CN103900976A - Extraction method for ozone stress plant leaf pigment and spectral information - Google Patents

Abstract

The invention relates to an extraction method for plant leaf pigment and spectral information, and in particular relates to an extraction method for ozone stress plant leaf pigment and spectral information. In the process of plant cultivation, stress treatment is carried out through O3, in a plant growth season, plant spectral information is measured according to different plant growth periods, meanwhile physiological indexes in different plant growth periods are measured by adopting a direct extraction method (free from grinding); afterwards, the measured plant spectral information is converted into spectral parameters, and correlation analysis is carried out on the spectral parameters and corresponding leaf physiological indexes through SPSS (Statistic Package for Social Science) software, wherein the spectral parameters which are significantly correlated with the leaf physiological indexes are taken as spectral characteristic factors representing plant ozone stress for extracting the spectral information. Compared with a conventional method, the extraction method for pigment content is easy and convenient to operate, and effectively avoids measurement error brought about by lighting decomposition, pigment residue and plant tissue residue in the operation process. The extraction method for spectral information is wide in application, can be used for early identification of ozone stress of various plants, and can provide a theoretical basis for remote sensing monitoring of plant ozone stress.

Description

The extracting method of a kind of ozone stress pigment in plant leaf and spectral information

Technical field

The present invention relates to a kind of pigment and withdrawing spectral information method of plant leaf blade, specifically the extracting method of a kind of ozone stress pigment in plant leaf and spectral information.

Background technology

Ozone (O in the atmosphere of troposphere ₃) be main secondary air pollution thing, be to be generated through photochemical reaction by the disposable pollutant such as oxides of nitrogen that enters atmosphere, be the principal ingredient that affects greenhouse gases and the photo-chemical smog of surface layer temperature.In recent years, the artificial origin such as motor vehicle exhaust emission, commercial production causes O near the ground ₃concentration increases day by day, estimates that 2015 to the O in the year two thousand fifty troposphere ₃concentration will increase by 20% – will increase by 40% – 60% for 25%, 2100 year.O in troposphere ₃be increased to a certain degree and not only can damage human health, and crop growth is also produced to profound influence.It can cause crop stomatal closure, the destruction of Photosynthetic structure and function, suppress the transmission of photosynthetical system electronics, change pigment content and composition, reduce ribulose-1,5-bisphosphate, content and the activity of 5-diphosphonic acid Carboxylase/oxygenase, cause photosynthetic capacity to decline, and further reduce crop biomass and output.

Pigment in plant leaf contamination is the key factor that affects blade visible waveband spectral information, therefore, extracting in the process of Spectra of The Leaves information, is necessary leaf pigment concentration carry out extraction and determination and measurement result degree of accuracy is had relatively high expectations.Traditional blade pigment detection method mainly adopts grinding-colourimetry, often in operating process there is following problem: grind (1) has partial pigment decompose because of illumination in filter process, simultaneously also can residual a small amount of pigment on mortar, filter paper, these all can cause measurement result on the low side; (2) grind the plant tissue remaining in leaching liquor after filtration and can make liquid muddiness, affect measurement result.

The O of plant ₃coerce that to carry out EARLY RECOGNITION be the important step that it is prevented and treated, and monitoring method based on plant spectral technology have the plurality of advantages such as the visual field is wide, information wide, in real time dynamic, has the incomparable superiority of conventional method aspect pollution monitoring.Plant is at O ₃in growth and development process under coercing can there is complicated variation in physiological ecological, and these can reflection to some extent on spectrum.Therefore, in conjunction with the actual conditions of different vegetation types, accurate evaluation plant leaf blade spectrum at different-waveband to O ₃the response of coercing, and then select specificity spectrum index, effectively extract characteristic spectrum information, to plant O ₃the EARLY RECOGNITION of coercing and control all have great importance.

Summary of the invention

The object of the invention is to provide a kind of and is widely used, is easy to the ozone stress pigment in plant leaf of popularization and the extracting method of spectral information.

For achieving the above object, the technical solution used in the present invention is:

An extracting method for ozone stress pigment in plant leaf and spectral information passes through O in plant culture process ₃carry out Stress treatment, vegetation season, measure plant spectral information breeding time by different plant, adopt direct extraction (exempting to grind) to measure different plant physical signs breeding time simultaneously; Afterwards the plant spectral information of mensuration is converted into spectrum parameter, by SPSS software, spectrum parameter is carried out to correlation analysis with corresponding Physiological Index in Leaves, wherein be the spectrum parameter of significant correlation with Physiological Index in Leaves, extract spectral information as the spectral signature factor that characterizes plant ozone stress.

Plant culture is carried out opening in top air chamber (OTC), in plant culture process, passes into O to the process chamber of OTC air chamber setting ₃, 7-8 hour ventilates every day.

Described opening in top air chamber (OTC) established respectively control group and O ₃coerce, group; The about 40nmolmol of natural ozone concentration in contrast ^-1, O ₃during concentration raises and processes, ozone concentration can be controlled in 60-110nmolmol ^-1.

In vegetation season, adopt the spectral information of spectroradio spectrophotometer plant leaf at Visible-to-Near InfaRed wave band, different plant each spotting METHOD FOR CONTINUOUS DETERMINATION breeding time 5-10 time breeding time by different plant.

Spectral information is converted into spectral signature parameter, spectral signature parameter is respectively: normalized differential vegetation index NDVI, position, red limit parameters R EP, the relevant pigment indices SIPI of structure, near-infrared band spectral reflectivity change integrated value A, moisture index WI, and computing formula is as follows:

NDVI=(R ₉₀₀-R ₆₈₀)/(R ₉₀₀+R ₆₈₀) (1)

$\mathrm{REP}=\mathrm{\λ}\max \frac{\mathrm{dR}}{\mathrm{d\λ}}---\left(2\right)$

SIPI=(R ₄₃₀-R ₈₀₀)/(R ₆₈₀-R ₈₀₀) (3)

$A={\∫}_{{\mathrm{\λ}}_2}^{{\mathrm{\λ}}_1}{R}_{\mathrm{\λ}}{d}_{\mathrm{\λ}}---\left(4\right)$

WI=(R ₉₇₀/R ₉₀₀) (5)

In formula: R _λfor plant is at the Leaf reflectance of af at wavelength lambda; λ ₁with λ ₂be respectively the two ends wavelength node of selected near infrared spectrum wave band, site position can specifically be determined according to vegetation type and spectrometer resolution;

for the spectral reflectivity derivative value of af at wavelength lambda.

Described employing direct extraction (exempting to grind) is measured different plant physical signs breeding time, first take the fresh sample 0.2-0.5g of plant leaf blade, be cut into width and be less than little of 1cm, put into 10ml tool plug test tube, add 10ml80% acetone soln, cover test tube plug, with tinfoil by tight test tube parcel in order to avoid printing opacity, test tube is placed after 24 hours in dark place, directly carries out colorimetric, measures chlorophyll a, chlorophyll b, chlorophyll a+b and carotenoid content.

The invention has the advantages that:

1. compared with classic method, it is easy and simple to handle that direct extraction is measured pigment content, without grinding, filtering, effectively avoided in operating process because illumination decomposition, pigment are residual, remaining the brought error at measurment of plant tissue.

2. withdrawing spectral information method is widely used, and can be used for the EARLY RECOGNITION of various plants ozone stress, can provide fundamental basis for the remote sensing monitoring of plant ozone stress.

Embodiment

1. condition control and plant culture: plant culture is carried out opening in top air chamber (OTC), the bottom surface of OTC air chamber can be regular polygon or circle, air chamber base area and height can be according to concrete vegetation type designs, air chamber top is intilted inclined-plane, angle inclination angle is 40-50 °, is filled with gas and blows out from top to reduce.OTC air chamber is whole mosaic glass around, and the moment keeps clean to reach higher transmittance.Seal seam everywhere with environment-friendly type glass cement, keep the good impermeability in air chamber bottom, prevent that gas scatters and disappears.

OTC air chamber condition can be established 2-4 processing, is respectively contrast and O ₃concentration raises, the about 40nmolmol of natural ozone concentration in contrast ^-1, O ₃during concentration raises and processes, ozone concentration can be controlled in 60-110nmolmol ^-1, the 7-8 hour that ventilates every day, each processing repeats for 3 times.Selecting to intend research plant by demand cultivates.

2. spectrum data gathering: in vegetation season, measure plant spectral information breeding time by different plant, each OTC air chamber is selected the above plant of 3 strain, application spectroradio spectrophotometer plant leaf spectral reflectivity, gather tested plant Visible-to-Near InfaRed wave band spectral information, each spotting METHOD FOR CONTINUOUS DETERMINATION 5-10 time.

3. spectrum parameter calculates: spectral information is converted into spectral signature parameter, and spectral signature parameter is respectively: normalized differential vegetation index NDVI, position, red limit parameters R EP, the relevant pigment indices SIPI of structure, near-infrared band spectral reflectivity change integrated value A, moisture index WI.Computing formula is as follows:

NDVI=R( ₉₀₀-R ₆₈₀)/(R ₉₀₀+R ₆₈₀) (1)

$\mathrm{REP}=\mathrm{\λ}\max \frac{\mathrm{dR}}{\mathrm{d\λ}}---\left(2\right)$

SIPI=(R ₄₃₀-R ₈₀₀)/(R ₆₈₀-R ₈₀₀) (3)

$A={\∫}_{{\mathrm{\λ}}_2}^{{\mathrm{\λ}}_1}{R}_{\mathrm{\λ}}{d}_{\mathrm{\λ}}---\left(4\right)$

WI=(R ₉₇₀/R ₉₀₀) (5)

In formula: R _λfor plant is at the Leaf reflectance of af at wavelength lambda; λ ₁with λ ₂be respectively the two ends wavelength node of selected near infrared spectrum wave band, site position can specifically be determined according to vegetation type and spectrometer resolution;

for the spectral reflectivity derivative value of af at wavelength lambda.

4. leaf pigment concentration is measured: extracting after Spectra of The Leaves information, herborization sample, adopts direct extraction analysis to measure leaf pigment concentration.First take the fresh sample 0.2-0.5g of plant leaf blade, be cut into width and be less than little of 1cm, put into 10ml tool plug test tube, add 10ml80% acetone soln, cover test tube plug, with tinfoil by test tube parcel tight in order to avoid printing opacity, test tube is placed after 24 hours in dark place, leaching liquor directly carries out colorimetric without filtering, and measures chlorophyll a, chlorophyll b, chlorophyll a+b and carotenoid content.

5. vane thickness and moisture determination: measure vane thickness with vane thickness instrument, oven drying method is measured leaf water content.

6. withdrawing spectral information: adopt statistical software to carry out correlation analysis to spectrum parameter with corresponding Physiological Index in Leaves and (be respectively NDVI, REP and chlorophyll a+b, SIPI and carotenoid/chlorophyll a, A and vane thickness, WI and leaf water content), calculate spectrum parameter and corresponding Physiological Index in Leaves related coefficient, and gained related coefficient is carried out to significance test, choose the spectrum parameter that is wherein significant correlation with Physiological Index in Leaves, extract spectral information as the spectral signature factor that characterizes plant ozone stress.

Case study on implementation

1. condition control and plant culture: test is carried out at Shenyang Inst. of Applied Ecology, Chinese Academy of Sciences's ecological experiment station, and take wheat as research object, OTC air chamber condition is established 2 processing, is respectively contrast and O ₃concentration raises, the about 40nmolmol of natural ozone concentration in OTC contrast ^-1, O ₃during concentration raises and processes, ozone concentration is 60nmolmol ^-1, to ventilate every day 7 hours, each processing repeats for 3 times.

2. spectrum data gathering: in wheat growth season, respectively at wheat tillering phase and jointing stage collection Spectra of The Leaves information, each OTC air chamber is selected the above plant of 3 strain, gather wheat leaf blade 400-1000nm wave band spectral information, each spotting METHOD FOR CONTINUOUS DETERMINATION 5 times.

3. spectrum parameter calculates: spectral information is converted into spectral signature parameter, and the spectral signature parameter of calculating is respectively: normalized differential vegetation index NDVI, position, red limit parameters R EP, the relevant pigment indices SIPI of structure, near-infrared band spectral reflectivity change integrated value A, moisture index WI.

4. leaf pigment concentration is measured: take the fresh sample 0.5g of wheat leaf blade, put into 10ml tool plug test tube, add 10ml80% acetone soln, cover test tube plug, with tinfoil by tight test tube parcel in order to avoid printing opacity, test tube is placed after 24 hours in dark place, and leaching liquor directly carries out colorimetric without filtering, and measures chlorophyll a, chlorophyll b, chlorophyll a+b and carotenoid content.

5. vane thickness and moisture determination: measure vane thickness with vane thickness instrument, oven drying method is measured leaf water content.

6. withdrawing spectral information: adopt SPSS analysis software to carry out correlation analysis to spectrum parameter with corresponding Physiological Index in Leaves and (be respectively NDVI, REP and chlorophyll a+b, SIPI and carotenoid/chlorophyll a, A and vane thickness, WI and leaf water content), calculate spectrum parameter and corresponding Physiological Index in Leaves related coefficient, and gained related coefficient is carried out to significance test, result shows, in the spectrum parameter of choosing, be the NDVI that has, REP and the A (table 1) of significant correlation with Physiological Index in Leaves.Can be used as the spectral signature factor extraction spectral information that characterizes this confession examination wheat ozone stress.

Table 1 wheat spectrum parameter and Physiological Index in Leaves related coefficient

Claims (6)

1. an extracting method for ozone stress pigment in plant leaf and spectral information, is characterized in that: in plant culture process, pass through O ₃carry out Stress treatment, vegetation season, measure plant spectral information breeding time by different plant, adopt direct extraction (exempting to grind) to measure different plant physical signs breeding time simultaneously; Afterwards the plant spectral information of mensuration is converted into spectrum parameter, by SPSS software, spectrum parameter is carried out to correlation analysis with corresponding Physiological Index in Leaves, wherein be the spectrum parameter of significant correlation with Physiological Index in Leaves, extract spectral information as the spectral signature factor that characterizes plant ozone stress.

2. by the extracting method of ozone stress pigment in plant leaf claimed in claim 1 and spectral information, it is characterized in that: plant culture is carried out opening in top air chamber (OTC), in plant culture process, passes into O to the process chamber of OTC air chamber setting ₃, 7-8 hour ventilates every day.

3. by the extracting method of ozone stress pigment in plant leaf claimed in claim 2 and spectral information, it is characterized in that: described in open in top air chamber (OTC) and establish respectively control group and O ₃coerce, group; The about 40nmolmol of natural ozone concentration in contrast ^-1, O ₃during concentration raises and processes, ozone concentration can be controlled in 60-110nmolmol ^-1.

4. by the extracting method of ozone stress pigment in plant leaf claimed in claim 1 and spectral information, it is characterized in that: in vegetation season, adopt the spectral information of spectroradio spectrophotometer plant leaf at Visible-to-Near InfaRed wave band, different plant each spotting METHOD FOR CONTINUOUS DETERMINATION breeding time 5-10 time breeding time by different plant.

5. by the extracting method of ozone stress pigment in plant leaf claimed in claim 1 and spectral information, it is characterized in that: spectral information is converted into spectral signature parameter, spectral signature parameter is respectively: normalized differential vegetation index NDVI, position, red limit parameters R EP, the relevant pigment indices SIPI of structure, near-infrared band spectral reflectivity change integrated value A, moisture index WI, and computing formula is as follows:

NDVI=(R ₉₀₀-R ₆₈₀)/(R ₉₀₀+R ₆₈₀) (1)

$\mathrm{REP}=\mathrm{\λ}\max \frac{\mathrm{dR}}{\mathrm{d\λ}}---\left(2\right)$

SIPI=(R ₄₃₀-R ₈₀₀)/(R ₆₈₀-R ₈₀₀) (3)

$A={\∫}_{{\mathrm{\λ}}_2}^{{\mathrm{\λ}}_1}{R}_{\mathrm{\λ}}{d}_{\mathrm{\λ}}---\left(4\right)$

WI=(R ₉₇₀/R ₉₀₀) (5)

In formula: R _λfor plant is at the Leaf reflectance of af at wavelength lambda; λ ₁with λ ₂be respectively the two ends wavelength node of selected near infrared spectrum wave band, site position can specifically be determined according to vegetation type and spectrometer resolution;

for the spectral reflectivity derivative value of af at wavelength lambda.

6. by the extracting method of ozone stress pigment in plant leaf claimed in claim 1 and spectral information, it is characterized in that: described employing direct extraction (exempting to grind) is measured different plant physical signs breeding time, first take the fresh sample 0.2-0.5g of plant leaf blade, be cut into width and be less than little of 1cm, put into 10ml tool plug test tube, add 10ml80% acetone soln, cover test tube plug, with tinfoil by tight test tube parcel in order to avoid printing opacity, test tube is placed after 24 hours in dark place, directly carry out colorimetric, measure chlorophyll a, chlorophyll b, chlorophyll a+b and carotenoid content.