CN108872108A - Plant young leaflet tablet pigment detection method - Google Patents
Plant young leaflet tablet pigment detection method Download PDFInfo
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- CN108872108A CN108872108A CN201810709795.1A CN201810709795A CN108872108A CN 108872108 A CN108872108 A CN 108872108A CN 201810709795 A CN201810709795 A CN 201810709795A CN 108872108 A CN108872108 A CN 108872108A
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/17—Systems in which incident light is modified in accordance with the properties of the material investigated
- G01N21/25—Colour; Spectral properties, i.e. comparison of effect of material on the light at two or more different wavelengths or wavelength bands
- G01N21/31—Investigating relative effect of material at wavelengths characteristic of specific elements or molecules, e.g. atomic absorption spectrometry
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- G—PHYSICS
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- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/286—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q involving mechanical work, e.g. chopping, disintegrating, compacting, homogenising
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/286—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q involving mechanical work, e.g. chopping, disintegrating, compacting, homogenising
- G01N2001/2866—Grinding or homogeneising
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Abstract
The present invention provides a kind of measuring method of plant young leaflet tablet pigment comprising test sample preparation and colorimetric determination.Test sample preparation step includes：Appropriate plant young leaflet tablet is weighed into the centrifuge tube equipped with steel ball, pouring into liquid nitrogen makes plant leaf blade rapid condensation, in being crushed rapidly in vortex instrument, centrifuge tube is wrapped with masking foil, solvent is added by certain mass volume ratio, it extracts under certain temperature and illumination condition and is centrifuged after a certain period of time, supernatant is test sample.Colorimetric determination step includes：Extraction solvent is used to carry out baseline correction as blank ultraviolet specrophotometer, the absorbance value under measurement 663nm, 646nm and 470nm calculates chlorophyll a, chlorophyll b and carotenoid content using Wellburn formula (1984).Operation of the present invention is easy, and high-efficient, the time is short, and measurement result is accurate, stablizes, and can be used in scientific research and industrial production to the rapidly extracting of plant young leaflet tablet pigment and accurate measurement, promotion and application value with higher.
Description
Technical field
The invention belongs to plant physiological techniques fields, are related to a kind of side for efficiently quickly measuring plant young leaflet tablet pigment
Method is suitable for measuring plant young leaflet tablet leaf in particular to using the method for ultraviolet specrophotometer measurement pigment in plant leaf
Green element a, chlorophyll b and carotenoid content.
Background technique
The composition of Leaves of The Higher Plants pigment mainly includes Chlorophylls and Carotenoids, and chlorophyll mainly includes chlorophyll a
And chlorophyll b, carotenoid are broadly divided into carrotene and lutein two major classes.Chlorophyll is that plant progress is photosynthetic heavy
Want ingredient, and the important indicator of reflection plant physiology situation.Carotenoid serves not only as the ginseng of the accessory pigments in photosynthesis
With the absorption and transmitting of luminous energy, also has the function of anti-oxidant, anticancer and anti-radiation.Therefore, Chlorophylls and Carotenoids
It extracts and measures and is extremely important in production and research work.
The primary organic solvent for extracting pigment in plant tissue includes chloroform, petroleum ether, acetone, ethyl alcohol and dimethyl sulfoxide
Deng carrying out mixing or its extraction with aqueous solution phytochrome by a certain percentage usually using acetone and ethyl alcohol.Currently, with acetone extraction
Method is most widely used, most proposes (Mackinney G.Absorption of light by by Mackinney earlier than nineteen forty-one
chlorophyll solutions[J].Journal of Biological Chemistry,1941,140(2):315-
322.), (Arnon D I.Copper enzymes in isolated is explained and derived through Arnon within 1949
chloroplasts.Polyphenoloxidase in Beta vulgaris[J].Plant Physiology,1949,24
(1):1-15.), earliest chlorophyll saponification measuring method has been reformed in the generation of the method, and operation is relatively easy, and time-consuming substantially shortens.
Acetone extraction mainly uses 80% acetone to carry out milling and extracting, then through washing turn, filtering and the step such as constant volume
Suddenly, process is complex, and pigment detection value is relatively low.This method is improved and proposes chlorophyll by Soviet Union's exhibition et al. simplifies extraction
Method, i.e., directly using the extraction of 80% acetone, (Soviet Union's exhibition, Zhang Xiaosu, Chen Juan wait chlorophyll content after shredding plant green tissues
Simplification [J] Plant Physiology Communications of measurement, 2010,46 (4):399-402.).This method enormously simplifies operating process,
The dosage of vegetable material and reagent is saved, but extraction time is too long, generally requiring could extract completely for 24-72 hours.Qiu Nian
It is big et al. to have invented two step rapid fractionation method of DMSO-80% acetone, chlorophyll first is extracted with DMSO high temperature, then dilute with 80% acetone
Extraction is released, evaluated error can be effectively reduced, accelerates finding speed, 3 hours (Qiu Nianwei, Wang Xiu will be shortened to extraction time
Suitable, Yang Fabin waits the rapidly extracting and micrometric measurement [J] Botany Gazette of chlorophyll, 2016,51 (5):667-678.).Wu Zhi
Rising sun et al. uses hot acetone extraction chlorophyll instead, and extraction time is made to shorten to 1.5-2.5 hours (Wu Zhixu, Zhang Yayan phytoplanktons
Internal chlorophyll a Improvement in determination method [J] chemical analysis metering, 2002,11 (6):37-38.).Nonetheless, with top
Method is still relatively complicated, and extraction time is longer or there are security risks, is objectively difficult to and is protected from light operation, thus pigment extracts effect
Rate is not high.
Summary of the invention
The technical problem to be solved in the present invention：Existing Chlorophylls and Carotenoids extraction process is complicated, time-consuming and effect
The problems such as rate is low.In order to solve the above technical problems, present patent application inventor is known according to literature survey result, there is no both at home and abroad
Utilize the report of liquid nitrogen auxiliary measuring pigment in plant leaf.
In order to simplify the extraction step of pigment, the extraction time for shortening pigment and the extraction efficiency etc. for improving pigment are asked
Topic, the present invention provide a kind of plant young leaflet tablet pigment detection method.The present invention is by exploring optimal pigment extracting mode, sieve
Optimum extraction time, temperature and mass volume ratio are selected, to realize the rapidly extracting and accurate measurement of plant young leaflet tablet pigment.
The purpose of the present invention is be achieved through the following technical solutions：
Plant young leaflet tablet pigment detection method, including test sample preparation step and colorimetric determination step, described in
Test sample preparation step includes：Using the cooling plant leaf blade of liquid nitrogen, with the pigment in solvent extraction blade, extracting solution after crushing
Centrifugation, takes supernatant, and test sample is made.
One of preferred embodiment as technical solution of the present invention, the test sample preparation step are specifically wrapped
It includes：By plant leaf blade merging equipped in the centrifuge tube of steel ball, pouring into liquid nitrogen condenses plant leaf blade, in crushing in vortex instrument, adds
Enter solvent, extract under certain temperature and illumination condition and be centrifuged after a certain period of time, supernatant is test sample.
One of preferred embodiment as technical solution of the present invention, the solvent select acetone or ethyl alcohol or third
The mixed solution of the mixed solution or acetone of ketone and ethyl alcohol, second alcohol and water.
It is further preferred that the solvent selects the mixed solution of acetone and ethyl alcohol, with volume basis, acetone:Ethyl alcohol=
1:1 or acetone:Ethyl alcohol=2:1 or acetone:Ethyl alcohol=1:2；The solvent selects the mixed solution of acetone, second alcohol and water, with
Volume basis, acetone:Ethyl alcohol:Water=6:3:1 or acetone:Ethyl alcohol:Water=3:6:1 or acetone:Ethyl alcohol:Water=4.5:4.5:1.
Result verification through the embodiment of the present invention, the optimum extraction solvent of pigment in plant leaf are 95% ethyl alcohol (volume ratio).
One of preferred embodiment as technical solution of the present invention, the extraction temperature are -20~37 DEG C.Into one
Preferably, the optimum extraction temperature of pigment in plant leaf is 4 DEG C to step.
One of preferred embodiment as technical solution of the present invention, the extraction intensity of illumination be 0~
1000lux.It is further preferred that the optimum extraction illumination condition of pigment in plant leaf is 0lux (dark condition).It is specific real
The mode of applying is that the effective masking foil of centrifugation wraps, and is extracted under dark condition.
One of preferred embodiment as technical solution of the present invention, the extraction time are 1~60min.Into one
Preferably, the optimum extraction time of pigment in plant leaf is 5min to step.
One of preferred embodiment as technical solution of the present invention, in terms of g/mL, plant leaf blade to be measured and extraction
The mass volume ratio range of solvent is 1/400~1/25.It is further preferred that the optimum extraction quality volume of pigment in plant leaf
Than for 1/100 (blade and solvent, g/mL).
For plant young leaflet tablet pigment detection method of the present invention, the colorimetric determination step includes：It surveys
Determine absorbance value of the test sample at 663nm, 646nm and 470nm, calculates chlorophyll a, Ye Lv using Wellburn improved method
The content of plain b and carotenoid.
Plant young leaflet tablet pigment detection method of the present invention, is particularly suitable for the efficient of plant young leaflet tablet pigment
Quickly measurement.
Optimization based on above-mentioned parameter, the present invention provide a kind of plant young leaflet tablet pigment detection method of optimization, packet
Include test sample preparation step and colorimetric determination step：
(1) prepared by test sample
Appropriate plant young leaflet tablet is weighed into the centrifuge tube equipped with steel ball, pouring into liquid nitrogen makes plant leaf blade rapid condensation,
In being crushed rapidly in vortex instrument, wrap centrifuge tube with masking foil, optimum solvent be added, suitable temperature, illumination condition and
It is extracted certain time under suitable quality volume, extracting solution centrifugation, supernatant is test sample.
(2) colorimetric determination
Test sample made from step (1) is detected using colorimetric method, measures it in 663nm, 646nm and 470nm
The absorbance value at place uses Wellburn improved method (Wellburn A R, Lichtenthaler H K.Formulae and
program to determine total carotenoids and chlorophylls a and b of leaf
extracts in different solvents[M].1984:Springer Netherlands.9-12.) calculate pigment contain
Amount.
The formula that Wellburn improved method calculates pigment content is as follows：
Ca(mg/g FW)=(12.21A663–2.81A646)×V/(1000×W)；
Cb(mg/g FW)=(20.13A646–5.03A663)×V/(1000×W)；
CAlways(mg/g FW)=(17.32A646+7.18A663)×V/(1000×W)；
Cx·c(mg/g FW)=[(1000A470–3.27Ca–104Cb)/229]×V/(1000×W)。
In formula：
Ca--- Chlorophyll-a Content；
Cb--- content of chlorophyll b；
CAlways--- total chlorophyll content；
Cx·c--- carotenoid content；
A663--- absorbance value of the sample at 663nm；
A646--- absorbance value of the sample at 646nm；
A470--- absorbance value of the sample at 470nm；
V --- sample volume, unit are milliliter (mL)；
W --- fresh weight, unit are gram (g)；
The content unit of mg/g FW --- leaf pigment, mg/g fresh weight.
The present invention combines the advantage and disadvantage of the above-mentioned prior art, provides a kind of efficiently quick measurement plant young leaflet tablet color
The method of element, abbreviation liquid nitrogen method cool down rapidly plant leaf blade using liquid nitrogen, using steel ball in powder quick in vortex instrument
It is broken, it reuses solvent and carries out being protected from light extraction.The method can accomplish to be protected from light completely for the pigment extraction of tender leaf tissue, lead to
Screening suitable Extraction solvent, temperature, illumination condition, time and best in quality volume ratio are crossed, pigment dissolution can be effectively improved
Efficiency will foreshorten within 5min extraction time.Research is extracted compared to previous pigment, the method extraction process is simple, the time
It is short, it is high-efficient, measurement result stablize, can be used in research work and industrial production in plant young leaflet tablet material pigment it is fast
Speed is extracted and accurate measurement.
Specifically, the pigment of the present invention using in liquid nitrogen method measurement plant young leaflet tablet, at least has with following
Beneficial effect or advantage.
1) present invention carries out rapid condensation to plant leaf blade using liquid nitrogen, reduces leaf temperature, can effectively reduce pigment and exist
Degradation speed in crushing process keeps measurement result more accurate.
2) present invention, can be by blade powder in fast-crushing in vortex instrument in such a way that steel ball replaces traditional manual grinding
Broken is more abundant, and the time of blade crushing is greatly decreased, and accelerates the measurement of pigment.
3) present invention is by screening optimal Extraction solvent, temperature, illumination condition, time and the quality of blade and solvent
Volume ratio, the links during optimization measurement pigment effectively shorten extraction time of pigment, increase the dissolution effect of pigment
Rate.
4) present invention calculates chlorophyll a, chlorophyll b and class recklessly using the formula (1984) of Wellburn improvement Arnon
The content of radish element, improves the accuracy of pigment detection.
Detailed description of the invention
Fig. 1 is influence of the different solvents to Lettuce young leaflet tablet pigment extraction efficiency.
Fig. 2 is influence of the different solvents to grape young leaflet tablet pigment extraction efficiency.
Fig. 3 is influence of the different solvents to pokeroot young leaflet tablet pigment extraction efficiency.
Fig. 4 is influence of the different extraction times to Lettuce young leaflet tablet pigment extraction efficiency.
Fig. 5 is influence of the different extraction times to grape young leaflet tablet pigment extraction efficiency.
Fig. 6 is influence of the different extraction times to pokeroot young leaflet tablet pigment extraction efficiency.
Fig. 7 is influence of the mass volume ratio of Lettuce young leaflet tablet and solvent to pigment extraction efficiency.
Fig. 8 is influence of the mass volume ratio of grape young leaflet tablet and solvent to pigment extraction efficiency.
Fig. 9 is influence of the mass volume ratio of pokeroot young leaflet tablet and solvent to pigment extraction efficiency.
Specific embodiment
Below with reference to embodiment, the present invention is further illustrated, and following each embodiments are merely to illustrate the present invention, right
The present invention is not restricted.The those of ordinary skill of this research field can refer to the description of the present embodiment, or with the present embodiment
The method is that the present invention is realized in a manner of preferred according to the replacement or appropriate changes and combinations for carrying out technological means.
Embodiment 1
A kind of plant young leaflet tablet pigment detection method of the present embodiment, in the present invention, this measuring method is also referred to as liquid
Nitrogen method.The present embodiment using liquid nitrogen method measurement plant young leaflet tablet in pigment, the comparison including different pigment extracting modes with
And the optimal Extraction solvent of screening, temperature, the best in quality volume ratio of illumination condition, extraction time and blade and solvent.
One, materials and methods
(1) different extracting modes compare
Liquid nitrogen method is respectively adopted, 3 kinds of different modes of polishing and extraction extract the pigment in blade, to not
It is compared with pigment extracting mode.
Liquid nitrogen method：Appropriate blade is weighed into the centrifuge tube equipped with steel ball, a little liquid nitrogen is added, vortex instrument is sufficiently smashed,
Centrifuge tube is wrapped with masking foil, optimum extraction solvent is added and is protected from light extraction.
Polishing：The blade of certain mass is weighed into mortar, a little quartz sand is added and is fully ground, solvent is added and carries out
It washes and turns, is protected from light extraction.
Extraction：Blade is sufficiently shredded, the blade of certain mass is weighed into centrifuge tube, the solvent of certain volume is added
Extraction is for 24 hours.
(2) different solvents
The pigment of plant leaf blade is extracted using 8 kinds of different organic extraction solvents.This 8 kinds of solvents are respectively：
(1) 80% acetone；
(2) 95% ethyl alcohol；
(3) acetone：Ethyl alcohol=1:1；
(4) acetone：Ethyl alcohol=2:1；
(5) acetone：Ethyl alcohol=1:2；
(6) acetone：Ethyl alcohol：Water=6:3:1；
(7) acetone：Ethyl alcohol：Water=3:6:1；
(8) acetone：Ethyl alcohol：Water=4.5:4.5:1.
Blade is crushed using liquid nitrogen method, above 8 kinds of solvents is separately added into and wash and turn, extract, measurement pigment contains after 1d
Amount.
(3) varying environment factor
Study the influence that different temperature, illumination condition and extraction time extract different type pigment in plant leaf.
- 20 DEG C, 4 DEG C, 25 DEG C and 37 DEG C four gradients are respectively set in temperature, and 0lux, 100lux, 1000lux tri- is arranged in illumination condition
Seven gradients of 1min, 3min, 5min, 10min, 20min, 30min, 60min are arranged in a gradient, extraction time.With three kinds of plants
Blade (Lettuce young leaflet tablet, grape young leaflet tablet, pokeroot young leaflet tablet) is material, using optimal Extraction solvent and
The extraction of liquid nitrogen method progress pigment.
(4) different quality volume ratio
For determine pigment in plant leaf extraction process in leaf quality and Extraction solvent optimal proportion, by leaf quality
(g) it is respectively 1/25,1/50,1/100,1/200,1/400 to be configured with the ratio of the volume of Extraction solvent (mL), uses liquid
Nitrogen method is in optimum extraction temperature, optimal illumination condition and the best in quality volume for exploring pigment extraction under most suitable extraction time
Than.
(5) measurement of Chlorophylls and Carotenoids
Zhao is first bright et al., and (Zhao Xianming, Wang Yanxia, Du Xiao wait tealeaves chlorophyll mixed liquor restricted-access media condition optimizing
The agriculture journal in the southwest [J], 2,011 24 (4):001303-1308.) (it is green that Yang Min text quickly measures plant leaf blade leaf with Yang Minwen
The spectrographic laboratory discussion [J] of cellulose content method, 2002,19 (4):Result of study 478-481.) shows that chlorophyll exists
Optical absorption characteristics in 8 kinds of different solvents such as 80% acetone are similar, therefore Arnon formula can be used to calculate chlorophyll and class Hu trailing plants
The content of Bu Su.But some researches show that Arnon method has large error in terms of absorbing wavelength and calculation formula, in order to improve effect
Rate and research are convenient, this research is using Wellburn et al. in the calculation formula of modified Chlorophylls and Carotenoids in 1984：
Ca(mg/g FW)=(12.21A663–2.81A646)×V/(1000×W)；
Cb(mg/g FW)=(20.13A646–5.03A663)×V/(1000×W)；
CAlways(mg/g FW)=(17.32A646+7.18A663)×V/(1000×W)；
Cx·c(mg/g FW)=[(1000A470–3.27Ca–104Cb)/229]×V/(1000×W)。
In formula：
Ca--- Chlorophyll-a Content；
Cb--- content of chlorophyll b；
CAlways--- total chlorophyll content；
Cx·c--- carotenoid content；
A663--- absorbance value of the sample at 663nm；
A646--- absorbance value of the sample at 646nm；
A470--- absorbance value of the sample at 470nm；
V --- sample volume, unit are milliliter (mL)；
W --- fresh weight, unit are gram (g)；
The content unit of mg/g FW --- leaf pigment, mg/g fresh weight.
Two, results and discussion
(1) optimum extraction mode
Influence of the different extracting modes to leaf pigment extraction efficiency is shown in Table 1.The display of table 1, the extraction of three kinds of leaf pigments
Efficiency is all remarkably higher than traditional polishing and leaching with liquid nitrogen method highest, chlorophyll a, chlorophyll b and the carotenoid of measurement
Formulation.
Influence of the different extracting modes of table 1 to pigment extraction efficiency
(2) optimum extraction solvent
Fig. 1 is influence of the different solvents to Lettuce young leaflet tablet pigment extraction efficiency.Fig. 2 is different solvents to grape children
The influence of leaflet tablet pigment extraction efficiency.Fig. 3 is influence of the different solvents to pokeroot young leaflet tablet pigment extraction efficiency.It is comprehensive
Closing Fig. 1 to Fig. 3 can obtain, (1) 80% acetone of Extraction solvent；(2) 95% ethyl alcohol；(3) acetone：Ethyl alcohol=1:1；(4) acetone：Ethyl alcohol
=2:1；(5) acetone：Ethyl alcohol=1:2；(6) acetone：Ethyl alcohol：Water=6:3:1；(7) acetone：Ethyl alcohol：Water=3:6:1；(8) third
Ketone：Ethyl alcohol：Water=4.5:4.5:1 is able to achieve effective extraction of the pigment in the plant young leaflet tablet of the present embodiment.With regard to this reality
For applying example, the optimum extraction solvent of these three blades is 95% ethyl alcohol.
(3) optimum extraction temperature
Influence of the different Extracting temperatures to leaf pigment extraction efficiency is shown in Table 2.Table 2 shows that extraction temperature is -20~37 DEG C
It is able to achieve effective extraction of the pigment in the plant young leaflet tablet of the present embodiment.For the present embodiment, these three blades
Optimum extraction temperature is 4 DEG C.
Influence of 2 different temperatures of table to pigment extraction efficiency
(4) optimum illumination condition
Influence of the different illumination conditions to leaf pigment extraction efficiency is shown in Table 3.From table 3 it can be seen that extraction intensity of illumination
Effective extraction of the pigment in the plant young leaflet tablet of the present embodiment is able to achieve for 0~1000lux.For the present embodiment,
The illumination condition that these three leaf pigments extract is best to carry out in dark (0lux).
Influence of 3 intensity of illumination of table to pigment extraction efficiency
(5) the optimum extraction time
Fig. 4 is influence of the different extraction times to Lettuce young leaflet tablet pigment extraction efficiency.Fig. 5 is different extraction times
Influence to grape young leaflet tablet pigment extraction efficiency.Fig. 6 is that different extraction times extract pokeroot young leaflet tablet pigment
The influence of efficiency.Comprehensive fig. 4 to fig. 6 can obtain, and extraction time is the plant young leaflet tablet that 1~60min is able to achieve the present embodiment
In pigment effective extraction.For the present embodiment, the optimum extraction time that these three leaf pigments extract is 5min.
(6) best in quality volume ratio
Fig. 7 is influence of the mass volume ratio of Lettuce young leaflet tablet and solvent to pigment extraction efficiency.Fig. 8 is grape children
Influence of the mass volume ratio of leaflet tablet and solvent to pigment extraction efficiency.Fig. 9 is the matter of pokeroot young leaflet tablet and solvent
Measure influence of the volume ratio to pigment extraction efficiency.Complex chart 7 can be obtained to Fig. 9, in terms of g/mL, plant leaf blade and Extraction solvent to be measured
Mass volume ratio range be 1/400~1/25 pigment being able to achieve in the plant young leaflet tablet of the present embodiment effectively mentioning
It takes.For the present embodiment, the best in quality volume ratio that these three leaf pigments extract is 1/100.
Three, conclusion
The optimum extraction condition of three kinds of plant young leaflet tablet pigments is shown in Table 4.As shown in table 4, three kinds of young leaflet tablets are with liquid nitrogen
Method pigment extraction efficiency highest, the optimum extraction solvent of pigment are acetone：Ethyl alcohol：Water=4.5:4.5:1 mixed solution, most preferably
Extracting temperature is 4 DEG C, and it is to carry out in the dark best that pigment, which extracts illumination condition, and the most suitable extraction time of leaf pigment is
Pigment dissolution rate highest when the mass volume ratio of 5min, blade and solvent is 1/100.
The optimum extraction condition of 4 three kinds of plant young leaflet tablet pigments of table
Embodiment 2
Liquid nitrogen method in embodiment 1 is compared by the present embodiment with traditional polishing and extraction, takes tender crow
Liana certain kind of berries blade, the experimental material as the present embodiment.Respectively according to three kinds of liquid nitrogen method, polishing and extraction modes to black liana
Pigment in certain kind of berries blade is extracted and is measured, and compares the difference between distinct methods measurement plant young leaflet tablet pigment content.
Three kinds of different measuring methods are shown in Table 5 to the extraction efficiency of Japanese Cayratia Herb young leaflet tablet pigment.As can be seen from Table 5, three
The efficiency highest of pigment is extracted in kind measuring method with liquid nitrogen method, chlorophyll a, chlorophyll b and the carotenoid of measurement are aobvious
It writes and is higher than traditional polishing and extraction, the time is most short, and the Extraction solvent used is nontoxic, therefore can be used as plant young tender leaf
The best approach of piece pigment detection.
5 three kinds of different measuring methods of table compare the extraction efficiency of Japanese Cayratia Herb young leaflet tablet pigment
Further narration is done to the present invention above in conjunction with embodiment, but present invention is not limited to the embodiments described above,
Within the knowledge of one of ordinary skill in the art, it can also make without departing from the purpose of the present invention
Various change.
Claims (10)
1. plant young leaflet tablet pigment detection method, including test sample preparation step and colorimetric determination step, feature exist
In the test sample preparation step includes：Using the cooling plant leaf blade of liquid nitrogen, with the color in solvent extraction blade after crushing
Element, extracting solution centrifugation, takes supernatant, and test sample is made.
2. plant young leaflet tablet pigment detection method according to claim 1, which is characterized in that the test sample preparation
Step specifically includes：By plant leaf blade merging equipped in the centrifuge tube of steel ball, pouring into liquid nitrogen condenses plant leaf blade, in vortex instrument
Upper crushing is added solvent, extracts under certain temperature and illumination condition and be centrifuged after a certain period of time, supernatant is test sample.
3. plant young leaflet tablet pigment detection method according to claim 1 or 2, which is characterized in that the solvent is selected
The mixed solution of the mixed solution or acetone of acetone or ethyl alcohol or acetone and ethyl alcohol, second alcohol and water.
4. plant young leaflet tablet pigment detection method according to claim 2, which is characterized in that the extraction temperature be-
20~37 DEG C.
5. plant young leaflet tablet pigment detection method according to claim 2, which is characterized in that the extraction intensity of illumination
For 0~1000lux.
6. plant young leaflet tablet pigment detection method according to claim 2, which is characterized in that the effective tinfoil paper of centrifugation
Paper wraps, and is extracted under dark condition.
7. plant young leaflet tablet pigment detection method according to claim 2, which is characterized in that the extraction time is 1
~60min.
8. plant young leaflet tablet pigment detection method according to claim 1 or 2, which is characterized in that be measured in terms of g/mL
The mass volume ratio range of plant leaf blade and Extraction solvent is 1/400~1/25.
9. plant young leaflet tablet pigment detection method according to claim 1, which is characterized in that the colorimetric determination step
Suddenly include：Absorbance value of the test sample at 663nm, 646nm and 470nm is measured, calculates leaf using Wellburn improved method
The content of green element a, chlorophyll b and carotenoid.
10. plant young leaflet tablet pigment detection method according to claim 1, which is characterized in that the plant leaf blade is
Plant young leaflet tablet.
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| CN117664955A (en) * | 2024-02-01 | 2024-03-08 | 中国林业科学研究院热带林业研究所 | Method for rapidly identifying aquilaria sinensis and common aquilaria sinensis based on seedling stage She Biaoxing |
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