CN100399022C - Method for detecting low-molecular-weight organic acid in secretion of root system of plant - Google Patents
Method for detecting low-molecular-weight organic acid in secretion of root system of plant Download PDFInfo
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- CN100399022C CN100399022C CNB2004100114047A CN200410011404A CN100399022C CN 100399022 C CN100399022 C CN 100399022C CN B2004100114047 A CNB2004100114047 A CN B2004100114047A CN 200410011404 A CN200410011404 A CN 200410011404A CN 100399022 C CN100399022 C CN 100399022C
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Abstract
The present invention relates to a method for detecting low-molecular-weight organic acid in the secretion of a plant root system, which belongs to the technical field of agricultural production. The present invention improves components and doses for traditionally detecting low-molecular-weight organic acid; the present invention increases the components for detecting organic acid and improves the doses for detection; in the process of purification, purified substances are separated and concentrated; the purification process and the detection are integrated together; finally, the low-molecular-weight organic acid with multiple components and high precision well meet the needs of scientific research.
Description
Technical field
The invention belongs to purifying, the detection method of plant root exudation low molecular weight organic acid in the agricultural production skill field.
Technical background
Organic acid is the chief component of secretions from plant roots, and its Main Ingredients and Appearance is the potpourri of low molecular weight organic acid.Past, this to be paid attention to inadequately, detection means falls behind, and detected component is few, and precision is low.Now, along with the progress of science and technology, people recognize the importance of low molecular weight organic acid in the secretions from plant roots, and how its decision plant makes full use of nutrient, and the ability that conforms has also determined the growing power of plant.So the researchist of root system of plant nutrition science both domestic and external aspect pays much attention to the research of low molecular weight organic acid in the secretions from plant roots.Low molecular weight organic acid forms focus and breach into the root system of plant research work as plant root exudation organic acid core component.
The method of traditional analysis low molecular weight organic acid generally adopts collects liquid directly measurement or directly concentrated the measurement, interfering ion is many in this method measuring process, make the low organic acid of some content detect less than, it is few to detect the low molecular weight organic acid kind, generally detects the 2-3 kind.Cause the organic acid total amount on the low side, the important mark amount of part organic acid is out in the cold.The present invention is exactly at above shortcoming, improves, and takes secretions from plant roots is carried out resin Solid-Phase Extraction, purifying and combines together with testing process, and detected component is increased, and precision improves, and is the most feasible effective way at present.
Summary of the invention
The present invention is from above technical background, root exudates is carried out the cloudy two kinds of continuous Solid-Phase Extraction of resin column of sun, purifying, the highly purified low molecular weight organic acid potpourri that obtains is handled, obtained the different low molecular weight organic acid of more physical and chemical character.
Key of the present invention is to adopt positive negative resin continuous extraction to separate in the process of solid-liquid continuous extraction, and process and the sample purifying that extracts combined together, finally obtains many components low molecular weight organic acid, thereby better meets the scientific research needs.Specifically the purification assays method is according to the following steps:
A. early-stage preparations: liquid is collected in configuration: 800uM CaCl
2With 5uM H
3BO
4Mixed liquid; Moving phase 0.125M (NH
4)
2HPO
4, use H
3PO
4Accent PH is 2.1-2.5; Standard acid configuration: standard acid sample comprises, succinic acid, citric acid, L type L-lactic acid, malonic acid, the left-handed malic acid of L type, Tartaric acid and oxalic acid, and configuration concentration is 300mg/L;
B. the collection of root exudates: root system of plant is cleaned, put into 200ml and collect liquid, the morning, 10:00-12:00 collected 2 hours, and cryopreservation is to be purified;
C. purifying: the resin cation dry method that takes by weighing the 5g model and the be Amberlite IR-120S 180mm * 20mm resin column of packing into, connect one the resin column that the 2g model is the resin anion (R.A.) same specification of Dowex 1 * 8-200 is housed, use 10ml deionized water pre-leaching resin column respectively, liquid is collected in siphon, cross earlier kation, after through negative ion;
D. wash-out:, collect eluent in 50ml rotary distillation bottle with 20-30ml 1M HCl wash-out resin anion (R.A.) post:
E. concentrate: adopt rotary evaporator, rotate evaporate to dryness under 40 ℃ of conditions, use the 1000ul deionized water dissolving then after, put into refrigerator, 4 ℃ of preservations are to be measured;
F. high pressure liquid chromatographic analysis condition: with long 250mm, the C of diameter 4mm
18Analytical column, detect wavelength 215nm; Moving phase 0.125M (NH
4)
2HPO
4, PH is 2.1-2.5, flow velocity 0.8-1ml/min; Writing speed: 1cm/min; Sample size 20-25ul;
G. detect.
Embodiment
The present invention is applied in Hundred Talents Program (the plain nutrition of the crop root phosphorus) problem of Northeast Institute of Geography and Agricultural Ecology, Chinese Academy of Sc.In the carrying out of problem by the detection of this method to low molecular weight organic acid in the soybean root system secretion, 7 component low molecular weight organic acids have been obtained, adopt the conventional sense means to detect low molecular weight organic acid more than 4 kinds more at present more, improve and detect component more than 130%, precision improves 2-3 doubly, obtained effect preferably, concrete condition such as following table:
Low molecular weight organic acid negative ion testing result unit: umol/L in the soybean root system secretion
Between each as can be seen from the table organic acid that detects significant otherness is arranged, detect many components organic acid.Because this subject study is still underway, is not easy to openly too many relevant concrete data, but only from last table, can embodies the absolute predominance and the practicality of this method.
Claims (1)
1. the detection method of a secretions from plant roots low molecular weight organic acid is characterized in that adopting positive negative resin as Solid-Phase Extraction, and the process and the detection of purifying combined together, finally obtains many components, low molecular weight organic acid, and concrete steps are as follows:
A. early-stage preparations: liquid is collected in configuration: 800uM CaCl
2With 5uM H
3BO
4Mixed liquid; Moving phase 0.125M (NH
4)
2HPO
4, use H
3PO
4Accent PH is 2.1-2.5; The configuration of standard acid sample, standard acid comprises: succinic acid, citric acid, L type L-lactic acid, malonic acid, the left-handed malic acid of L type, Tartaric acid and oxalic acid, configuration concentration is 300mg/L;
B. the collection of root exudates: root system of plant is cleaned, put into 200ml and collect liquid, the morning, 10:00-12:00 collected 2 hours, and cryopreservation is to be purified;
C. purifying: the resin cation dry method that takes by weighing the 5g model and the be Amberlite IR-120B 180mm * 20mm resin column of packing into, connect one the resin column that the 2g model is the resin anion (R.A.) same specification of Dowex 1 * 8-200 is housed, use 10ml deionized water pre-leaching resin column respectively, liquid is collected in siphon, cross earlier kation, after through negative ion;
D. wash-out:, collect eluent in 50ml rotary distillation bottle with 20-30ml 1M HCl wash-out resin anion (R.A.) post:
E. concentrate: adopt rotary evaporator, rotate evaporate to dryness under 40 ℃ of conditions, use the 1000ul deionized water dissolving then after, put into refrigerator, 4 ℃ of preservations are to be measured;
F. high pressure liquid chromatographic analysis condition: with long 250mm, the C of diameter 4mm
18Analytical column, detect wavelength 215nm; Moving phase 0.125M (NH
4)
2HPO
4, PH is 2.1-2.5, flow velocity 0.8-1ml/min; Writing speed: lcm/min; Sample size 20-25ul;
G. detect.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB2004100114047A CN100399022C (en) | 2004-12-30 | 2004-12-30 | Method for detecting low-molecular-weight organic acid in secretion of root system of plant |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB2004100114047A CN100399022C (en) | 2004-12-30 | 2004-12-30 | Method for detecting low-molecular-weight organic acid in secretion of root system of plant |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN1721849A CN1721849A (en) | 2006-01-18 |
| CN100399022C true CN100399022C (en) | 2008-07-02 |
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|---|---|---|---|
| CNB2004100114047A Expired - Fee Related CN100399022C (en) | 2004-12-30 | 2004-12-30 | Method for detecting low-molecular-weight organic acid in secretion of root system of plant |
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Families Citing this family (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104833760A (en) * | 2015-05-25 | 2015-08-12 | 南京信息工程大学 | Method for using high-performance liquid chromatography to measure greenhouse tomato soil low-molecular-weight organic acids |
| CN105021761B (en) * | 2015-07-02 | 2017-09-12 | 华中农业大学 | A kind of method of the content and its true and false of a variety of organic acids in detection grape juice |
| CN105866310A (en) * | 2016-05-20 | 2016-08-17 | 大连大学 | Method for measuring contents of organic acids in blueberries |
| CN110967432B (en) * | 2019-12-25 | 2021-03-30 | 江南大学 | Method for measuring content of organic acid in root exudates |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| GB2174616A (en) * | 1985-04-24 | 1986-11-12 | Yokogawa Hokushin Electric | Method for analysis of organic acids and apparatus therefor |
| JPH02208559A (en) * | 1989-02-08 | 1990-08-20 | Hitachi Ltd | Analysis of organic acid |
| CN1532544A (en) * | 2003-03-20 | 2004-09-29 | 华中科技大学同济医学院附属同济医院 | Method for detecting organic acid/amino acid metabolite in urine by gas chromatography-mass spectrometry of filter paper sheet |
-
2004
- 2004-12-30 CN CNB2004100114047A patent/CN100399022C/en not_active Expired - Fee Related
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| GB2174616A (en) * | 1985-04-24 | 1986-11-12 | Yokogawa Hokushin Electric | Method for analysis of organic acids and apparatus therefor |
| JPH02208559A (en) * | 1989-02-08 | 1990-08-20 | Hitachi Ltd | Analysis of organic acid |
| CN1532544A (en) * | 2003-03-20 | 2004-09-29 | 华中科技大学同济医学院附属同济医院 | Method for detecting organic acid/amino acid metabolite in urine by gas chromatography-mass spectrometry of filter paper sheet |
Non-Patent Citations (3)
| Title |
|---|
| 反相高效液相色谱法测定植物组织及根分泌物中草酸. 俞乐等.分析化学，第30卷第9期. 2002 * |
| 植物根系分泌物中有机酸的分析方法. 杨红等.分析测试学报，第20卷第4期. 2001 * |
| 研究植物根系分泌物的方法. 郜红建等.植物生理学通讯，第39卷第1期. 2003 * |
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| CN1721849A (en) | 2006-01-18 |
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