阅读说明：本技术 一种用于检测橡胶树hcn的检测试剂及制备方法 (Detection reagent for detecting HCN of rubber tree and preparation method thereof ) 是由 杨洪 李德军 代龙军 王立丰 赵溪竹 于 2021-07-28 设计创作，主要内容包括：本发明提供一种用于检测橡胶树HCN的检测试剂,以氯仿为溶剂,每20～25mL氯仿含有4,4'-二氨基二苯甲烷10～15g、乙酰乙酸乙酯铜10～15g、海藻酸钠聚合物2～5g和聚乙烯吡咯烷酮5～8g。本发明制备的检测试剂可直接用于检测橡胶树树皮的HCN含量,该检测试剂具有检测时间快、结果准确、操作简单、成本低等优点,可适用于大量橡胶树树皮的氰化氢含量的检测。(The invention provides a detection reagent for detecting rubber tree HCN, which takes chloroform as a solvent, and each 20-25 mL of chloroform contains 10-15 g of 4,4' -diaminodiphenylmethane, 10-15 g of copper ethyl acetoacetate, 2-5 g of sodium alginate polymer and 5-8 g of polyvinylpyrrolidone. The detection reagent prepared by the invention can be directly used for detecting the HCN content of the rubber tree bark, has the advantages of short detection time, accurate result, simple operation, low cost and the like, and can be suitable for detecting the hydrogen cyanide content of a large amount of rubber tree barks.)

1. A detection reagent for detecting rubber tree HCN is characterized in that: chloroform is used as a solvent, and each 20-25 mL of chloroform contains 10-15 g of 4,4' -diaminodiphenylmethane, 10-15 g of copper ethyl acetoacetate, 2-5 g of sodium alginate polymer and 5-8 g of polyvinylpyrrolidone.

2. A detection reagent for detecting rubber tree HCN according to claim 1, characterized in that: the preparation method of the sodium alginate polymer comprises the following steps:

step S1: adding sodium alginate into a sodium periodate solution with the mass concentration of 0.25-0.30%, and stirring for 1-2 hours in a dark place to obtain a reaction solution;

step S2: adding a calcium chloride aqueous solution with the mass concentration of 5-8% into the reaction liquid, stirring for 50-60 min, adding a sodium polyacrylate solution with the mass concentration of 4-6%, and ultrasonically emulsifying at 25-35 ℃ for 5-10 min to obtain an emulsion;

step S3: and centrifuging the emulsion at the rotating speed of 3000-4000 rpm for 20-30 min to obtain a precipitate, and performing spray drying on the precipitate at the temperature of 75-85 ℃ for 25-35 min to obtain the sodium alginate polymer.

3. A detection reagent for detecting rubber tree HCN according to claim 2, characterized in that: the mass ratio of the sodium alginate to the sodium periodate solution to the calcium chloride aqueous solution to the sodium polyacrylate solution is 0.02-0.025: 0.2-0.3: 55-65: 0.5 to 0.8.

4. The method for preparing a detection reagent for detecting rubber tree HCN according to any one of claims 1 to 3, characterized in that: the method comprises the following steps:

(1) adding 4,4' -diaminodiphenylmethane and copper ethyl acetoacetate into chloroform, and stirring for 10-15 min to obtain a mixed solution;

(2) taking the mixed solution, adding polyvinylpyrrolidone, and carrying out ultrasonic treatment to obtain a primary solution;

(3) and adding a sodium alginate polymer into the primary liquid, thermally shearing, and cooling to obtain the detection reagent.

5. The method for producing a detection reagent for detecting rubber tree HCN according to claim 4, characterized in that: the ultrasonic frequency of the ultrasonic treatment is 2.5-3 MHz, and the time is 25-30 min.

6. The method for producing a detection reagent for detecting rubber tree HCN according to claim 4, characterized in that: the thermal shearing is shearing at the temperature of 50-55 ℃, the shearing rate is 5000-8000 rpm, and the time is 12-18 min.

7. The use method of the detection reagent for detecting rubber tree HCN according to any one of claims 1 to 6, characterized in that: and dropwise adding a detection reagent onto the bark of the rubber tree, standing for 5-10 min, and completing detection of the rubber tree HCN.

Technical Field

The invention relates to the technical field of biochemical detection, in particular to a detection reagent for detecting rubber tree HCN and a preparation method thereof.

Background

Natural rubber has the irreplaceable advantages of synthetic rubber in the aspects of elasticity, wear resistance, ductility and the like, and has wide application in some important industrial fields (such as aviation, aerospace, heavy-duty automobile and other manufacturing industries). Hevea brasiliensis (Hevea brasiliensis, abbreviated as Hevea rubber tree) is the major source of natural rubber. The hevea brasiliensis is a plant of the euphorbiaceae family and is also a main economic crop in tropical regions of China. The traditional breeding cycle of the hevea brasiliensis is about 30 years, and early yield prediction is the key to shortening the breeding cycle. Dead skin (tapping panel drought) is a major factor limiting the yield of rubber trees. According to statistics, the cumulative incidence rate of dead skin in main rubber planting areas in China is between 20% and 40%, the incidence rate of dead skin in serious areas is more than 50%, and the direct economic loss caused by dead skin of rubber trees is up to 30 hundred million yuan each year. In addition, with the popularization of high-yield clone and ethephon stimulation rubber collection technology, the incidence rate and the severity of dead bark of rubber trees tend to increase year by year. Therefore, early prediction, timely prevention and intervention of the occurrence of dead skin are the most effective means for reducing the harm of dead skin at present.

The Brazilian rubber tree is a typical cyanogenic plant, and HCN released by trunk bark tissues is higher when the rubber tree has dead bark symptoms, namely HCN is related to dead bark occurrence. Thus, the dead bark can be early predicted by detecting HCN in the bark tissue. The HCN test paper can be used for directly detecting the leaves of the rubber trees to determine the content of HCN in the leaves, the internal tissues of the barks of the rubber trees are different from the leaves of the rubber trees, the inner tissues of the barks of the rubber trees have various hierarchical structures and have deep colors, and the HCN test paper is not suitable for detecting the content of HCN in the barks of the rubber trees and is easy to cause inaccurate detection results, so that a detection reagent suitable for detecting the content of HCN in the barks of the rubber trees needs to be prepared, and an important basis is provided for early accurate prediction of dead barks of the rubber trees.

Disclosure of Invention

Therefore, the invention aims to provide a detection reagent for detecting rubber tree HCN and a preparation method thereof.

The technical scheme of the invention is realized as follows:

4,4' -diaminodiphenylmethane and copper ethyl acetoacetate react with hydrogen cyanide to form colored compounds, so that the colored compounds can be used for detecting the content of hydrogen cyanide in substances, and the chemical reaction equation is as follows:

2CuAc₂+C₁₇H₂₂N₂+2HCN→2CuCN+[C₁₇H₂₁N₂]Ac+3HAc

the bark is divided into multi-level organizational structures such as exocuticle, cambium and phloem from outside to inside, sodium alginate polymer can make the component of detect reagent form cross-linked interpenetrating network, polyvinyl pyrrolidone can remove substances such as protein, tannin in the bark, reduce the influence of nutrient composition to developer color development effect in the bark, have the effect of surfactant agent simultaneously, sodium alginate polymer and polyvinyl pyrrolidone can play stably and promote the multi-level organizational structure that detect reagent infiltrates into the bark, be favorable to detect reagent and the inside HCN of bark tissue to act on fully.

A detection reagent for detecting rubber tree HCN uses chloroform as a solvent, and each 20-25 mL of chloroform contains 10-15 g of 4,4' -diaminodiphenylmethane, 10-15 g of copper ethyl acetoacetate, 2-5 g of sodium alginate polymer and 5-8 g of polyvinylpyrrolidone.

Further, the preparation method of the sodium alginate polymer comprises the following steps:

step Sa: adding sodium alginate into a sodium periodate solution with the mass concentration of 0.25-0.30%, and stirring for 1-2 hours in a dark place to obtain a reaction solution;

and Sb: adding a calcium chloride aqueous solution with the mass concentration of 5-8% into the reaction solution, stirring for 50-60 min, adding a sodium polyacrylate solution with the mass concentration of 4-6%, and ultrasonically emulsifying at 25-35 ℃ for 5-10 min to obtain an emulsion;

step Sc: centrifuging the emulsion at 3000-4000 rpm for 20-30 min, and spray drying the precipitate at 75-85 ℃ for 25-35 min to obtain the sodium alginate polymer.

Further, the mass ratio of the sodium alginate to the sodium periodate solution to the calcium chloride aqueous solution to the sodium polyacrylate solution is 0.02-0.025: 0.2-0.3: 55-65: 0.5 to 0.8.

Further, the preparation method of the detection reagent for detecting the rubber tree HCN comprises the following steps:

(1) adding 4,4' -diaminodiphenylmethane and copper ethyl acetoacetate into chloroform, and stirring for 10-15 min to obtain a mixed solution;

(2) taking the mixed solution, adding polyvinylpyrrolidone, and carrying out ultrasonic treatment at the ultrasonic frequency of 2.5-3 MHz for 25-30 min to obtain a primary solution;

(3) and adding a sodium alginate polymer into the primary liquid, shearing at the temperature of 50-55 ℃, with the shearing rate of 5000-8000 rpm and the shearing time of 12-18 min, and cooling to obtain the detection reagent.

Further explaining, the use method of the detection reagent for detecting the rubber tree HCN comprises the steps of dropwise adding the detection reagent to the bark of the rubber tree, standing for 5-10 min, and completing detection of the rubber tree HCN.

Compared with the prior art, the invention has the beneficial effects that: the invention can realize the detection of the content of HCN in the bark of the rubber tree by scientifically prepared HCN detection reagent, and has clear detection result and high prediction accuracy.

The sodium alginate polymer and the polyvinylpyrrolidone are adopted, so that substances such as protein, tannin and the like in the bark can be removed, the influence of nutrient components in the bark on the color development effect of the color developing agent is reduced, the polyvinylpyrrolidone can improve the compatibility among components of the detection reagent and improve the dispersibility of liquid, the effect of stabilizing the detection reagent is achieved, the sodium alginate polymer and the polyvinylpyrrolidone are cooperated with each other, the detection reagent is promoted to rapidly and uniformly permeate into the bark tissue, the detection reagent and HCN in the bark are promoted to fully act, the color development effect is more obvious, and the result is more accurate.

Drawings

FIG. 1 is a schematic view of the present invention showing bark divided into inside and outside

Detailed Description

In order to better understand the technical content of the invention, specific examples are provided below to further illustrate the invention.

The experimental methods used in the examples of the present invention are all conventional methods unless otherwise specified.

The materials, reagents and the like used in the examples of the present invention can be obtained commercially without specific description.

Example 1

A detection reagent for detecting rubber tree HCN uses chloroform as a solvent, and each 22mL of chloroform contains 12g of 4,4' -diaminodiphenylmethane, 12g of copper acetoacetic ester, 3g of sodium alginate polymer and 5g of polyvinylpyrrolidone.

The preparation method of the sodium alginate polymer comprises the following steps:

step Sa: according to the mass ratio of sodium alginate to sodium periodate solution to calcium chloride solution to sodium polyacrylate solution of 0.022: 0.2: 60: 0.6, taking 0.022g of sodium alginate, adding 0.2g of sodium periodate solution with the mass concentration of 0.28%, and stirring for 2 hours in a dark place to obtain a reaction solution;

and Sb: adding 60g of calcium chloride aqueous solution with the mass concentration of 6% into the reaction solution, stirring for 55min, adding 0.6g of sodium polyacrylate solution with the mass concentration of 5%, and ultrasonically emulsifying at 30 ℃ for 8min to obtain an emulsion;

step Sc: centrifuging the emulsion at 3500rpm for 25min, collecting precipitate, and spray drying at 80 deg.C for 30min to obtain sodium alginate polymer.

A preparation method of a detection reagent for detecting rubber tree HCN comprises the following steps:

(1) adding 4,4' -diaminodiphenylmethane and copper ethyl acetoacetate into chloroform, and stirring for 12min to obtain a mixed solution;

(2) adding polyvinylpyrrolidone into the mixed solution, and performing ultrasonic treatment at frequency of 2.8MHz for 28min to obtain primary solution;

(3) and adding the sodium alginate polymer into the primary liquid, shearing at the temperature of 55 ℃, with the shearing rate of 6500rpm and the shearing time of 15min, and cooling to obtain the detection reagent.

A use method of a detection reagent for detecting rubber tree HCN comprises the steps of dropwise adding the detection reagent to the bark of a rubber tree, standing for 8min, and completing detection of the rubber tree HCN.

Example 2

A detection reagent for detecting rubber tree HCN uses chloroform as a solvent, and each 20mL of chloroform contains 10g of 4,4' -diaminodiphenylmethane, 10g of copper ethyl acetoacetate, 2g of sodium alginate polymer and 5g of polyvinylpyrrolidone.

The preparation method of the sodium alginate polymer comprises the following steps:

step Sa: according to the mass ratio of sodium alginate to sodium periodate solution to calcium chloride solution to sodium polyacrylate solution of 0.025: 0.3: 65: 0.8, taking 0.025g of sodium alginate, adding 0.3g of sodium periodate solution with the mass concentration of 0.28%, and stirring for 2 hours in a dark place to obtain reaction liquid;

and Sb: adding 65g of calcium chloride aqueous solution with the mass concentration of 6% into the reaction solution, stirring for 55min, adding 0.8g of sodium polyacrylate solution with the mass concentration of 5%, and ultrasonically emulsifying at 30 ℃ for 8min to obtain an emulsion;

step Sc: centrifuging the emulsion at 3500rpm for 25min, collecting precipitate, and spray drying at 80 deg.C for 30min to obtain sodium alginate polymer.

Example 3

A detection reagent for detecting rubber tree HCN uses chloroform as a solvent, and each 25mL of chloroform contains 15g of 4,4' -diaminodiphenylmethane, 15g of copper ethyl acetoacetate, 5g of sodium alginate polymer and 8g of polyvinylpyrrolidone.

The preparation method of the sodium alginate polymer comprises the following steps:

step Sa: according to the mass ratio of sodium alginate to sodium periodate solution to calcium chloride aqueous solution to sodium polyacrylate solution of 0.020: 0.2: 55: 0.5, taking 0.020g of sodium alginate, adding 0.3g of sodium periodate solution with the mass concentration of 0.30%, and stirring for 2 hours in a dark place to obtain reaction liquid;

and Sb: adding 55g of calcium chloride aqueous solution with the mass concentration of 8% into the reaction solution, stirring for 60min, adding 0.5g of sodium polyacrylate solution with the mass concentration of 5%, and ultrasonically emulsifying at 35 ℃ for 8min to obtain an emulsion;

step Sc: centrifuging the emulsion at 4000rpm for 25min, and spray drying the precipitate at 85 deg.C for 30min to obtain sodium alginate polymer.

The preparation method of the detection reagent for detecting the rubber tree HCN of the embodiment 2 and the embodiment 3 comprises the following steps:

(1) adding 4,4' -diaminodiphenylmethane and copper ethyl acetoacetate into chloroform, and stirring for 12min to obtain a mixed solution;

(2) adding polyvinylpyrrolidone into the mixed solution, and performing ultrasonic treatment at a frequency of 3MHz for 30min to obtain a primary solution;

(3) and adding the sodium alginate polymer into the primary liquid, shearing at the temperature of 50 ℃, with the shearing rate of 8000rpm and the shearing time of 18min, and cooling to obtain the detection reagent.

A use method of a detection reagent for detecting rubber tree HCN comprises the steps of dropwise adding the detection reagent to the bark of a rubber tree, standing for 10min, and completing detection of the rubber tree HCN.

Comparative example 1

A detection reagent for detecting hevea brasiliensis HCN according to example 1, which is different in that: 0.5g of sodium alginate polymer and 2g of polyvinylpyrrolidone.

Comparative example 2

A detection reagent for detecting hevea brasiliensis HCN according to example 1, which is different in that: 5g of 4,4' -diaminodiphenylmethane, 8g of copper ethyl acetoacetate and 15mL of chloroform.

Comparative example 3

A detection reagent for detecting hevea brasiliensis HCN according to example 1, which is different in that: 18g of 4,4' -diaminodiphenylmethane, 20g of ethyl acetoacetate copper, 30mL of chloroform, 10g of sodium alginate polymer and 12g of polyvinylpyrrolidone.

Comparative example 4

A detection reagent for detecting hevea brasiliensis HCN according to example 1, which is different in that: the preparation method of the detection reagent is different, and chloroform is taken, 4' -diaminodiphenylmethane and copper ethyl acetoacetate are added, and the mixture is stirred for 10-15 min to obtain a mixed solution; taking the mixed solution, adding a sodium alginate polymer, carrying out thermal shearing, and cooling to obtain a primary solution; and adding polyvinylpyrrolidone into the primary solution, and performing ultrasonic treatment to obtain the detection reagent.

First, the experimental process

Respectively selecting barks of 5 rubber trees H1-H5 without dead bark and 5 rubber trees T1-T5 with dead bark in different degrees, wherein T1 represents that the cut surface dead bark length of the rubber trees is less than 2cm, T2 represents that the cut surface dead bark length is 2cm to 1/4 secant length, T3 represents that the cut surface dead bark length accounts for 1/4 to 1/2 of the secant length, T4 represents that the cut surface dead bark length accounts for 1/2 to 3/4 of the secant length, T5 represents that the cut surface dead bark length accounts for 3/4 to full line dead bark of the secant length, dividing the barks into an inner part and an outer part according to a graph 1, and respectively carrying out a color reaction experiment, wherein the experiment results are shown in a table 1:

note: + indicating color development and-indicating no color development

As can be seen from the above table, the color development part is the inner part of the bark, the bark of H1-H5 does not develop color, the bark of T1-T5 presents different degrees of blue-green color development effect, the rubber trees of the dead bark of examples 1-3 present different colors, comparative example 1 only develops color of the rubber trees of T4 and T5, comparative example 2 only develops color of T5, and comparative example 3 does not develop color, and comparative example 4T 3-T5 develops color.

In conclusion, the invention controls the proportion of the 4,4' -diaminodiphenylmethane, the copper ethyl acetoacetate, the chloroform, the sodium alginate polymer and the polyvinylpyrrolidone, so that the prepared detection reagent can be directly used for detecting the HCN content of the rubber tree bark.

The above description is only for the purpose of illustrating the preferred embodiments of the present invention and is not to be construed as limiting the invention, and any modifications, equivalents, improvements and the like that fall within the spirit and principle of the present invention are intended to be included therein.