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Improvement and exploration of the experimental method for identification of nucleic acid chemical components

Date:2021/01/06 13:58:27
Biochemistry experiments are an important part of biochemistry teaching. Through experiments, students' rigorous scientific attitude and style are cultivated, and students' ability to analyze and solve problems are improved. It is an important way to cultivate students' hands-on ability and innovation ability. Through experiments, they can consolidate and deepen biochemistry. The basic knowledge of theoretical courses enables students to better combine theory with practice. The identification of nucleic acid chemical components is one of the important contents in the teaching of biochemistry experiments. Higher vocational colleges have few experimental hours. I have consulted a lot of textbooks, consulted a lot of materials, and done many experiments to find out what is suitable for higher vocational education. The best experiment method for college students. At the same time, the amount of experimental reagents has also been reformed in line with the principles of green chemistry, which saves reagents, reduces pollution, and achieves environmental protection. The experimental methods are summarized as follows:
1. Experimental principle
Nucleic acid is a biological information macromolecule with nucleotide as the basic unit. It is a biological macromolecular compound formed by the polymerization of many nucleotides. It is one of the most basic substances of life. Nucleic acid is widely present in all animal and plant cells and microorganisms. According to the different chemical composition, nucleic acid can be divided into ribonucleic acid (RNA for short) and deoxyribonucleic acid (DNA for short), which are composed of ribose, phosphoric acid, and base. This experiment yeast as the sample. Yeast nucleic acid contains more RNA. The chemical composition of the nucleic acid is identified based on the different colored compounds produced by the reaction of the nucleic acid in strong acid and the corresponding reagent.
1. Ribose is heated in a sulfuric acid solution to convert it into a sugar aldehyde derivative, which reacts with a deglycol reagent to generate a green compound to identify the presence of ribose.
2. Deoxyribose produces ω-hydroxy-γ-ketovaleraldehyde in the acidic solution of ribose. This compound reacts with diphenylamine reagent to produce a blue compound to identify the presence of deoxyribose.
3. Phosphoric acid and ammonium molybdate reagent react to generate yellow ammonium phosphomolybdate to identify the presence of phosphorus in nucleic acids.
4. Purine base reacts with silver hydroxide reagent to generate flocculent purine silver compound to identify the presence of purine silver.
2. Experimental method
1. Yeast hydrolysis
Take 200 mg of dry yeast powder (500 mg of fresh yeast), put it into a large test tube (20*200mm), add 20 ml of 5% sulfuric acid, stir gently with a glass rod, and then put it in a water bath at 100°C for 30 minutes. When the water bath is about 15 minutes, stir once with a glass rod. After 30 minutes in the water bath, take out the test tube and filter it with a long-necked funnel. Leave the filtrate for the following experiment.
2. Identification of hydrolysate
Ribose determination: take 0.5 ml of the filtrate, add 0.5 ml of diethylphenol reagent, and bathe in a water bath at 100°C for 10 minutes to observe any changes in color.
Deoxyribose determination: Take 0.5 ml of the filtrate, add 1 ml of diphenylamine reagent, and put in a water bath at 100°C for 15 minutes to observe the color change.
Purine base determination: Take 1 ml of the filtrate, add 1 ml of 2% silver nitrate, add 3-4 drops of ammonia until the precipitation disappears, mix the test tube, and then stand for about 5 minutes to observe whether the test tube produces flocculent purine silver compounds.
Determination of phosphoric acid: take 0.5 ml of the filtrate, add 0.5 ml of ammonium molybdate reagent, and bath in a constant temperature water bath at 100°C for 5 minutes to observe the color change.
Three, experimental equipment
Electric heating constant temperature water bath, large test tube (20*200mm), middle test tube (15*150mm), long-necked funnel, iron stand, filter paper, test tube rack, glass rod, dropper, 50ml beaker
Four, experimental reagents
1. A pack of 200 mg of dry yeast, pack it before class.
2.5% sulfuric acid
3.10% Ammonium Molybdate Nitrate Reagent
Weigh 4 grams of ammonium molybdate and dissolve it in 100 ml of 10% nitric acid.
4. Diethylphenol reagent
Measure 100 ml of concentrated hydrochloric acid, add 100 mg of ferric chloride, and add 100 mg of diethyl phenol after dissolution. This reagent is ready when. Diethylphenol is also known as: moss black powder, 3,5-dihydroxytoluene.
5. Diphenylamine reagent
Weigh 2 g of diphenylamine and dissolve it in 200 ml of glacial acetic acid, then slowly add 5.5 ml of concentrated sulfuric acid, mix well and store in a brown bottle. The color of the reagent is light yellow. If the reagent is blue-green, it indicates that the glacial acetic acid is impure and cannot.
6. Ammonia
7.2% silver nitrate
Note: Purine base reacts with silver nitrate in alkaline solution to generate flocculent purine silver compound. Purine is an organic compound that is particularly rich in nitrogen. The nitrogen atom on the purine ring has a lone electron pair. The lone electron pair of the silver ion can accept the atom to form a purine silver complex ion. If there is hydroxide ion in the solution, it will coordinate with the purine silver ion. Combined, the purine silver complex ion can become purine silver compound, forming a white flocculent precipitate under weak alkaline conditions. Note that the ammonia water must not be excessive. Purine bases can only form flocculent purine silver compounds with silver nitrate under weakly alkaline (PH8-8.5) conditions. If the ammonia is excessive, silver diammine hydroxide is formed, which is a complex compound, and this substance has no phenomenon.
Five, experimental discussion
The reagents in this experiment include diphenylamine, diphenylamine, ammonium molybdate, concentrated hydrochloric acid, glacial acetic acid, concentrated nitric acid. The reagents are highly volatile and toxic. After adding the reagents, a 100℃ water bath is required. According to green chemistry and conservation Based on economic principles, after many experiments, the experimental reagents have been improved. The amount of improved experimental reagents has been reduced by 3/4. The experimental results are very good, which reduces environmental pollution and avoids volatile reagents on teachers and students. The harm effect has realized the greening of experimental teaching, and has also improved the environmental protection awareness of teachers and students; on the other hand, it has reduced experimental consumption and reagent consumption, reduced experimental costs, and improved the quality and efficiency of talent training.
The improved experimental method is simple and easy to operate, with fewer experimental instruments, and obvious experimental phenomena. Through the experiment, the students’ basic operating skills have been exercised, and the knowledge has also been expanded, so that the students will realize that the experiment must be operated in a serious and meticulous manner. For example, in the determination of purine base, another substance is formed by adding excessive ammonia. During the experiment, we must carefully observe and comprehensively analyze the phenomena and results of the experiment.

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