Application of selenium supplement agent containing organic selenium glucose in production of selenium-rich oranges
阅读说明：本技术 含有机硒葡萄糖的补硒剂在生产富硒橙子中的应用 (Application of selenium supplement agent containing organic selenium glucose in production of selenium-rich oranges ) 是由 王宇光 沈泓舸 吴梦静 于 2021-08-30 设计创作，主要内容包括：本发明自制了含式(I)所示的有机硒代葡萄糖的补硒剂,进一步配制成水溶液,使用输液,在脐橙树干上进行输液,取得了良好的富硒效果。本发明采用输液法给橙树进行补硒,吸收率高,补硒剂利用率高,避免了象喷洒法补硒的浪费大、利用率小的缺点。(The invention prepares the selenium supplement agent containing the organic selenium glucose shown in the formula (I) by self, and further prepares the selenium supplement agent into aqueous solution, and the infusion is carried out on the navel orange trunk by using the infusion, thereby obtaining good selenium-rich effect. The invention adopts the infusion method to supplement selenium for orange trees, has high absorption rate and high utilization rate of the selenium supplement agent, and avoids the defects of large waste and small utilization rate of selenium supplement like a spraying method.)
1. An application of selenium supplement containing organic selenium glucose shown in formula (I) in producing selenium-rich orange,
2. the use of claim 1, wherein: the selenium supplement is used in the form of an aqueous solution, and the concentration of the organic selenium glucose shown in the formula (I) is 0.103-0.308 g/L in terms of selenium.
3. Use according to claim 2, characterized in that: the concentration of the organic selenium glucose shown in the formula (I) is 0.15-0.25 g/L in terms of selenium.
4. Use according to claim 3, characterized in that: the concentration of the organic selenium glucose shown in the formula (I) as the selenium supplement is 0.205g/L in terms of selenium.
5. The use of claim 1, wherein: the selenium supplement also comprises glucose and/or water.
6. The use according to claim 1, characterized in that the method of application is: the selenium supplement agent is used in the form of aqueous solution and is input into the trunk of the orange tree by adopting an infusion method.
7. The use of claim 1, wherein the selenium supplement is prepared by a process comprising:
under the protection atmosphere, stirring Se and sodium borohydride used as substrates in ethanol A in an ice water bath for 4-8 hours, adding glucose and ethanol B into the obtained reaction solution, stirring at normal temperature for 12-36 hours, removing two thirds of ethanol by rotary evaporation, removing the remaining ethanol by vacuum filtration, washing a filter cake with petroleum ether, and draining the petroleum ether to obtain the selenium supplement;
the mass ratio of Se, sodium borohydride and glucose is 1: 0.5-2: 12 to 20.51.
8. The use of claim 7, wherein: the volume of ethanol A is 2.56mL/mmol, calculated as Se species.
9. The use of claim 7, wherein: the volume of ethanol B is 1.03mL/mmol based on the amount of Se.
10. The use of claim 7, wherein: the protective atmosphere is a nitrogen atmosphere.
Technical Field
The invention relates to an application of a selenium supplement containing organic selenium glucose in producing selenium-rich oranges.
Background
In 1817, the swedish chemist jones jacobia bejersey discovered selenium. Beijerius suggests naming this element as selenium, derived from the greek language seleni, meaning "moon".
Selenium is a very important nutrient. Animals with inadequate dietary selenium may experience muscle weakness. But excess selenium is toxic. The supplement of selenium to the human body is mainly through food, because selenium is naturally present in grains, grains and meat. Adults need to ingest 70ug of selenium every day to maintain health and have been shown to be an essential trace element for humans. It is well known that elemental selenium is practically non-toxic. However, hydrogen selenide and other organic selenium reagents are highly toxic. Excessive intake of selenium can lead to lung fluid accumulation, dyspnea, bronchitis, nausea, headache, sore throat and many other health problems, even death.
Selenium is also the only nutrient associated with viruses, and its function of resisting viral infection is direct, affecting viral pathogenicity by preventing mutations in viral genes, which is not possessed by other vitamins and trace elements. The united states Food and Drug Administration (FDA) indicates that selenium not only can reduce the risk of cancer, but also has an anticancer effect. Besides anti-inflammation, thyroid hormone production and liver protection, selenium also has a significant antioxidant effect.
Selenium supplementation to animals is generally carried out in the form of inorganic salt solids. The problem of high toxicity generally exists, and common selenium products cannot be dissolved in water, so that the use of selenium in a farm is greatly limited.
Selenium-rich foods are generally classified into natural selenium-rich foods and exogenous selenium-rich foods (also called artificial organic selenium foods). Among natural selenium-rich sources, most are organic selenium. Inorganic selenium, such as selenite, although widely used as a nutritional supplement, is not naturally present in traditional diets. When selenite is absorbed, it cannot be directly stored in normal body proteins. The food rich in organic selenium maintains selenase activity for a longer period of time than the food containing inorganic selenium during selenium deficiency.
With the attention of people on health, the selenium-rich food gradually gets into the sight of people.
Brazil nuts, which are the highest selenium containing foods. The selenium content can reach 512ug/g, and the nut can be found in Parmesan rainforest, such as Boriveria, Ecuador, Peru, Guiyan, and Brazil.
Selenium-rich broccoli is a common vegetable capable of accumulating selenium. The selenium content of broccoli is up to about 1g/kg when planted in synthetic soilless mixtures containing sodium selenite, the selenium in broccoli being mainly present in the form of selenomethylselenocysteine (SeMSC). The selenium-rich agricultural method is developed in broccoli varieties with natural agricultural conditions, and the health care performance of the broccoli can be further improved.
Selenium-enriched green tea, selenium is an important component of many antioxidant enzymes, such as glutathione peroxidase and thioredoxin reductase. The selenium-rich green tea extract has a higher protective effect on lipid peroxidation in vitro and a higher scavenging activity on free radicals, compared with the common green tea, the selenium-rich green tea extract has a stronger protective effect physiological function on lipid peroxidation, and selenium also influences the flavor of the green tea, because the selenium-rich green tea shows better sweetness and fragrance, but has less bitter taste.
Selenium-rich spirulina is blue-green microalgae, contains high-quality protein and various nutrient substances, has the characteristics of oxidation resistance, free radical removal and strong anti-inflammatory property, and is considered to be a promising dietary selenium supplement. The phycocyanin containing selenium has effective proliferation inhibiting effect on human melanoma A375 cell and human breast cancer MCF-7 cell.
With the development of the times, the high-price toxic selenium additive SeO2，Na2SeO3，Na2SeO4And gradually quitting the historical stage. And with the abundant appearance of selenium-rich foods in life, the indispensable character of the selenium element is directly explained. Indirectly reflects the desideratum of developing a good selenium additive. The selenium-enriched orange production method adopts the selenium glucose aqueous solution as the selenium supplement agent, and adopts the transfusion application method to apply the selenium glucose aqueous solution to the production of selenium-enriched navel oranges, so that the selenium-enriched orange agricultural product is developed, the daily selenium supplement of people can be met, and a certain health care function is achieved. Promoting the further development of the selenium additive industry and the selenium-rich agricultural product industry.
Disclosure of Invention
The invention provides an application of a selenium supplement containing organic selenium glucose in producing selenium-rich oranges in order to produce the selenium-rich navel oranges. The invention prepares the selenium supplement agent containing the organic selenium glucose shown in the formula (I) by self, and further prepares the selenium supplement agent into aqueous solution, and the infusion is carried out on the navel orange trunk by using the infusion, thereby obtaining good selenium-rich effect.
In order to achieve the purpose, the invention adopts the following technical scheme:
the invention provides an application of a selenium supplement containing organic selenium glucose shown in a formula (I) in producing selenium-enriched oranges,
further, the selenium supplement agent is used in the form of an aqueous solution, and the concentration of the organic selenium glucose shown in the formula (I) is 0.103-0.308 g/L, preferably 0.15-0.25 g/L, and particularly preferably 0.205g/L in terms of selenium.
Preferably, the selenium supplement further comprises glucose and/or water.
Preferably, the method of application is: the selenium supplement agent is used in the form of aqueous solution and is input into the trunk of the orange tree by adopting an infusion method.
Specifically, the selenium supplement agent is prepared by the following method:
under protective atmosphere (such as inert gas or nitrogen, preferably nitrogen), with Se and sodium borohydride (NaBH)4) Stirring in an ice water bath for 4-8h (preferably 6h) in ethanol A as a substrate, adding glucose and ethanol B into the obtained reaction solution, stirring for 12-36 (preferably 24h) at normal temperature, performing rotary evaporation to remove two thirds of ethanol, performing vacuum filtration to remove the remaining ethanol, washing a filter cake with petroleum ether, and draining the petroleum ether to obtain the selenium supplement (off-white solid);
the mass ratio of Se, sodium borohydride and glucose is 1: 0.5-2: 12 to 20.51 (preferably 1: 0.65: 10.26).
The Se is completely reacted to generate the organic selenium glucose. The off-white solid contains almost exclusively organic selenium glucose and glucose.
Further, the volume of ethanol A is 2.56mL/mmol based on the amount of Se as a substance; the volume of ethanol B is 1.03mL/mmol based on the amount of Se. Ethanol a and ethanol B are both ethanol and are labeled A, B only to distinguish the different stages of ethanol without other special meanings.
Preferably, the selenium supplement agent is further prepared into an aqueous solution, the mass of the Se is 0.103-0.308 g/L (preferably 0.15-0.25 g/L, particularly preferably 0.205g/L) of the aqueous solution of the selenium supplement agent, and good selenium enrichment effect is achieved by using infusion and carrying out infusion on the navel orange trunks.
Compared with the prior art, the invention has the beneficial effects that:
(1) an effective navel orange selenium enrichment method is developed, and a self-made organic selenium glucose compound (I) water solution is used as a selenium supplement agent, so that the advantages of the water aqua are reflected: convenient use, cleanness and environmental protection, and simultaneously, the selenium-glucose absorption enhancer conforms to the characteristic that orange trees can absorb aqueous solution more easily and promotes the absorption of selenium glucose.
(2) The invention adopts the infusion method to supplement selenium for orange trees, has high absorption rate and high utilization rate of the selenium supplement agent, and avoids the defects of large waste and small utilization rate of other methods such as the spraying method for supplementing selenium.
(3) The organic selenium glucose aqueous solution is used as a selenium supplement agent, has low preparation cost, simple post-treatment and good water solubility, and is easy to prepare into a water aqua which has low cost and is easy to be absorbed by plants.
Drawings
FIG. 1 is a mass spectrum of organoselenium glucose (I): MS (EI) M/z 244 (M)+-H2O),262(M+)。
FIG. 2 is a standard curve of net selenium content versus net fluorescence.
Detailed Description
The invention will be further described with reference to specific examples, but the scope of the invention is not limited thereto.
In the present invention, 2,3-Diaminonaphthalene (2,3-Diaminonaphthalene, abbreviated as DAN, CAS:771-97-1) is purchased from Sahn's chemical technology (Shanghai) Co., Ltd.
Example 1: synthesis of organic selenium glucose Compound (I)
To a 250mL reaction flaskTo this was added 1.14g (30mmol) of NaBH4And 1.54g (19.5mmol) of Se powder, flushing nitrogen into a reaction bottle, adding 50mL of ethanol under the condition of ice-water bath, stirring, and placing the reaction bottle at normal temperature and stirring for 6 hours when no air bubbles emerge. Then, 36g (200mmol) of glucose and 20mL of ethanol were added thereto, and the mixture was stirred at room temperature for 24 hours (complete reaction of Se powder). Approximately two-thirds of the ethanol was evaporated in a rotary evaporator, the remaining ethanol was vacuum filtered using a buchner funnel, the filter cake was washed with 30mL of petroleum ether, and the petroleum ether was drained to give 37.05g of organic selenized glucose (I) as an off-white solid. The mass spectrum is characterized as follows: MS (EI) M/z 244 (M)+-H2O),262(M+).
Example 2: preparation of selenium-rich orange by infusion method
(1) Preparation of aqueous glucose selenium solution
5.000g, 10.000g and 15.000g of the selenoglucose prepared in example 1 were weighed out and dissolved in 1.9L of distilled water, and then dissolved in 2L (the selenium concentration was 0.103g/L, 0.205g/L and 0.308 g/L).
(2) Application of selenium supplement agent of glucose selenium aqueous solution
At the position about 80 centimeters above the tree trunk soil in the fruiting period of Gannan navel orange in Ganci, Jiangxi city, Renjin county, a drill bit with the diameter of 1mm is used for drilling downwards in an inclined mode at an angle of 45 degrees, and the drilling depth is about 2-3 cm. And opening a flow rate control switch of the fruit and vegetable infusion apparatus to allow air in the tube to flow out, and forcibly inserting the needle head into the hole. Hanging the aqueous solution of glucose and selenium prepared in the step (1) in the embodiment 2 at a high position to ensure that the selenium supplement agent and the needle hole are kept at a relative height, so that the infusion apparatus for fruits and vegetables can help the aqueous solution of glucose and selenium to flow the selenium supplement agent onto the trunk of the orange tree for 24 hours to enable the orange tree to absorb the selenium for 20 days; the above operation was repeated once on each of day 21, day 41 and day 61.
The above experiments were divided into three groups according to the concentration of the selenium supplement: 0.103g/L group, 0.205g/L group, 0.308g/L group; three orange trees are selected from each group for three parallel experiments.
Example 3: determination of selenium content of selenium-rich orange
The invention adopts a national standard method to measure the selenium content of the selenium-enriched orange.
The detection principle is as follows: will try outDigesting with mixed acid to convert selenium compound into inorganic selenium Se4+Se under acidic conditions4 +Reaction with 2,3-Diaminonaphthalene (2,3-Diaminonaphthalene, abbreviated as DAN) produced 4, 5-benzokohlrabi selenol (4, 5-benzopimaselenol) which was then extracted with cyclohexane and evaluated on the machine. 4,5-benzo kohlrabi selenium brain emits fluorescence with the wavelength of 520nm under the action of excitation light with the wavelength of 376nm, and the fluorescence intensity is measured and compared with a standard series for quantification.
Preparing a reagent:
hydrochloric acid solution (1%): 5mL of hydrochloric acid (12mol/L) was weighed out, diluted with water to 500mL, and mixed well.
DAN reagent (1 g/L): the reagent was prepared in a dark room. 0.2g of DAN was weighed into a conical flask with a cap, and 200mL of a hydrochloric acid solution (1%) was added thereto, followed by shaking for about 15min to dissolve all the components. About 40mL of cyclohexane was added and shaking was continued for 5 min. Pouring the solution into a separating funnel filled with glass wool (or absorbent cotton), filtering a cyclohexane layer after layering, collecting a DAN solution layer, and repeatedly purifying with cyclohexane until the fluorescence in the cyclohexane is reduced to the minimum (about 5-6 times). The purified DAN solution was stored in a brown bottle, and cyclohexane having a thickness of about 1cm was added to cover the surface layer, and the mixture was stored at 0 ℃ to 5 ℃. If necessary, purified once more with cyclohexane before use.
Nitric acid-perchloric acid mixed acid (9+ 1): 900mL of nitric acid was mixed with 100mL of perchloric acid.
Hydrochloric acid solution (6 mol/L): weighing 50mL of hydrochloric acid, slowly adding into 40mL of water, cooling, adding water to a constant volume of 100mL, and mixing uniformly.
Aqueous ammonia solution (1+ 1): 5mL of water was mixed with 5mL of aqueous ammonia (14.8 mol/L).
EDTA mixed solution:
EDTA solution (0.2 mol/L): EDTA-2Na74g was weighed, added water and heated to dissolve completely, cooled and diluted to 1000mL with water.
Hydroxylamine hydrochloride solution (100 g/L): 10g of hydroxylamine hydrochloride is weighed and dissolved in water, diluted to 100mL and mixed evenly.
Cresol red indicator (0.2 g/L): weighing 50mg of cresol red, dissolving in a small amount of water, adding 1 drop of ammonia water solution (1+1), adding water to dilute to 250mL after completely dissolving, and mixing uniformly; .
50mL of each of the EDTA solution (0.2mol/L) and the hydroxylamine hydrochloride solution (100g/L) was taken, 5mL of cresol red indicator (0.2g/L) was added, and the mixture was diluted to 1L with water and mixed well.
Hydrochloric acid solution (1+9), 100mL of hydrochloric acid is measured, slowly added into 900mL of water, and mixed evenly.
And (3) standard substance:
selenium standard solution (1000 mg/L): 2.198g of sodium selenite is taken and is made into 1L by hydrochloric acid solution (1%).
Preparation of standard solution:
selenium standard intermediate (100 mg/L): accurately sucking 1.00mL of selenium standard solution (1000mg/L) into a 10mL volumetric flask, adding hydrochloric acid solution (1%) to a constant volume to a scale, and uniformly mixing.
Accurately sucking 0.50mL of selenium standard intermediate solution (100mg/L), diluting to 1000mL with hydrochloric acid solution (1%), and mixing.
Preparation of a standard curve:
0mL, 0.0912mL, 0.2736mL, 0.4560mL, 0.9120mL, 1.296mL, 1.824mL, 2.280mL, 6.840mL and 11.400mL (corresponding to the mass of selenium contained: 0. mu.g, 0.00456. mu.g, 0.01368. mu.g, 0.0228. mu.g, 0.0456. mu.g, 0.0684. mu.g, 0.0912. mu.g, 0.114. mu.g, 0.342. mu.g and 0.570. mu.g) were pipetted into an EP tube, followed by addition of a hydrochloric acid solution (1+9) to 15mL, addition of 60mL of an EDTA mixture, and adjustment to a light reddish orange color (pH1.5 to 2.0) with an aqueous ammonia solution (1+1) and a hydrochloric acid solution (1+9), respectively. The procedure was carried out in a dark room by adding 9mL of DAN reagent (1g/L), mixing, heating in a boiling water bath for 5min, taking out, cooling, adding 3mL of cyclohexane, shaking for 4min, transferring the whole solution into a separatory funnel, separating the layers, discarding the aqueous layer, carefully transferring the cyclohexane layer from the top of the separatory funnel to a test tube with a lid, and not adding water drops into the cyclohexane. The fluorescence intensity was measured by placing 300. mu.L of each of the above cyclohexane samples in a 96-well plate.
Standard curve:
note: the net fluorescence is fluorescence-6495, and the fluorescence at selenium content of 0 is taken as zero point.
Selenium content determination of selenium-enriched orange
Digestion of selenium-enriched orange sample
0.5g to 3g of oranges matured after the selenium supplement is applied in example 2 are accurately weighed, cut into pieces with scissors, placed in a conical flask, added with 10mL of nitric acid-perchloric acid mixed acid (9+1) and a few glass beads, and covered with a watch glass for cold digestion overnight. Heating on an electric heating plate the next day, heating at 180 deg.C for about 30min, and timely adding nitric acid to prevent evaporation. When the solution becomes clear and colorless with white smoke, the solution is continuously heated to the residual volume of about 2mL, the solution is cut and can not be evaporated to dryness, 5mL of hydrochloric acid solution (6mol/L) is added after the solution is cooled, the solution is continuously heated until the solution becomes clear and colorless with white smoke, the solution is continuously heated to the residual volume of about 2mL, and the solution is cooled. The same operation method is used for digesting the orange sample in the blank control group, and the blank control group is Gannan navel orange in Gannan county of Gannan city in the general Jiangxi province without adding the selenium supplement.
Selenium content determination of selenium-enriched orange
After 2mL of the digested sample was added with a hydrochloric acid solution (1.2mol/L) to 5mL, 20mL of an EDTA mixture was added, and the mixture was adjusted to light red orange color (pH1.5 to 2.0) with an ammonia solution (50%) and a hydrochloric acid solution (1.2 mol/L). The procedure was carried out in a dark room by adding 3mL of DAN reagent (1g/L), mixing, heating in a boiling water bath for 5min, taking out, cooling, adding 3mL of cyclohexane, shaking for 4min, transferring the whole solution into a separatory funnel, separating the layers, discarding the aqueous layer, carefully transferring the cyclohexane layer from the top of the separatory funnel to a test tube with a lid, and not adding water drops into the cyclohexane. And (3) respectively taking 300 mu L of the cyclohexane samples in the series, measuring the fluorescence intensity, and obtaining the selenium content according to the relation equation y between the net selenium content and the net fluorescence intensity, which is obtained in the previous step, of 90139 x-140.89. The same operation method is used for measuring the selenium content of the oranges in the blank control group, and the blank control group is Gannan navel orange in Gancin county of Gancin city in the general Jiangxi without adding a selenium supplement.
The experimental results are as follows:
the selenium concentration of the first group of selenium supplement agents is 0.103 g/L.
Navel orange sample a
1
2
3
Average
Weighing gram number g
2.1237
2.3291
2.0199
2.157566667
Fluorescence value
36171
41465
36337
37991
Selenium content of mu g
0.402843275
0.461574790
0.404684876
0.423034314
Per gram of selenium content mug
0.189689351
0.198177317
0.200348966
0.196071878
Blank spaceControl groupb
1
2
3
Average
Weighing gram number g
2.3208
2.1511
2.3402
2.2707
Fluorescence value
31218
30358
33938
31838
Selenium content of mu g
0.347894807
0.338353987
0.378070425
0.354773073
Selenium content ug per gram
0.149902967
0.157293472
0.161554749
0.156250396
Subtracting the content of selenium ug per gram of empty
0.039786384
0.040883845
0.038794217
0.039821482
Note: a, adding a selenium supplement agent to obtain mature navel orange with the selenium concentration of 0.205 g/L;
and b, the blank control group is the common Gannan navel orange without the selenium supplement.
Second group: the selenium concentration of the selenium supplement is 0.205 g/L.
Navel orange samplea
1
2
3
Average
Weighing gram number g
2.1723
2.2945
2.3451
2.270633333
Fluorescence value
39105
41127
42289
40840.33333
Selenium content of mu g
0.435393004
0.457825026
0.470716227
0.454644752
Per gram of selenium content mug
0.200429501
0.199531500
0.200723307
0.200228102
Blank control groupb
1
2
3
Average
Weighing the gram number
2.1231
2.0209
2.3732
2.1724
Fluorescence value
28858
27164
32112
29378
Selenium content of mu g
0.321713021
0.302919824
0.357812822
0.327481889
Selenium content ug per gram
0.151529848
0.149893525
0.150772300
0.150731891
Subtracting the content of selenium ug per gram of empty
0.048899653
0.049637975
0.049951007
0.049496211
Note: a, adding a selenium supplement agent to obtain mature navel orange with the selenium concentration of 0.205 g/L;
and b, the blank control group is the common Gannan navel orange without the selenium supplement.
Third group: the selenium concentration of the selenium supplement is 0.308 g/L.
Navel orange samplea
1
2
3
Average
Weighing the gram number
2.2112
2.3237
2.1705
2.235133333
Fluorescence value
39567
44453
40676
41565.33333
Selenium content ug
0.440518422
0.494723594
0.452821642
0.462687886
Selenium content ug per gram
0.199221428
0.212903384
0.208625497
0.20691677
Blank control groupb
1
2
3
Average
Weighing the gram number
2.1468
2.1122
2.3458
2.20160000
Fluorescence value
27000
28905
31082
28995.66667
Selenium content ug
0.301100412
0.322234438
0.346386026
0.323240292
Selenium content ug per gram
0.140255455
0.152558677
0.147662216
0.146825449
Subtracting the content of selenium ug per gram of empty
0.058965973
0.060344707
0.060963281
0.06009132
Note: a, adding a selenium supplement agent to obtain mature navel orange with the selenium concentration of 0.308 g/L;
and b, the blank control group is the common Gannan navel orange without the selenium supplement.
From the above three groups of experimental data, the effect of the glucose selenium aqueous solution selenium supplement agent and the administration method (infusion method) adopted in the present embodiment is very obvious: the first group is a low concentration (0.103g/L) group, and the selenium supplement result is as follows: the selenium content in navel orange is 0.03982 ug/g; the second group is a medium concentration (0.205g/L) group, and the selenium supplement result is as follows: the selenium content in navel orange is 0.04950 ug/g; the third group is a high concentration (0.308g/L) group, and the selenium supplement result is as follows: the selenium content in navel orange is 0.06009 ug/g. With the increase of the concentration of the selenium supplement, the selenium content in the navel orange is correspondingly increased, but according to the national standard of the people's republic of China: the selenium limit sanitary standard (GB 13105-91) in food specifies that the Se content in fruits is less than or equal to 0.05mg/kg, namely less than or equal to 0.05ug/g, so the selenium supplement scheme of the invention preferably selects the second group, namely the selenium concentration of the selenium supplement is 0.205 g/L.
Example 4: preparation method of selenium-rich orange by spraying method
(1) Preparation of aqueous glucose selenium solution
5.000g, 10.000g and 15.000g of the selenoglucose prepared in example 1 were weighed out and dissolved in 1.9L of distilled water, and then dissolved in 2L (the selenium concentration was 0.103g/L, 0.205g/L and 0.308 g/L).
(2) Spray application of glucose selenium aqueous solution selenium supplement agent
Adding the prepared 2L glucose selenium aqueous solution in the step (1) into an automatic sprayer, spraying the mixture to the crown of the selected Gannan navel orange in Ganci, Heijin county of Gancisco in the fruit bearing period for four times, wherein the spraying speed is about 500mL/5 minutes each time, and the spraying interval is 1 hour each time, so that the orange trees are absorbed for 20 days; the above operation was repeated once on each of day 21, day 41 and day 61.
The above experiments were divided into three groups according to the concentration of the selenium supplement: 0.103g/L group, 0.205g/L group, 0.308g/L group; three orange trees are selected from each group for three parallel experiments.
Example 5: determination of selenium content in selenium-rich orange (prepared by spraying method)
(1) Digestion of selenium-enriched orange sample
0.5g to 3g of oranges ripened by the selenium supplement applied in example 4 are weighed out accurately, cut into pieces with scissors, placed in a conical flask, added with 10mL of mixed acid of nitric acid and perchloric acid (9+1) and a few glass beads, covered on a surface dish and digested overnight in a cold state. Heating on an electric heating plate the next day, heating at 180 deg.C for about 30min, and timely adding nitric acid to prevent evaporation. When the solution becomes clear and colorless with white smoke, the solution is continuously heated to the residual volume of about 2mL, the solution is cut and can not be evaporated to dryness, 5mL of hydrochloric acid solution (6mol/L) is added after the solution is cooled, the solution is continuously heated until the solution becomes clear and colorless with white smoke, the solution is continuously heated to the residual volume of about 2mL, and the solution is cooled. The same operation method is used for digesting the orange sample in the blank control group, and the blank control group is Gannan navel orange in Gannan county of Gannan city in the general Jiangxi province without adding the selenium supplement.
(2) Selenium content determination of selenium-enriched orange
After 2mL of the digested sample was added with a hydrochloric acid solution (1.2mol/L) to 5mL, 20mL of an EDTA mixture was added, and the mixture was adjusted to light red orange color (pH1.5 to 2.0) with an ammonia solution (50%) and a hydrochloric acid solution (1.2 mol/L). The procedure was carried out in a dark room by adding 3mL of DAN reagent (1g/L), mixing, heating in a boiling water bath for 5min, taking out, cooling, adding 3mL of cyclohexane, shaking for 4min, transferring the whole solution into a separatory funnel, separating the layers, discarding the aqueous layer, carefully transferring the cyclohexane layer from the top of the separatory funnel to a test tube with a lid, and not adding water drops into the cyclohexane. The fluorescence intensity of 300. mu.L cyclohexane samples in the above series were measured in a 96-well plate, and the selenium content was obtained by the equation (y) of net selenium content to net fluorescence obtained in example 3, 90139 x-140.89. The same operation method is used for measuring the selenium content of the oranges in the blank control group, and the blank control group is Gannan navel orange in Gancin county of Gancin city in the general Jiangxi without adding a selenium supplement.
The experimental results are as follows:
the experimental results are as follows:
the selenium concentration of the first group of selenium supplement agents is 0.103 g/L.
Note: a, adding a selenium supplement agent to obtain mature navel orange with the selenium concentration of 0.205 g/L;
and b, the blank control group is the common Gannan navel orange without the selenium supplement.
Second group: the selenium concentration of the selenium supplement is 0.205 g/L.
Navel orange samplea
1
2
3
Average
Weighing gram number g
2.3733
1.9947
2.4051
2.2577
Fluorescence value
35199
28267
35084
32850
Selenium content of mu g
0.39205993
0.315156481
0.390784122
0.366000178
Per gram of selenium content mug
0.165196111
0.157996932
0.162481445
0.161891496
Blank control groupb
1
2
3
Average
Weighing the gram number
2.1336
2.2257
2.3451
2.2348
Fluorescence value
30662
30612
33237
31503.66667
Selenium content of mu g
0.341726556
0.341171857
0.370293547
0.351063986
Selenium content ug per gram
0.160164302
0.153287441
0.157900962
0.157117568
Subtracting the content of selenium ug per gram of empty
0.005031809
0.004709491
0.004580483
0.004773927
Note: a, adding a selenium supplement agent to obtain mature navel orange with the selenium concentration of 0.205 g/L;
and b, the blank control group is the common Gannan navel orange without the selenium supplement.
Third group: the selenium concentration of the selenium supplement is 0.308 g/L.
Navel orange samplea
1
2
3
Average
Weighing the gram number
2.3113
2.4146
2.3002
2.342033333
Fluorescence value
33139
33447
34449
33678.33333
Selenium content ug
0.369206337
0.372623282
0.383739447
0.375189689
Selenium content ug per gram
0.159739686
0.154320915
0.166828731
0.160296444
Blank control groupb
1
2
3
Average
Weighing the gram number
2.3328
2.0197
2.2652
2.2059
Fluorescence value
32237
26945
32736
30639.33333
Selenium content ug
0.35919957
0.300490243
0.364735464
0.341475092
Selenium content ug per gram
0.153977868
0.148779642
0.161016892
0.154591467
Subtract blankThe content of selenium per gram
0.005761818
0.005541273
0.005811839
0.005704977
Note: a, adding a selenium supplement agent to obtain mature navel orange with the selenium concentration of 0.308 g/L;
and b, the blank control group is the common Gannan navel orange without the selenium supplement.
From the above three groups of experimental data, the effect of the glucose selenium aqueous solution selenium supplement agent and the application method (spraying method) adopted in the present embodiment is not obvious: the first group is a low concentration (0.103g/L) group, and the selenium supplement result is as follows: the selenium content in navel orange is 0.003568 ug/g; the second group is a medium concentration (0.205g/L) group, and the selenium supplement result is as follows: the selenium content in navel orange is 0.004774 ug/g; the third group is a high concentration (0.308g/L) group, and the selenium supplement result is as follows: the selenium content in navel orange is 0.005705 ug/g. Although the concentration of the selenium supplement agent is increased, the selenium content in the navel orange is also not obviously increased, so the selenium supplement effect of the selenium supplement agent applied by the glucose selenium aqueous solution by the spraying method adopted in the embodiment is poor.
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