CN111096116B - Method for treating evodia lepta seeds - Google Patents
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Abstract
The invention relates to a treatment method of evodia lepta seeds. The method comprises the following steps: a: pretreatment of seed coats: (A1) dry rubbing: taking dry and clean evodia lepta seeds, and repeatedly kneading; (A2) screening: screening out the wax powder adhered to the evodia lepta seed coat by a sieve; (A3) wet rubbing: scrubbing the evodia lepta seeds in the step (A2) in clear water, and fishing out; b: stratification or gibberellin treatment: stratifying the evodia lepta seeds of step (A3) with wet sand at 20-40 ℃; or soaking in gibberellin solution, and taking out. The method combines the pretreatment of the seed coat with the wet sand accumulation or the gibberellin soaking, promotes the germination of the seeds together, and greatly increases the germination rate of the evodia lepta seeds.
Description
Technical Field
The invention relates to the field of seed biology, in particular to a method for treating evodia lepta seeds.
Background
Tape-white (Melipope pteleifolia (Champion ex Bentham) T.G.Hartley) is a plant of the genus Meloidogus of the family Rutaceae, and tape-white is mainly originated in thin forests or shrubs in hills, plains, mountains, linea creeks from low to medium altitudes. The Guangdong is distributed in various places in the south of China, mainly produced in Guangdong, Guangxi, Hainan, south of Jiangxi, Fujian, Taiwan, Guizhou, south of Yunnan and the like, the Guangdong is produced in various mountain counties, and India, Malaysia, Indonesia and Philippines are also distributed.
The herb of Dillenwu is a common Chinese herbal medicine in Lingnan, recorded in the book "Lingnan medicine gathering", and recorded by the section of the "Chinese pharmacopoeia" 1977 and other literatures. The evodia lepta is taken as a medicine by drying stems and twigs with leaves, has the effects of bitter taste and cold property, has multiple effects of clearing heat and detoxicating, promoting qi circulation and relieving pain, eliminating dampness and relieving itching and the like, is mainly used for treating febrile high fever and persistent pyrexia, sore throat, heat-toxin sore swelling, rheumatic arthralgia, damp-fire osteodynia, epigastric pain, traumatic swelling and pain and the like, and is externally used for treating skin damp-heat sore and eruption, skin pruritus, hemorrhoids, insect and snake bite, carbuncle swelling, sore and furuncle and the like. The research on the chemical components of the evodia lepta discovers that the evodia lepta medicinal material contains a plurality of components such as flavonoid, alkaloid, volatile oil, chromene and the like. Modern pharmacological research shows that the evodia lepta has the effects of bacteriostasis, anti-inflammation, antipyresis, analgesia, antioxidation, liver protection, blood sugar and blood fat regulation and the like.
The evodia lepta is a raw medicinal material of various formulated preparations such as a biond magnolia rhinitis pill, a Xiaojiean capsule, a cold medicine, a Sanjiuweitai granule, a Sanjin tablet and the like, is one of common medicinal and edible plants, has higher edible value, and is an important raw material of Guangdong herbal tea. Therefore, domestic enterprises have a great demand for the evodia lepta. Although the distribution area of the evodia lepta is wider, wild resources are sporadically distributed. Therefore, the total amount of the wild resource of the evodia lepta is deficient, the acquisition difficulty of the wild resource is high, and the cost is high. At present, many traditional Chinese medicine enterprises mainly meet the needs of the enterprises by importing the evodia lepta from southeast Asia countries. In recent years, a plurality of enterprises begin to plant the evodia lepta in a large area so as to fill the gap of the demand.
At present, two modes of cutting propagation and seed propagation are mainly adopted for cultivating most traditional Chinese medicinal materials. The cost of the cuttage propagation of the evodia lepta is high, and the number of branches suitable for cuttage of the evodia lepta is small. The amount of the evodia lepta seeds is large and easy to obtain, so most enterprises adopt a seed propagation mode to carry out the seedling raising of the evodia lepta. However, the culture period of the evodia lepta seeds is longer than that of cutting propagation, and the germination rate is lower (less than 5%). One of the main reasons for the low germination rate of the evodia lepta seeds is that the evodia lepta seeds have a dormancy time of 5-6 months. In order to improve the seed propagation efficiency and reduce the propagation cost, the traditional treatment method of the thin leaf evodia seeds adopts chemical reagents such as a potassium permanganate solution and a sodium hydroxide solution to soak the seeds, which may cause potential adverse effects on the embryo and the endosperm of the thin leaf evodia seeds and the growth of subsequent seedlings, and the method has the advantages of complex treatment steps, long treatment time and little improvement of the germination rate of the thin leaf evodia seeds obtained after treatment.
Disclosure of Invention
Based on the method, the invention provides a method capable of remarkably improving the germination rate of the evodia lepta seeds.
The specific technical scheme is as follows:
a treatment method of evodia lepta seeds comprises the following steps:
a: pretreatment of seed coats:
(A1) dry rubbing: taking dry and clean evodia lepta seeds, and repeatedly kneading;
(A2) screening: screening out the wax powder adhered to the evodia lepta seed coat by a sieve;
(A3) wet rubbing: scrubbing the evodia lepta seeds in the step (A2) in clear water, and fishing out;
b: stratification or gibberellin treatment: layering the evodia lepta seeds obtained in the step (A3) at 20-40 ℃ by using wet sand; or soaking in gibberellin solution, and taking out.
In one embodiment, in the step (a3), the method for scrubbing the evodia lepta seeds in clear water is as follows: and (4) rubbing in clear water until the water becomes turbid, then changing the water for continuous rubbing until the water does not become turbid, and fishing out.
In one embodiment, in the step (B), the temperature of the lamination is 25-35 ℃.
In one embodiment, in the step (B), the layering time is 10-20 days.
In one embodiment, in the step (B), the layering time is 15-20 days.
In one embodiment, in step (B), the gibberellin solution is an aqueous gibberellin solution.
In one embodiment, in step (B), the concentration of the gibberellin aqueous solution is 100 mg.L-1～400mg·L-1。
In one embodiment, in step (B), the concentration of the gibberellin aqueous solution is 200 mg.L-1～300mg·L-1。
In one embodiment, in the step (B), the gibberellin solution is soaked for 20 to 30 hours.
In one embodiment, in the step (A1), the seeds of the evodia lepta are kneaded until the surface of the evodia lepta seeds is free from wax shedding.
Compared with the prior art, the invention has the following beneficial effects:
the method for treating the evodia lepta seeds comprises the steps of firstly rubbing the seeds by dry rubbing, and grinding off a wax layer on the surfaces of the seeds, so that water can enter seed coats conveniently; then the seeds are rubbed in clean water to wash off grease rich in the seed coats, so that the seed coats are loosened, the air permeability and the water permeability of the seed coats are increased, the mechanical constraint of the seed coats on embryos is reduced, and the physical dormancy of the evodia lepta seeds is effectively relieved. Furthermore, the physiological dormancy of the evodia lepta seeds can be rapidly and effectively relieved by wet sand accumulation at the temperature of 20-40 ℃ and gibberellin solution soaking. The method combines the pretreatment of the seed coat with the wet sand accumulation or the gibberellin soaking, promotes the germination of the seeds together, and greatly increases the germination rate of the evodia lepta seeds. In addition, in the seed coat pretreatment process of the evodia lepta seeds, a chemical reagent is not needed to soak the seed coat, so that the damage of the chemical reagent to the endosperm and embryo tissues in the evodia lepta seeds is avoided.
Drawings
FIG. 1 is a germination diagram of Dillenia indica seeds before and after wet sand stratification in a greenhouse;
FIG. 2 is a diagram of germination of thin evodia seeds before and after treatment with gibberellin solution in a greenhouse;
FIG. 3 is a germination diagram of the evodia lepta seeds before and after pretreatment of the seed coats in the greenhouse.
Detailed Description
The present invention will be described in further detail with reference to specific examples.
1 materials and methods
1.1 Experimental materials
Collecting the evodia lepta fruits with naturally dried and cracked peels on trees in GAP industrialized demonstration bases of Dongyuan county in Heyuan city of Guangdong province, removing the peels and collecting seeds. The gibberellin solution is prepared as follows:
100mg·mL-1gibberellin mother liquor (stored at 4 ℃ refrigerator): 1g of gibberellin powder was weighed on an electronic balance and added to 10mL of ethanol to be sufficiently dissolved.
400mg·L-1Aqueous gibberellin solution (fresh formula): 2mL of the gibberellin mother liquor was measured by a pipette, added to 498mL of purified water, and stirred thoroughly with a stirrer.
300mg·L-1Aqueous gibberellin solution (fresh formula): 1.5mL of gibberellin mother liquor was measured by a pipette, added to 498.5mL of purified water, and stirred thoroughly with a stirrer.
200mg·L-1Aqueous gibberellin solution (fresh formula): 1mL of gibberellin mother liquor was measured by a pipette, added to 499mL of pure water, and stirred thoroughly with a stirrer.
100mg·L-1Aqueous gibberellin solution (fresh formula): 0.5mL of gibberellin mother liquor was measured by a pipette, added to 499.5mL of purified water, and stirred thoroughly with a stirrer.
Example 1
And (3) placing the collected thin evodia seeds in a clean container, and repeatedly rubbing the thin evodia seeds by using two hands until no wax falls off from the surfaces of the thin evodia seeds. And (4) screening coarse wax powder adhered to the evodia lepta seed coats by using a sieve until no obvious wax powder leaks from the sieve. And pouring the evodia lepta seeds of which the wax powder is completely sieved into clear water for scrubbing, and changing the water after the water becomes turbid to continue scrubbing until the water does not become turbid. The layers were stacked on wet sand at 30 ℃ for 5 days.
Example 2
Putting the collected thin evodia seeds into a clean container, and repeatedly rubbing the thin evodia seeds by using two hands until no wax falls off from the surfaces of the thin evodia seeds. And (4) screening coarse wax powder adhered to the evodia lepta seed coats by using a sieve until no obvious wax powder leaks from the sieve. And pouring the evodia lepta seeds of which the wax powder is completely sieved into clear water for scrubbing, and changing water after the water becomes turbid to continue scrubbing until the water does not become turbid. Wet sand stratification was carried out at 30 ℃ for 10 days.
Example 3
Putting the collected thin evodia seeds into a clean container, and repeatedly rubbing the thin evodia seeds by using two hands until no wax falls off from the surfaces of the thin evodia seeds. And (4) screening coarse wax powder adhered to the evodia lepta seed coats by using a sieve until no obvious wax powder leaks from the sieve. And pouring the evodia lepta seeds of which the wax powder is completely sieved into clear water for scrubbing, and changing the water after the water becomes turbid to continue scrubbing until the water does not become turbid. Wet sand stratification was carried out at 30 ℃ for 15 days.
Example 4
Putting the collected thin evodia seeds into a clean container, and repeatedly rubbing the thin evodia seeds by using two hands until no wax falls off from the surfaces of the thin evodia seeds. Removing the adhesion by sievingCoarse wax powder on the seed coat of the evodia lepta till no obvious wax powder leaks out of the screen. And pouring the evodia lepta seeds of which the wax powder is completely sieved into clear water for scrubbing, and changing water after the water becomes turbid to continue scrubbing until the water does not become turbid. At 100 mg.L-1Soaking in gibberellin solution for 24 hr.
Example 5
And (3) placing the collected thin evodia seeds in a clean container, and repeatedly rubbing the thin evodia seeds by using two hands until no wax falls off from the surfaces of the thin evodia seeds. And (4) sieving the coarse paraffin fragments adhered to the evodia lepta seed coats by using a sieve until no obvious paraffin fragments leak from the sieve. And pouring the evodia lepta seeds of which the wax powder is completely sieved into clear water for scrubbing, and changing the water after the water becomes turbid to continue scrubbing until the water does not become turbid. At 200 mg. L-1Soaking in gibberellin solution for 24 hr.
Example 6
Putting the collected thin evodia seeds into a clean container, and repeatedly rubbing the thin evodia seeds by using two hands until no wax falls off from the surfaces of the thin evodia seeds. And (4) screening coarse wax powder adhered to the evodia lepta seed coats by using a sieve until no obvious wax powder leaks from the sieve. And pouring the evodia lepta seeds of which the wax powder is completely sieved into clear water for scrubbing, and changing the water after the water becomes turbid to continue scrubbing until the water does not become turbid. At 300 mg.L-1Soaking in gibberellin solution for 24 hr.
Example 7
Putting the collected thin evodia seeds into a clean container, and repeatedly rubbing the thin evodia seeds by using two hands until no wax falls off from the surfaces of the thin evodia seeds. And (4) screening coarse wax powder adhered to the evodia lepta seed coats by using a sieve until no obvious wax powder leaks from the sieve. And pouring the evodia lepta seeds of which the wax powder is completely sieved into clear water for scrubbing, and changing the water after the water becomes turbid to continue scrubbing until the water does not become turbid. At 400 mg.L-1Soaking in gibberellin solution for 24 hr.
Comparative example 1
And (3) placing the collected thin evodia seeds in a clean container, and repeatedly rubbing the thin evodia seeds by using two hands until no wax falls off from the surfaces of the thin evodia seeds. And (4) sieving the coarse paraffin fragments adhered to the evodia lepta seed coats by using a sieve until no obvious paraffin fragments leak from the sieve. And pouring the evodia lepta seeds of which the wax powder is completely sieved into clear water for scrubbing, and changing the water after the water becomes turbid to continue scrubbing until the water does not become turbid.
Comparative example 2
The harvested evodia lepta seeds are not treated at all.
Comparative example 3
Directly layering the collected evodia lepta seeds in wet sand at 30 ℃ for 15 days.
Comparative example 4
Putting the collected thin evodia seeds into a clean container, and repeatedly rubbing the thin evodia seeds by using two hands until no wax falls off from the surfaces of the thin evodia seeds. And (4) screening coarse wax powder adhered to the evodia lepta seed coats by using a sieve until no obvious wax powder leaks from the sieve. And pouring the evodia lepta seeds of which the wax powder is completely sieved into clear water for scrubbing, and changing the water after the water becomes turbid to continue scrubbing until the water does not become turbid. Wet sand stratification was carried out at 4 ℃ for 15 days.
Comparative example 5
Directly collecting collected evodia lepta seeds at 300 mg.L-1Soaking in gibberellin solution for 24 hr.
Test group one: research on treatment method of evodia lepta seeds in laboratory
Soaking the evodia lepta seeds treated in the embodiments 1-7 and the comparative examples 1-5 in clear water for 24h to swell the seeds. Then, a germination test is carried out in a climatic incubator by adopting a sand-on-sand method (the germination bed is a sand bed, the size of sand grains is uniform, the diameter of the sand grains is 0.4-0.8 mm, and the sand grains are washed by clear water and sterilized at high temperature before use), the test adopts a completely randomized design, 3 times of repetition are arranged, and 50 seeds are repeated every time. The germination conditions were as follows: temperature, 30 ℃ (12h)/20 ℃ (12 h); illumination, 6000LX (12h)/0LX (12 h); humidity, 60% RH. The experiment used a fully randomized design with three biological replicates per group, each replicate 50 seeds. The results are shown in Table 1.
Experiment group two: research on treatment method of evodia lepta seeds in greenhouse
The tape seeds obtained in comparative examples 1-2 and examples 1, 3, 4 and 6 were subjected to germination tests in a greenhouse. Pouring the beautiful soil on clean cement ground, fully mixing with clean water and uniformly stirring, holding the mixture into a ball by hand, stopping adding water, and pouring the uniformly wetted beautiful soil into the strip-shaped flowerpot. Respectively and uniformly sowing untreated and differently treated mature evodia lepta seeds in the strip-shaped flowerpot. The experiment used a fully randomized design, three biological replicates per treatment, 200 seeds per replicate. The treated seeds were imbibed in clear water for 24 hours before germination testing. The germination test is carried out in a greenhouse with sufficient illumination and good ventilation, and watering is carried out every day according to the dry and wet conditions of soil. The germination period is 3 months. The results are shown in Table 2.
TABLE 1 research results of treatment method of Ixeris sonchifolia seeds in laboratory
P <0.01 compared to comparative example 1, P <0.05 compared to comparative example 1;
in table 1, in examples 2 and 3, the germination rates of the thin evodia seeds after the seed coat pretreatment in warm stratification for 10 days and 15 days are 67.33 ± 5.03% and 82.00 ± 3.46%, respectively, and both have statistical differences (P <0.01) compared with the germination rate of comparative example 1 (42.67 ± 4.62%) which is not warm stratification. The germination rate of the evodia lepta seeds subjected to the pretreatment of the seed coats and subjected to warm temperature (30 ℃) stratification for 15 days is the highest, and compared with the germination rate of the evodia lepta seeds subjected to warm temperature stratification for 5 and 10 days, the germination rate has statistical differences (P is less than 0.01, and P is less than 0.01). In comparative example 4, when the dielletia indica seeds after the seed coat pretreatment are subjected to wet sand stratification at a low temperature of 4 ℃, the germination rate is 46.00 +/-4.00%, and the germination rate is not statistically different from that of comparative example 1 (the germination rate is 42.67 +/-4.62%), so that the temperature of the wet sand stratification has a great influence on the treatment effect of the dielletia indica seeds.
In Table 1, in examples 4 to 7, the seed coat was pretreatedThe amount of the seeds of the evodia lepta ranges from 100, 200, 300 and 400 mg.L-1The germination rates of the gibberellin solution of (1) after soaking for 24 hours are 55.33 + -8.08%, 66.67 + -5.77%, 77.33 + -5.03% and 62.00 + -3.46%, respectively, and all have statistical differences (P is 42.67 + -4.62%) compared with comparative example 1 (germination rate is 42.67 + -4.62%)<0.05，P<0.01，P<0.01，P<0.01). Wherein the amount of the evodia lepta seeds subjected to the seed coat pretreatment is 300 mg.L-1The germination rate of the gibberellin solution is the highest after being soaked for 24 hours, and the germination rate is between 100, 200 and 400 mg.L-1Compared with the germination rate of the gibberellin solution after being soaked for 24 hours, the germination rates of the gibberellin solution have statistical difference (P)<0.01，P<0.05，P<0.01). The results show that the germination rate of the evodia lepta seeds can be further improved by treating the evodia lepta solution after the seed coat pretreatment, and when the concentration of the evodia lepta solution is 300 mg.L-1The effect is best.
In Table 1, in comparative example 1, the germination rates of the Ixeris sonchifolia seeds after the seed coat pretreatment are 42.67 + -4.62%, respectively, and compared with comparative example 2 (the germination rate is 4.00 + -2.00%), there is a statistical difference (P)<0.01). The result shows that the germination rate of the evodia lepta seeds after the seed coat pretreatment is obviously improved. In comparative example 3, the germination rate of Dillenia indica seeds directly warm stratified for 15 days is 4.67 + -1.15%, and compared with comparative example 2 (germination rate is 4.00 + -2.00%), there is no statistical difference (P)>0.05). In comparative example 5, the amount of the seeds of Evodia indica is 300 mg. L-1The germination rates of the gibberellin solutions of (1) were 2.67 + -1.15% respectively for 24 hours, and there was no statistical difference (P) from comparative example 2 (germination rates of 4.00 + -2.00%)>0.05). The results show that the germination rate of the evodia lepta seeds cannot be improved by the warm stratification and the gibberellin aqueous solution treatment without the seed coat pretreatment.
Therefore, the seed coat pretreatment process is combined with wet sand stratification or gibberellin soaking respectively, so that the germination rate of the evodia lepta seeds can be obviously improved; all the steps of the invention are complementary, and the lack of any one step can not achieve the purpose of obviously improving the germination rate of the evodia lepta seeds.
TABLE 2 research results of treatment method of evodia lepta seeds in greenhouse
P <0.01 compared to comparative example 1, P <0.05 compared to comparative example 1;
in table 2, the germination pattern results of thin evodia seeds before and after the wet sand stratification treatment in examples 1, 3 and comparative example 1 are shown in fig. 1. The results show that under the greenhouse conditions, the germination rate of the warm-temperature stratification for 15 days is 57.83 +/-6.83%, and compared with the germination rate of the warm-temperature stratification for 0 day group (the germination rate is 33.17 +/-2.57%), the germination rate is statistically different (P)<0.01). The results of germination rates of thin evodia seeds before and after gibberellin solution treatment in examples 4 and 6 and comparative example 1 are shown in fig. 2. The results show that under the greenhouse condition, 100 mg-L-1The germination rate of the gibberellin-treated group is 45.50 + -5.27%, and 0 mg. L-1Compared with the gibberellin-treated group (germination rate of 33.17 +/-2.57%), the gibberellin-treated group has statistical difference (P)<0.01)；300mg·L-1The germination rate of the gibberellin-treated group is 53.00 + -5.07%, and 0 mg. L-1Compared with the gibberellin-treated group, the group has statistical difference (P)<0.01). In addition, in comparative examples 1 and 2, the germination patterns of the evodia lepta seeds before and after the seed coat pretreatment are shown in fig. 3, and the results show that the seed coat pretreatment has statistical difference (P) compared with the seed coat pretreatment without under the greenhouse condition<0.05)。
From the results in table 2, it can be seen that in the greenhouse, the treatment method of the present invention is consistent with the results obtained in the laboratory, and the method for treating the thin leaf tickclover seeds by combining the seed coat pretreatment process with stratification or gibberellin soaking can significantly improve the germination rates of the thin leaf tickler seeds in different environments.
The technical features of the embodiments described above may be arbitrarily combined, and for the sake of brevity, all possible combinations of the technical features in the embodiments described above are not described, but should be considered as being within the scope of the present specification as long as there is no contradiction between the combinations of the technical features.
The above-mentioned embodiments only express several embodiments of the present invention, and the description thereof is more specific and detailed, but not construed as limiting the scope of the invention. It should be noted that, for a person skilled in the art, several variations and modifications can be made without departing from the inventive concept, which falls within the scope of the present invention. Therefore, the protection scope of the present patent shall be subject to the appended claims.
Claims (8)
1. A method for treating evodia lepta seeds is characterized by comprising the following steps:
a: pretreatment of seed coats:
(A1) dry rubbing: taking dry and clean evodia lepta seeds, and repeatedly kneading;
(A2) screening: sieving waxy powder adhered to the evodia lepta seed coat by a sieve;
(A3) wet rubbing: scrubbing the evodia lepta seeds in the step (A2) in clear water, and fishing out;
b: stratification or gibberellin treatment: layering the evodia lepta seeds obtained in the step (A3) with wet sand at 20-40 ℃ for 10-20 days; or soaking in gibberellin solution, and taking out; the gibberellin solution is a gibberellin water solution, and the concentration of the gibberellin water solution is 100 mg.L-1～400mg·L-1The soaking time of the gibberellin solution is 20 to 30 hours.
2. A method for treating Dillenia indica seeds as claimed in claim 1, wherein in step (A3), said method for scrubbing Dillenia indica seeds in clear water is as follows: and (4) rubbing in clear water until the water becomes turbid, then changing the water for continuous rubbing until the water does not become turbid, and fishing out.
3. A method for treating evodia lepta seeds according to claim 1, wherein in step (B) the temperature of said stratification is 25-35 ℃.
4. A method for treating evodia lepta seeds according to claim 1, wherein in step (B) said stratification time is 15-20 days.
5. A method for treating Evodia indica seeds as claimed in any one of claims 1 to 4, wherein in step (B), the temperature of said stratification is 30 ℃ and the time of said stratification is 15 days.
6. A method for treating the seeds of Evodia indica as claimed in any one of claims 1 to 4, wherein in step (B), the concentration of said gibberellin aqueous solution is 200 mg-L-1～300mg·L-1。
7. A method for treating the seeds of Evodia indica as claimed in any one of claims 1 to 4, wherein in step (B), the concentration of said gibberellin aqueous solution is 300 mg-L-1And the soaking time of the gibberellin solution is 24 hours.
8. A method for treating the Dillenia indica Linn seeds as claimed in any claim 1 to 4, wherein the step (A1) is carried out by kneading until the surface of the Dillenia indica Linn seeds is free from wax shedding.
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| CN104285538A (en) * | 2014-09-19 | 2015-01-21 | 华润三九医药股份有限公司 | Treatment method of Melicope ptelefolia (Champ.ex Benth.) Hartley seeds and Melicope ptelefolia (Champ.ex Benth.) Hartley breeding method |
| CN104365220A (en) * | 2014-12-15 | 2015-02-25 | 湖南科技大学 | Cyclocarya paliurus seed treatment method |
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| CN104285538A (en) * | 2014-09-19 | 2015-01-21 | 华润三九医药股份有限公司 | Treatment method of Melicope ptelefolia (Champ.ex Benth.) Hartley seeds and Melicope ptelefolia (Champ.ex Benth.) Hartley breeding method |
| CN104365220A (en) * | 2014-12-15 | 2015-02-25 | 湖南科技大学 | Cyclocarya paliurus seed treatment method |
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