阅读说明：本技术 一种提升三叶青块根中黄酮含量的室内光照控制方法 (Indoor illumination control method for improving flavone content in radix tetrastigme root tuber ) 是由 韩素芳 程诗明 成亮 吴翠蓉 徐梁 徐秀荣 于 2021-10-15 设计创作，主要内容包括：本发明公开了一种提升三叶青块根中黄酮含量的室内光照控制方法,对三叶青单株进行1500-2000lux白光+0.02-0.03μW/cm～(2)紫外光的光照组合照射；其中,三叶青单株为将一年生三叶青苗栽植于控根容器中,在0.6～0.7郁闭度的竹林下栽培1年所得；通过控制光照强度和光照时间,提高了黄酮的合成量。(The invention discloses an indoor illumination control method for improving flavone content in radix tetrastigme root tuber, which is used for carrying out 1500-2000lux white light +0.02-0.03 mu W/cm on a single radix tetrastigme plant 2 Illumination combination irradiation of ultraviolet light; wherein, the radix tetrastigme single plant is obtained by planting an annual radix tetrastigme seedling in a root control container and cultivating for 1 year in a bamboo forest with 0.6-0.7 canopy density; by controlling the illumination intensity and illumination time, the synthesis amount of flavone is improved.)

1. An indoor illumination control method for improving flavone content in radix tetrastigme root tuber is characterized in that 1500-2000lux white light +0.02-0.03 mu W/cm is carried out on a single radix tetrastigme plant²Illumination combination irradiation of ultraviolet light; wherein the radix tetrastigme single plant is obtained by planting annual radix tetrastigme seedlings in a root control container and cultivating the seedlings under a bamboo forest with 0.6-0.7 canopy density for 1 year.

2. The indoor illumination control method for improving the flavone content in the radix tetrastigme root tuber as claimed in claim 1, wherein the irradiation time of the white light is 8-16 points; the irradiation time of the ultraviolet light is 11 to 13 points; irradiation was continued for 45 days.

Technical Field

The invention relates to the technical field of radix tetrastigme planting, in particular to an indoor illumination control method for improving flavone content in radix tetrastigme tuberous roots.

Background

Tetrastigma hemsleyanum Diels et Gilg is a grape family (Vitacvae) cliff climbing plant (Tetrastigma) which is a folk common Chinese herbal medicine and is also called anoectochilus roxburghii calabash, trifoliate flatvine and the like. Radix Apioris Fortunei root tuber (growing period 3-4 years) has antitumor, antiviral, liver protecting, antiinflammatory and analgesic effects. Modern medical research shows that: radix Apioris Fortunei contains abundant radix Apioris Fortunei flavone, polysaccharide, and kaempferol. Research shows that radix tetrastigme flavone, beta-sitosterol, polysaccharide, kaempferol and the like can effectively inhibit the proliferation of tumor cells and promote the apoptosis of the tumor cells, has good anti-tumor effect and has no toxic or side effect. The traditional Chinese medicine composition has a good treatment effect on most primary cancers, metastatic cancers and the like, has an obvious improvement effect on various adverse reactions caused by tumor chemotherapy, such as low appetite, vomiting, nausea, hair loss, leukopenia and the like, has a pain relieving effect on late-stage patients, and can effectively improve the immune function of a human body; the radix tetrastigme has broad spectrum (suitable for patients with various solid tumors and non-solid tumors), high efficiency (capable of quickly improving malignant symptoms such as cancerous fever, pain, hydrothorax and ascites, leucopenia, anorexia, hypodynamia and the like), quick response and high clinical total effective rate), no toxicity (being a pure natural plant extract, having no toxic or side effect after tests), no drug resistance (being capable of inhibiting tumors in all directions, multiple sections and multiple targets, completely different from the cytotoxic effect of chemotherapeutic drugs, and having no drug resistance in clinical application).

The main active ingredients of the radix tetrastigme are flavonoids which mainly comprise procyanidine B1, catechin, procyanidine B2, rutin, isoquercitrin, kaempferol-3-O-rutinoside, astragalin, quercitrin and the like, but the composition of the radix tetrastigme flavone in different producing areas is obviously different. For a long time, the radix tetrastigme medicinal material always depends on wild resources, and on one hand, non-regenerative mining causes the storage amount of the wild radix tetrastigme resources to be sharply reduced, and many traditional production areas have no medicine to be mined; on the other hand, the variation of the effective components of the wild radix tetrastigme in different producing areas is obviously higher than that of the wild radix tetrastigme cultured artificially, and the quality of the radix tetrastigme is influenced. Therefore, the enhancement of the research on the radix tetrastigme wild-imitating cultivation technology is an urgent need and an effective way for the sustainable development of the radix tetrastigme.

The radix tetrastigme is suitable for growing in high-humus forests such as valleys and shrubs with about 0.7 canopy density, occasionally grows in places with direct sunlight, but the growth is poor. With the first crop of wild-simulated cultivated radix tetrastigme entering the harvesting period, the phenomena of low yield, large quality difference and even no root tuber formation gradually draw attention of forest farmers and researchers. The accurate cultivation technology based on the research of the influence of environmental factors such as illumination, temperature, moisture and the like on the growth and development of the radix tetrastigme becomes a difficult point which needs to be solved urgently at present. The genuine medicinal materials are products adapting to environmental stress for a long time, and the light environment under the forest is more complex than that in a greenhouse and a nursery land due to the existence of a canopy, so that the light factor becomes one of important factors influencing the genuine property of the medicinal materials of the radix tetrastigme under the forest. An accurate light control technology is established, which is not only beneficial to improving the wild-imitating cultivation technology of the radix tetrastigme under the forest, but also beneficial to improving the indoor cultivation quality.

Therefore, the problem that needs to be solved by the technical personnel in the field is how to provide a light control method for improving the flavone content in the radix tetrastigme tuberous root and ensuring the tuberous root yield.

Disclosure of Invention

In view of the above, the invention provides an indoor illumination control method for improving the flavone content in radix tetrastigme root tuber, and the flavone content in the root tuber is improved by controlling the illumination intensity and the illumination time.

In order to achieve the purpose, the invention adopts the following technical scheme:

an indoor illumination control method for increasing flavone content in radix Apioris Fortunei root tuber comprises subjecting radix Apioris Fortunei single plant to 1500-2000lux white light +0.02-0.03 μ W/cm²Illumination combination irradiation of ultraviolet light; wherein the radix tetrastigme single plant is obtained by planting annual radix tetrastigme seedlings in a root control container and cultivating the seedlings under a bamboo forest with 0.6-0.7 canopy density for 1 year.

Plants respond by regulating the expression of series enzyme genes such as Phenylalanine Ammonia Lyase (PAL) and chalcone synthase (CHS), and regulating the synthesis and accumulation of flavone, and the activity and molecular synthesis level of these enzymes are influenced by environmental factors such as illumination, temperature and water. PAL and CHS are two key enzymes in the anabolic pathway of flavones. PAL is a key enzyme and a rate-limiting enzyme which are connected with the primary metabolism and the phenylpropanoid metabolism and catalyze the first step reaction of the phenylpropanoid metabolism, has the molecular weight of 220-330kDa and is an acidic protein. PAL is an enzyme which is most studied on the influence of light stress on flavone accumulation at present. The content of total flavone has a trend substantially consistent with that of PAL enzyme activity, and slightly lags behind the change of enzyme activity. The light stress first induces the expression of PAL enzyme to realize the adjustment of downstream branch of flavone anabolism pathway. CHS is the first rate-limiting enzyme in the flavonoid substance biosynthesis pathway, has the molecular mass of 42-45 kDa, and catalyzes the condensation of 3 acetate groups of malonyl-coenzyme A and one acetate group of p-hydroxyphenylacryloyl-coenzyme to generate chalcone. The light intensity and light quality affect the enzyme activity and gene expression in the plant body, and the activity is increased along with the increase of the light intensity. PAL activity is mainly regulated by ultraviolet light, mRNA generated by CHS gene expression is regulated by blue light and ultraviolet light, and the response of the PAL activity and the mRNA generated by CHS gene expression to light is more obvious than that of other enzymes. Until now, researchers in related fields have studied response rules of light stress flavone of various plants such as arabidopsis thaliana, ginseng, tartary buckwheat and the like, and the influence of light intensity on different plant flavonoids is different, and the content of most plant flavonoids is increased along with the increase of the light intensity. The photoplasmic effect is that short-band light can promote the accumulation of flavone substances, and especially ultraviolet light UV-B (280-315nm) can significantly influence the accumulation of flavonoid compounds (called UV-B filter).

As the preferred technical scheme of the invention, the irradiation time of the white light is 8-16 points; the irradiation time of the ultraviolet light is 11 to 13 points; irradiation was continued for 45 days.

The specific implementation mode is as follows:

the technical solutions in the embodiments of the present invention are clearly and completely described below, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.

The embodiment of the invention discloses a method for improving the content of flavone in radix tetrastigme, which comprises the following specific steps:

example 1 biomass and flavone accumulation rule under white light stress of leaves, stems and root tubers of radix tetrastigme;

single white light irradiation experiment: adopting cultivation shelves, placing 50 radix Apioris Fortunei seedlings (2 years old, obtained by planting annual radix Apioris Fortunei seedlings in a root control container under a bamboo forest with 0.6-0.7 canopy density for 1 year, the vine length is about 30 centimeters, the leaf color is more consistent, using wire netting to build a platform at the middle lower part of each cultivation shelf, spreading radix Apioris Fortunei on the wire netting, and uniformly receiving light irradiation, installing a white light source (40W white light fluorescent tube, measuring non-ultraviolet light intensity by an ultraviolet irradiator), measuring light intensity by an illumination intensity measuring instrument (measuring the illumination intensity of four corners and central point of the cultivation shelf respectively, and taking the average value as the illumination intensity value of the layer), adjusting the number of the open tubes and the distance between the radix Apioris Fortunei and the tubes, treating for 8h every day, continuously irradiating for 45 days, and repeating three times for each treatment. Every 15 days, 5 plants are selected, and leaves, stems, roots and root tubers are collected to be tested.

For four white light laboratory groups: the tetrastigma hemsleyanum Diels et Gilg of the illumination experiment with 1000-.

The experimental results are shown in table 1;

TABLE 1

As can be seen from Table 1: the 1000-plus 1500lux experimental group comprises 12.29g of the fresh weight of single leaf of radix tetrastigme, 12.05g of the fresh weight of stem, 2.42g of the fresh weight of root, 6.61g of the fresh weight of root tuber and 31.16g of the total fresh weight of single plant; the 1500 plus 2000lux experimental group has the fresh weight of single leaf of radix tetrastigme 23.48g, the fresh weight of stem 14.86g, the fresh weight of root 7.13g, the fresh weight of root tuber 4.30g and the total fresh weight of single plant 49.76 g. The 2500-plus 3000lux experimental group has the fresh weight of single leaf of radix tetrastigme of 15.24g, the fresh weight of stem of 11.01g, the fresh weight of root of 5.22g, the fresh weight of root tuber of 2.91g and the total fresh weight of single plant of 34.37 g. The 1500 plus 2000lux experiment group shows that the fresh weights of the leaves, stems and roots of the tetrastigma hemsleyanum Diels et Gilg are the highest of the three groups, but the total weight of tubers is lower than the 1000 plus 1500lux group, because the high illumination intensity promotes photosynthesis, the growth of plants on the ground is promoted, and the nutrition of underground tubers is upwards conveyed, so that the growth of the tubers is slowed down. The size and area of the blade were measured. As the intensity of light increases, the leaf length of the radix tetrastigme tends to increase, while the width tends to decrease. The average leaf area was the largest for the 1000- ­ 1500lux (found 1223lux) group, followed by the 2500- ­ 3000lux group and the smallest for the 1500- ­ 2000lux group.

Under different illumination, the total content of six kinds of flavones, namely quercetin, quercitrin, isoquercitrin, rutin, orientin and isoorientin, in a single plant is measured, and the result is shown in table 2; the highest group was the 1500-2000lux group, in which the average content in leaves was 0.5035mg/g, the average content in stems was 0.2723mg/g, the average content in roots was 0.0884mg/g, and the average content in tuberous roots was 0.0638 mg/g.

TABLE 2

Example 2 the PAL and CHS response law of leaves, stems and tuberous roots of radix tetrastigme to UV-B stress;

white light + UV-B composite illumination experiment: the cultivation method comprises the steps of placing 50 radix tetrastigme seedlings (grown for 2 years, obtained by planting annual radix tetrastigme seedlings in a root control container for 1 year in a bamboo forest with the closing degree of 0.6-0.7, the stems are about 30 centimeters long and the colors of the leaves are consistent, using a wire netting for building a platform at the middle lower part of each cultivation frame, paving the radix tetrastigme on the wire netting for uniformly receiving light irradiation, installing an experimental light source (40W white light fluorescent lamp tube; 40W purple light fluorescent lamp tube with the emission spectrum area of 300-320nm), using a light intensity measuring instrument and an ultraviolet irradiation instrument to measure the light intensity (respectively measuring the light intensity of four corners and the central point of the cultivation frame, and taking the average value as the intensity value of the layer), and adjusting the number of the white light starting lamp tubes and the distance between the wire netting and the lamp tubes to enable the irradiation intensity to reach a set value. The method is characterized in that white light treatment is carried out for 8 hours every day, ultraviolet light irradiation is carried out for 2 hours, and continuous irradiation is carried out for 45 days according to the law of field sunlight. Every 15 days, 5 plants are selected, and leaves, stems, roots and underground root tubers are collected to be tested.

The method specifically comprises the following steps:

ultraviolet light stress experiments were performed on 50 plants per group. According to the ultraviolet intensity characteristics in the forest, white light of 1500 plus 2000lux is adopted in combination with 0.01 mu W/cm²，0.02μW/cm²，0.05μW/cm²，0.1μW/cm²The ultraviolet light of (1) is used as an experimental group, and white light of 1500-. The ultraviolet light irradiation time is 11-13 points per day, and the white light irradiation time is 8-16 points per day. After 45 days of irradiation, 3 pots were sampled from each experimental group, and the plants were decomposed into leaves, young stems, old stems, main roots, fibrous roots, and tuberous roots, and the Phenylalanine Ammonia Lyase (PAL) and chalcone synthase (CHS) activities were measured. The results showed PAL activity in leaves and young stems at 0.1. mu.W/cm²The groups are strongest and respectively 23.63U/g/min and 16.58U/g/min, which shows that the PAL activity in the new tissue is strong under the high ultraviolet irradiation so as to promote the capability of the plant for resisting the external stress. The old stem has the highest PAL activity in the blank experiment, which indicates that the old stem is less affected by the stress of PAL activity. PAL activity in main root, fibrous root and root tuber is significantly higher than blank, wherein 0.02 μ W/cm²The PAL activity of the main root in the experimental group is higher than 198.91U/g/min and 0.01 mu W/cm²The root tuber PAL activity in the group is as high as 103.17U/g/min. The CHS activity of the old stem is highest in blank, which indicates that the old stem is less affected by ultraviolet irradiation. 0.02 mu W/cm²The highest CHS activity of the leaves in the experimental group is 1.36IU/g, the highest CHS activity of the tender stems is blank, and the second CHS activity is 0.01 mu W/cm²Experimental group. The CHS activity is the strongest and is 0.02 mu W/cm²Root tuber in experimental group.

The embodiments in the present description are described in a progressive manner, each embodiment focuses on differences from other embodiments, and the same and similar parts among the embodiments are referred to each other.

The previous description of the disclosed embodiments is provided to enable any person skilled in the art to make or use the present invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the generic principles defined herein may be applied to other embodiments without departing from the spirit or scope of the invention. Thus, the present invention is not intended to be limited to the embodiments shown herein but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.