Red soil compound substrate suitable for gold wire orchid planting and application

Abstract

This invention discloses a red soil composite substrate suitable for the cultivation of *Anoectochilus roxburghii* and its application, belonging to the field of *Anoectochilus roxburghii* cultivation technology. It addresses the limitations of existing technologies, such as difficulty in seed germination, slow growth, and dwindling wild resources, as well as the poor aeration and root rot issues in red soil distribution areas during artificial cultivation. These limitations hinder the large-scale and efficient cultivation of *Anoectochilus roxburghii* in red soil regions. The red soil composite substrate provided by this invention is composed of red soil, peat moss, perlite, and bark mixed in a specific volume ratio, specifically (2-4):1:1:1. Through the synergistic effect of the components, it leverages the water and fertilizer retention properties of red soil while improving its aeration. This scientifically and thoroughly solves the technical problems of "acidic, infertile, and compacted" soil, poor aeration, and root rot in the vast red soil areas of southern China, making red soil a suitable cultivation substrate for *Anoectochilus roxburghii*.

Description

Technical Field

[0001] This invention relates to the field of Anoectochilus roxburghii cultivation technology, specifically to a red soil compound substrate suitable for Anoectochilus roxburghii cultivation and its application. Background Technology

[0002] Anoectochilus roxburghii is a perennial herbaceous plant belonging to the genus Anoectochilus in the family Orchidaceae, mainly distributed in areas of my country at altitudes of 50-1600 meters. With its small and beautiful plant shape, elegant leaves, and dark purple foliage, Anoectochilus roxburghii has high medicinal and ornamental value, and market demand is increasing. However, due to the tiny size of its seeds and the lack of endosperm, Anoectochilus roxburghii seeds are extremely difficult to germinate naturally, leading to a continuous decline in wild resources.

[0003] To meet the growing demand for Anoectochilus roxburghii, the artificial cultivation industry for this orchid is developing rapidly. Breeding techniques mainly include artificial culture medium cultivation, protected cultivation, and semi-wild forest cultivation. Although asexual reproduction techniques for Anoectochilus roxburghii are quite mature, large-scale asexual reproduction has led to problems such as severe germplasm degradation, poor resistance to adverse conditions, and reduced medicinal components.

[0004] Yunnan Province has a complex and diverse climate, exhibiting characteristics of monsoon, low-latitude, and plateau climates. It is characterized by small seasonal temperature variations, large daily temperature variations, distinct dry and wet seasons, diverse climate types, and significant vertical climate features. These climatic conditions are generally suitable for the growth of Anoectochilus roxburghii, a plant that prefers cool and humid conditions. However, the cultivation industry for Anoectochilus roxburghii is mainly concentrated in Zhejiang and Fujian provinces, primarily because Yunnan's soil is mostly red soil. Red soil has a granular surface structure, with a deeper layer cemented by iron and aluminum oxides, forming a hardened layer. Its texture is mostly clay or clay loam, with high porosity but few large pores, resulting in limitations such as low mineral nutrient content, high acidity, susceptibility to aluminum and manganese toxicity, and poor fertilizer retention.

[0005] Anoectochilus roxburghii is a small herb with very shallow roots, native to the topsoil of forests rich in humus. It has very high requirements for soil aeration, drainage, and nutrients. Its natural habitat is not red soil, but rather forest undergrowth humus. It cannot grow in normal red soil, which is characterized by being acidic (far more so than the native soil of Anoectochilus roxburghii), infertile (extremely barren for Anoectochilus roxburghii), and compacted (the roots cannot grow normally). Therefore, the climate and soil characteristics of Yunnan are very contradictory for cultivating Anoectochilus roxburghii.

[0006] Therefore, the improvement of red soil is paramount in developing the Anoectochilus roxburghii cultivation industry in Yunnan. Improving the cultivation of Anoectochilus roxburghii with a formula soil based primarily on red soil has significant practical and economic value. Summary of the Invention

[0007] To address the shortcomings and deficiencies of existing technologies, this invention provides a red soil compound substrate suitable for planting Anoectochilus roxburghii, formulated in a specific ratio. This substrate maximizes the activation of secondary metabolic pathways in Anoectochilus roxburghii while ensuring an extremely high transplant survival rate, significantly promoting the biosynthesis of core effective components such as Anoectochilus roxburghii glycosides, polysaccharides, and flavonoids.

[0008] To achieve the above objectives, the present invention provides a red soil composite substrate suitable for planting Anoectochilus roxburghii. The composite substrate is made by mixing red soil, peat moss, perlite and bark in a specific volume ratio. Specifically, the volume ratio is: red soil: peat moss: perlite: bark = (2-4):1:1:1; the perlite has a particle size distribution of 3-6 mm; the peat moss is fine fiber with a size of 0-25 mm.

[0009] Preferably, the volume ratio of each component in the compound matrix is: red soil: peat soil: perlite: bark = 3:1:1:1.

[0010] Preferably, the bark is the dried bark of any one or more of pine, chestnut, and sand trees, and the bark is obtained by pretreatment as follows: after air drying and crushing, the size of the crushed bark is 10-25mm.

[0011] Preferably, the peat soil is finely fibrous with a size of 0-10 mm.

[0012] Preferably, the red soil is clean and free from pests and diseases; the red soil treatment method is to remove pests, sterilize and disinfect it before use; the removal of pests, sterilization and disinfection before use is carried out by sun exposure, and the pH value of the red soil is in the range of 5.5-6.5, without the need for additional adjustment.

[0013] Preferably, the preparation method of the above-mentioned red soil composite substrate suitable for planting Anoectochilus roxburghii includes the following: Raw material preparation: Prepare clean, pest-free red soil, peat moss, perlite and bark. The red soil and peat moss should be in a semi-dry and loose state. Preparation of compound substrate: According to the volume ratio, accurately proportion and mix the red soil, peat moss, perlite and bark. After mixing them evenly, the substrate moisture content should be such that it can be formed into a ball when squeezed in the hand, but crumbles when released. This will give you a red soil compound substrate suitable for planting Anoectochilus roxburghii.

[0014] The above-mentioned red soil compound substrate suitable for planting Anoectochilus roxburghii is used to improve the red soil suitable for planting Anoectochilus roxburghii.

[0015] The preferred indoor seedling cultivation method includes the following steps: S1. Selecting the cultivation site and environment for Anoectochilus roxburghii. Temperature: Golden Thread Orchid prefers cool temperatures, with the average daily temperature maintained between 18-25℃; Air humidity: Prefers high humidity, with relative humidity maintained between 70% and 85%; Light: Shading is required; light intensity should be controlled at 5000-10000 Lux; daily light duration should be 8-10 hours. Cultivation location: Inside greenhouses or cultivation facilities with shade nets; S3. Transplanting of Anoectochilus roxburghii tissue culture seedlings: Carefully transplant healthy and well-maintained Anoectochilus roxburghii tissue culture seedlings into a prepared red soil composite substrate for cultivation and post-cultivation management.

[0016] This invention provides a red soil composite substrate suitable for planting Anoectochilus roxburghii. It has the following beneficial effects: The red soil compound substrate of this invention, suitable for planting Anoectochilus roxburghii, combines red soil with bark, peat moss, perlite and other substrates through substrate improvement technology. This not only gives full play to the water and fertilizer retention properties of red soil, but also improves its aeration. It scientifically and thoroughly solves the technical problems of "acidic, infertile, compacted" soil, poor aeration and easy root rot in the vast red soil areas of southern China, making red soil a suitable cultivation substrate for Anoectochilus roxburghii.

[0017] This invention improves survival rate and effective ingredient content by optimizing substrate formula, significantly enhancing planting efficiency and increasing local farmers' income. Furthermore, the substrate of this invention transforms local red soil resources into a high-quality cultivation substrate. Combined with a controllable facility environment, it forms a complete, low-cost, and scalable standardized production scheme for Anoectochilus roxburghii in red soil areas, laying the foundation for large-scale planting. It can further extend the industrial chain and upgrade the industrial structure, developing deep-processed Anoectochilus roxburghii products (such as lozenges, face masks, and tea) and ecotourism projects, forming a complete "planting-processing-cultural tourism" industrial chain and increasing the added value of the industry.

[0018] The technology of this invention does not require the occupation of high-quality arable land or the reclamation of forest land, avoids the destruction of the original forest stand structure, achieves a win-win situation for ecology and economy, and can provide a new land use model for red soil areas. Attached Figure Description

[0019] Figure 1 This is a statistical bar chart showing the entropy weight evaluation values ​​of eight growth indicators of *Anoectochilus roxburghii* under different red soil composite matrices in the experimental examples of this invention. Figure 2 This is a bar chart showing the statistical results of the content of roxburghii glycosides in *Anoectochilus roxburghii* grown under different red soil composite substrates in the experimental examples of this invention. Figure 3 This is a bar chart showing the statistical results of flavonoid content in *Anoectochilus roxburghii* grown under different red soil composite substrates in the experimental examples of this invention. Figure 4This is a bar chart showing the statistical results of polysaccharide content in *Anoectochilus roxburghii* grown under different red soil composite substrates in the experimental examples of this invention. Figure 5 This is a bar chart showing the survival rate of *Anoectochilus roxburghii* under different red soil composite matrices in the experimental examples of this invention. Figure 6 This is a bar chart showing the statistical results of *Anoectochilus roxburghii* plant height under different Red Ten compound substrates in the experimental examples of this invention; Figure 7 This is a bar chart showing the statistical results of root length of Anoectochilus roxburghii under different Red Ten compound substrates in the experimental examples of this invention; Figure 8 This is a bar chart showing the statistical results of the stem diameter of *Anoectochilus roxburghii* under different red soil composite substrates in the experimental examples of this invention; Figure 9 This is a bar chart showing the statistical results of the number of leaves per plant of *Anoectochilus roxburghii* under different red soil composite substrates in the experimental examples of this invention. Figure 10 This is a bar chart showing the statistical results of the maximum leaf area of ​​*Anoectochilus roxburghii* under different red soil composite substrates in the experimental examples of this invention. Figure 11 This is a bar chart showing the statistical results of the fresh weight of *Anoectochilus roxburghii* under different Red Ten compound matrices in the experimental examples of this invention; Figure 12 This is a bar chart showing the statistical results of the dry weight of Anoectochilus roxburghii under different Red Earth compound matrices in the experimental examples of this invention; Figure 13 This is a bar chart showing the statistical results of the dry weight percentage of Anoectochilus roxburghii under different Red Ten compound matrices in the experimental examples of this invention.

[0020] Note: In the statistical bar chart, no symbol indicates P>0.05, meaning there is no significant difference between the two groups; * indicates P≤0.05, meaning the difference is relatively significant; ** indicates P≤0.01, meaning the difference is significant; *** indicates P≤0.001, meaning the difference is extremely significant. Detailed Implementation

[0021] The technical solutions of the present invention will be clearly and completely described below with reference to the accompanying drawings of the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments.

[0022] Example 1 The present invention provides a red soil compound substrate suitable for planting Anoectochilus roxburghii. The compound substrate is composed of red soil, peat moss, perlite and bark mixed in a specific volume ratio. Specifically, the volume ratio of red soil: peat moss: perlite: bark is (2-4):1:1:1.

[0023] The perlite grain size distribution is 3-6 mm; Peat moss is composed of fine fibers with a size of 0-25 mm.

[0024] The bark is the dried bark of any one or more of the following trees: pine, chestnut, and sand tree. The bark is obtained after pretreatment by the following method: air-drying and crushing, and the size of the crushed bark is 10-25mm.

[0025] Peat moss is composed of fine fibers with a size of 0-10 mm.

[0026] The red soil is clean and free of pests and diseases; the red soil is treated by removing pests, sterilizing and disinfecting it before use; the removal of pests, sterilizing and disinfecting it before use is done by sun exposure, and the pH value of the red soil is in the range of 5.5-6.5, which does not require additional adjustment.

[0027] Raw material preparation: Prepare clean, pest-free red soil, peat moss, perlite, and bark.

[0028] The specific methods of cultivation and planting include the following: First, the cultivation location: inside a greenhouse or a cultivation facility with shade netting; Choose a well-drained, sheltered, and sunny gentle slope to build a single plastic greenhouse equipped with shade nets (70%-80% shading rate) and drip irrigation or micro-sprinkler irrigation system, and build elevated seedbeds 60-80cm off the ground. When cultivating, use local red soil that has been simply disinfected. The red soil and peat moss are in a semi-dry and loose state. Mix the red soil, peat moss, perlite and bark according to the specified volume ratio. Adjust the humidity to the state of "forming a ball when squeezed in the hand, and crumbling when released". The resulting red soil mixture substrate suitable for planting Anoectochilus roxburghii is then filled into seedling trays.

[0029] Transplant the hardened Anoectochilus roxburghii tissue culture seedlings into seedling trays and water them thoroughly to help them settle down. After transplanting, the environment is precisely controlled using facilities: the daily temperature inside the greenhouse is maintained at 18-25℃. Air humidity: Prefers high humidity, with relative humidity maintained between 70% and 85%; Light: Shading is required; light intensity should be controlled at 5000-10000 Lux; daily light duration should be 8-10 hours. Under these conditions, water according to the principle of "watering when dry" and apply diluted liquid fertilizer every 10-15 days; After 7 months of cultivation and management, high-quality Anoectochilus roxburghii plants can be harvested.

[0030] experimental group 1.1 Experimental group setup The red soil (CK), peat moss, perlite, bark, and vermiculite in the red soil compound substrate for planting Anoectochilus roxburghii were formulated in different volume ratios. The plants were then planted and cultivated according to the preparation method and cultivation plan in Example 1. The performance of the planted Anoectochilus roxburghii was then comprehensively evaluated.

[0031] Table 1 shows the components of the laterite composite matrix and the corresponding volume ratios of the T1-T24 groups and the control group (CK).

[0032] Each substrate (T1-T24) has three parallel replicates. After purchasing the tissue culture seedlings of *Anoectochilus roxburghii*, they are first placed indoors in a greenhouse and left to stand for 10 days, then the caps of the culture bottles are unscrewed and left to stand for 3 days, and finally, the bottles are left open to harden off for 5 days. During cultivation, 10 healthy *Anoectochilus roxburghii* tissue culture seedlings are selected from each pot and planted, ensuring even spacing between seedlings and uniform seedling condition between groups.

[0033] 2.1 Evaluation Methods for Experimental Cases This invention uses tissue-cultured Anoectochilus roxburghii seedlings as the cultivation object, which are transplanted into pure red soil (control group CK) and compound substrates containing different proportions of red soil (experimental groups T1-T24) for 7 months. During this period, environmental factors such as temperature and humidity are controlled and kept stable. The damage of Anoectochilus roxburghii seedlings is recorded to calculate the survival rate. After harvesting, the growth indicators and medicinal value indicators of Anoectochilus roxburghii are measured to evaluate the effect of compound substrates containing different proportions of red soil on the transplantation of Anoectochilus roxburghii tissue-cultured seedlings.

[0034] 2.2 The experimental design and corresponding performance analysis are as follows: (1) The effect of compound substrates containing different proportions of red soil on the survival rate of Anoectochilus roxburghii tissue culture seedlings after transplanting; (2) The effect of compound substrates containing different proportions of red soil on the growth and medicinal indicators of Anoectochilus roxburghii tissue culture seedlings after transplanting.

[0035] 2.3 Survival rate statistics of Anoectochilus roxburghii During the cultivation of Anoectochilus roxburghii, its survival status was observed and recorded regularly. Finally, the number of seedlings lost in each group was counted, and the survival rate of each group was calculated as: (final number of surviving seedlings / 30) × 100%.

[0036] 2.4 Measurement of various indicators of Anoectochilus roxburghii 2.4.1 Growth Indicators The main growth indicators of Anoectochilus roxburghii include plant height (measured with a ruler from the base to the top), number of leaves per plant, maximum leaf length and width (the largest leaf is selected and its maximum length and width are measured with a ruler), stem diameter (measured with calipers, with 1cm above the stem base as the baseline), root length (the main root system of Anoectochilus roxburghii is selected and its length from the base to the tip is measured with a ruler), fresh and dry weight per plant (weighed using an electronic balance in fresh and dry states), and dry weight percentage (dry weight of a single Anoectochilus roxburghii plant / fresh weight of a single Anoectochilus roxburghii plant × 100%).

[0037] 2.4.2 Medicinal Quality Indicators 2.4.2.1 Sample pretreatment: drying, grinding and ultrasonic extraction The foundation of medicinal quality determination lies in obtaining stable, homogeneous samples with complete component extraction. First, fresh samples of *Anoectochilus roxburghii* plants were dried at 50°C to constant weight to completely remove moisture and obtain dry matter suitable for precise calculations. Subsequently, the dried sample was pre-crushed manually and then finely ground using a high-frequency vibrating ball mill to produce a uniform and fine powder, significantly increasing the specific surface area for subsequent extraction. Finally, ultrasonic extraction was performed: the dried *Anoectochilus roxburghii* powder was ultrasonically treated with water as a solvent to prepare an aqueous extract for determining *Anoectochilus roxburghii* glycosides and polysaccharides; another portion of the dried powder was ultrasonically treated with 70% ethanol as a solvent to prepare an ethanolic extract for determining total flavonoids. Both extracts underwent centrifugation, filtration, and volume adjustment to obtain a clear and stable sample solution for testing.

[0038] 2.4.2.2 Determination scheme for Anoectochilus roxburghii glycoside content Anoectochilin, a characteristic iridoid glycoside of *Anoectochilus roxburghii*, is a core indicator for quality evaluation. This method employs high-performance liquid chromatography-evaporative light scattering (HPLC-ELSD) for its determination. A hydrophilic AQ-C18 column is used, with isocratic elution in a high-proportion aqueous methanol-water solution to achieve efficient chromatographic separation of anoectochilin. The ELSD detector is independent of the compound's UV absorption, making it particularly suitable for the detection of anoectochilin, offering high sensitivity and baseline stability. A standard curve is constructed by injecting a series of anoectochilin standard solutions. The aqueous extract of the sample is then analyzed, and the anoectochilin content in the sample is calculated based on the peak area. The results are expressed in grams per 100 grams of dry weight.

[0039] 2.4.2.3 Polysaccharide Content Determination Protocol Polysaccharides are important water-soluble active macromolecules in *Anoectochilus roxburghii*. This method uses the classic phenol-sulfuric acid colorimetric method to determine their total amount. The principle is that under the action of concentrated sulfuric acid, the polysaccharide hydrolyzes and dehydrates, producing a aldehyde derivative that reacts with phenol to form an orange-yellow compound with characteristic absorption at 490 nm. Glucose was used as a standard, and a standard curve was plotted between absorbance and sugar mass. In the determination, an appropriate amount of *Anoectochilus roxburghii* aqueous extract was reacted sequentially with phenol and concentrated sulfuric acid. After heating and color development, the absorbance was measured at 490 nm and substituted into the standard curve for calculation. The final result is expressed as glucose equivalent, i.e., the number of grams of polysaccharide contained per 100 grams of dry weight.

[0040] 2.4.2.4 Determination Scheme for Total Flavonoid Content Total flavonoids are an important class of polyphenolic active components in *Anoectochilus roxburghii*. This method uses the aluminum nitrate-sodium nitrite colorimetric method for determination. This method is based on the reaction of flavonoids with aluminum ions to form stable complexes under alkaline conditions, which exhibit strong absorption at 510 nm. Rutin was used as a standard, and a standard curve was plotted. In the sample determination, sodium nitrite, aluminum nitrate, and sodium hydroxide solutions were added sequentially to the *Anoectochilus roxburghii* alcohol extract, reacting stepwise to develop color, and then diluted to volume. The absorbance was measured at 510 nm. Substituting the absorbance value of the sample into the rutin standard curve, the total flavonoid content, expressed as "rutin equivalents," can be calculated. The result is usually expressed in milligrams per gram of dry weight.

[0041] 2.4.3 Statistical Analysis The growth and medicinal quality indicators of *Anoectochilus roxburghii* obtained in this experiment will be objectively and comprehensively evaluated and analyzed using the objective weighted entropy method. The core of this method lies in objectively calculating the weight of each indicator based on the amount of information provided by the data, thus avoiding bias caused by subjective assignment. Specifically, firstly, the raw data of all measured indicators will be standardized to eliminate the influence of dimensions and orders of magnitude. Then, the entropy value of each indicator will be calculated. The smaller the entropy value, the greater the degree of variation of the indicator, and the more information it can provide in the comprehensive evaluation; therefore, it should be assigned a greater weight. After calculating the weight of each indicator based on the entropy value, the weighted sum of each indicator in each experimental group can be obtained to obtain a comprehensive score. Finally, by comparing the comprehensive scores of all experimental groups, the optimal red soil compound substrate formula for promoting the growth and accumulation of active ingredients in *Anoectochilus roxburghii* can be scientifically and quantitatively screened.

[0042] 2.5 Growth Indicator Results Depend on Figures 2-13 The average data results of each group of Anoectochilus roxburghii under different Red Ten compound substrates include 8 morphological indicators (plant height, stem diameter, root length, leaf area, number of leaves, survival rate, fresh weight, and dry weight) and 3 medicinal indicators (polysaccharide content, flavonoid content, and Anoectochilus roxburghii glycoside content). The 11 indicators were evaluated using an entropy weight evaluation mathematical model to obtain the entropy weight evaluation value of the growth indicators of each group.

[0043] The specific calculation steps of the mathematical model for entropy weight evaluation are as follows: Given 25 sample treatments and 11 indicators measured, we obtain n=11 and m=25. Using m different irrigation and compounding substrate schemes combined with n different indicators, we can construct the evaluation matrix R=( ) m×n ,in Let represent the value of the j-th index under the i-th compound matrix treatment. The optimal value of each column in the matrix is ​​denoted as . And obtain standardized calculations based on the nature of the indicators. .

[0044] Calculate the proportion The formula is: =

[0045] Calculate information entropy The formula is: = - ln , j=1, 2, 3,……, n.

[0046] Calculate the entropy value The formula is: = , j=1, 2, 3,……, n.

[0047] Calculate objective weights and final indicator weights : First, calculate the objective weights. = ,in .certainly, Decision-maker's subjective weight It can be combined with objective weights to derive new indicator weights. = .

[0048] Calculate the entropy weight evaluation value The formula is: = , where i = 1, 2, 3, ..., m.

[0049] like Figure 1 The bar chart shown here, which evaluates the entropy weight scores for the eight growth morphology indicators mentioned above, visually demonstrates the comprehensive performance of *Anoectochilus roxburghii* growth indicators under different substrate ratios. Statistical results show that, compared to the pure red soil control group (CK), most of the compound substrate groups in this invention exhibit superior growth potential. Specifically, groups T21, T13, T19, T15, T10, T23, and T24 show relatively high evaluation values ​​for their growth indicators. This indicates that under this compound system, the physical pore structure and nutrient supply of the substrate achieve a good balance, which is beneficial for the root development, morphological development, and rapid accumulation of overall biomass in *Anoectochilus roxburghii* tissue culture seedlings.

[0050] 2.6 Results of Anoectochilus roxburghii glycoside index Anoectochilin is a core specific indicator for evaluating the medicinal quality of Anoectochilus roxburghii. For example... Figure 2Data shows that under the intervention of the compound matrix of this invention, group T10 showed an advantage in this indicator, followed by groups T23 and T24. This indicates that the specific red soil light matrix improvement scheme can provide an excellent physicochemical induction environment for the secondary metabolic pathway of Anoectochilus roxburghii, and significantly improve the yield of the core active ingredients.

[0051] 2.7 Flavonoid Index Results The evaluation results of total flavonoid content showed some fluctuation among different compound groups. For example... Figure 3 As shown, group T24 exhibited the highest level, indicating that when red soil is combined with perlite, which has good air permeability and water retention properties, and peat soil and bark rich in organic matter, it can effectively activate the polyphenol metabolic pathway in Anoectochilus roxburghii and promote the enrichment of antioxidant active substances.

[0052] 2.8 Polysaccharide Index Results The data distribution of polysaccharide indicators further validates the necessity of matrix improvement. For example... Figure 4 As shown, in the experimental groups, the polysaccharide evaluation scores of groups T21 and T13 were significantly higher than those of other groups. The optimized compound substrate improved the rhizosphere microenvironment, promoted the efficient absorption of water and nutrients by plants, and thus converted them into abundant polysaccharides.

[0053] 2.9 Comprehensive data on medicinal indicators, the statistical data results are shown in Table 2 below.

[0054] Table 2. Comprehensive data on the medicinal properties of Anoectochilus roxburghii under different red clay composite matrices.

[0055] In the polysaccharide determination experiment, after grinding the dried *Anoectochilus roxburghii* and sonicating the cells to obtain the contents, six samples in groups T13 and T21 were not adequately filtered, resulting in an excessively dark color of the extract. The values ​​detected by the microplate reader were significantly higher than those of other groups. This does not necessarily reflect their actual polysaccharide content. Since this occurred in all three replicates of each group, it was impossible to determine the true polysaccharide content of this treatment by eliminating outliers. The data from other groups were accurate because: the detection method itself was validated with standards; the results from the three samples within the same group were consistent, demonstrating good repeatability; and the overall data conformed to the expected experimental pattern.

[0056] 2.10 The entropy weight evaluation ranking of all indicators is shown in Table 3 below.

[0057] Table 3. Objective weights (entropy method) of all indicators of *Anoectochilus roxburghii* under different red soil composite matrices. Entropy weight evaluation ranking table.

[0058] The final comprehensive score, obtained by objectively weighting all growth and medicinal indicators using the entropy weight method, shows that the overall planting benefits of the compound substrate of this invention comprehensively surpass those of traditional pure red soil. Group T24 (red soil: perlite: perlite: bark = 3:1:1:1) ranked first with its outstanding performance in all individual indicators, demonstrating extremely high potential for large-scale promotion in ensuring high survival rates and stable medicinal quality. These detailed data provide strong scientific evidence for the efficient agricultural utilization of red soil resources.

[0059] In this invention's formula, red soil accounts for 60% of the volume, which, proportionally, is more than half, significantly reducing production costs compared to traditional methods that rely entirely on purchased lightweight substrates like peat moss. This formula retains the advantages of red soil, which is rich in iron and aluminum oxides and trace elements, enhancing the root system's stability and truly transforming "inferior soil" into "golden soil." The addition of peat moss supplements the high-quality humus lacking in red soil, buffering pH levels and improving the substrate's aggregate structure. Perlite, as a porous inorganic material, completely overcomes the physical limitation of red soil's tendency to compact, greatly improving the substrate's porosity and drainage / aeration capacity, ensuring that the delicate, fleshy roots of the *Anoectochilus roxburghii* can breathe freely. The bark perfectly simulates the evergreen broad-leaved forest understory habitat of the *Anoectochilus roxburghii* in its wild state. Its slow decomposition process not only maintains soil looseness for a long time but also provides sustained nutrient support as a slow-release fertilizer.

[0060] Based on the above experimental data, the red soil compound substrate of this invention, formulated in a specific ratio and suitable for planting Anoectochilus roxburghii, can maximize the activation of the secondary metabolic pathways of Anoectochilus roxburghii while ensuring an extremely high transplant survival rate, and significantly promote the biosynthesis of core effective components such as Anoectochilus roxburghii glycosides, polysaccharides, and flavonoids.

[0061] In summary, the red soil compound substrate of this invention, suitable for the cultivation of Anoectochilus roxburghii, is easy to operate, uses readily available raw materials, and is extremely well-suited to the climatic characteristics of large red soil distribution areas such as Yunnan. It not only provides a low-cost substrate solution for the large-scale, standardized cultivation of Anoectochilus roxburghii, but also lays a solid technical foundation for the subsequent development of deep-processed products of Anoectochilus roxburghii, the creation of a complete "planting-processing-cultural tourism" industrial chain, and the contribution to rural revitalization.

[0062] The above description is only a preferred embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. Any equivalent substitutions or modifications made by those skilled in the art within the scope of the technology disclosed in the present invention, based on the technical solution and inventive concept of the present invention, should be covered within the scope of protection of the present invention.

Claims

1. A red soil composite substrate suitable for planting Anoectochilus roxburghii, characterized in that, The compound matrix is ​​composed of red clay, peat moss, perlite and bark mixed in a specific volume ratio. Specifically, the volume ratio is: red clay: peat moss: perlite: bark = (2-4):1:1:1; the perlite has a particle size distribution of 3-6 mm; the peat moss is fine fiber with a size of 0-25 mm.

2. The red soil composite substrate suitable for planting Anoectochilus roxburghii according to claim 1, characterized in that, The volume ratio of each component in this compound matrix is: red soil: peat soil: perlite: bark = 3:1:1:

1.

3. The red soil composite substrate suitable for planting Anoectochilus roxburghii according to claim 2, characterized in that, The bark is the dried bark of any one or more of the following trees: pine, chestnut, and sand tree. The bark is obtained by pre-treatment as follows: after air drying and crushing, the size of the crushed bark is 10-25mm.

4. The red soil composite substrate suitable for planting Anoectochilus roxburghii according to claim 2, characterized in that, The peat soil is made of fine fibers with a size of 0-10 mm.

5. The red soil composite substrate suitable for planting Anoectochilus roxburghii according to claim 2, characterized in that, The red soil is clean and free from pests and diseases; the red soil is treated by removing pests, sterilizing and disinfecting it before use; the removal of pests, sterilizing and disinfecting it before use is done by sun exposure; the pH value of the red soil is in the range of 5.5-6.5 and does not require additional adjustment.

6. A red soil composite substrate suitable for planting *Anoectochilus roxburghii* according to any one of claims 1-5, characterized in that, The configuration methods include the following: Raw material preparation: Prepare clean, pest-free red soil, peat moss, perlite and bark. The red soil and peat moss should be in a semi-dry and loose state. Preparation of compound substrate: According to the volume ratio, accurately proportion and mix the red soil, peat moss, perlite and bark. After mixing them evenly, the substrate moisture content should be such that it can be formed into a ball when squeezed in the hand, but crumbles when released. This will give you a red soil compound substrate suitable for planting Anoectochilus roxburghii.

7. The application of a red soil compound substrate suitable for planting *Anoectochilus roxburghii* according to any one of claims 1-6, characterized in that, Red soil improvement for planting Anoectochilus roxburghii.

8. The application of the red soil compound substrate suitable for planting Anoectochilus roxburghii according to claim 7, characterized in that, The indoor seedling cultivation method includes the following steps: S1. Selecting the cultivation site and environment for Anoectochilus roxburghii. Temperature: Golden Thread Orchid prefers cool temperatures, with the average daily temperature maintained between 18-25℃; Air humidity: Prefers high humidity, with relative humidity maintained between 70% and 85%; Light: Shading is required; light intensity should be controlled at 5000-10000 Lux; daily light duration should be 8-10 hours. Cultivation location: Inside greenhouses or cultivation facilities with shade nets; S3. Transplanting of Anoectochilus roxburghii tissue culture seedlings: Carefully transplant healthy and well-maintained Anoectochilus roxburghii tissue culture seedlings into a prepared red soil composite substrate for cultivation and post-cultivation management.

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