A method for constructing a characteristic spectrum of a herba commelinae and a medicine preparation and application thereof

Abstract

This invention belongs to the field of quality testing technology for traditional Chinese medicine preparations, specifically relating to a characteristic chromatogram of Commelina communis and its pharmaceutical preparations, and further disclosing the method for constructing the characteristic chromatogram and the quality testing method for Commelina communis and its pharmaceutical preparations. Using Commelina communis medicinal material and its standard decoction as the research object, this invention identified 14 common characteristic peaks and identified 5 components: Peak 1: hypoxanthine; Peak 3: adenosine; Peak 4: guanosine; Peak 8: L-tryptophan; Peak 14: isopropanol. Under these chromatographic conditions, the established characteristic chromatogram, obtained through the detection of Commelina communis medicinal material and its related preparations, shows significant characteristic components, good separation effect, and is simple to use. It can easily and quickly identify the types and quantities of chemical components contained in Commelina communis medicinal material and its related preparations (standard decoction freeze-dried powder, formula granules), providing a more comprehensive, objective, and rapid quality evaluation method for Commelina communis medicinal material and its pharmaceutical preparations.

Description

Technical Field

[0001] This invention belongs to the field of quality testing technology for traditional Chinese medicine preparations, specifically relating to a characteristic spectrum of Commelina communis and its medicinal preparations, and further disclosing a method for constructing the characteristic spectrum, as well as a method for quality testing of Commelina communis and its medicinal preparations. Background Technology

[0002] The raw material for Commelina communis formula granules is the dried aerial parts of Commelina communis L., a plant of the Commelinaceae family. Commelina communis is a Chinese herbal medicine listed in the current pharmacopoeia. It is sweet, bland, and cold in nature, and enters the lung, stomach, and small intestine meridians. It has the effects of clearing heat and purging fire, detoxifying, and promoting diuresis and reducing swelling. It can be used for colds with fever, fever with thirst, sore throat, edema with scanty urine, painful urination due to heat, and carbuncles and boils.

[0003] The main chemical components of Commelina communis are flavonoids and phenolic acids, and the literature also describes the chemical components in Commelina communis. However, on the one hand, by determining or identifying the content of the above-mentioned single components, the quality of Commelina communis and its preparations (such as formulation granules) cannot be detected and controlled as a whole; on the other hand, determining the content of a single component in conjunction with the identification of other components in Commelina communis and its preparations is time-consuming and labor-intensive, and difficult to widely apply in production practice.

[0004] For example, Chinese patent CN117929590A discloses a method for constructing characteristic chromatograms of Commelina communis and its preparations. This method uses high-performance liquid chromatography (HPLC) with octadecylsilane-bonded silica gel as the packing material, methanol as mobile phase A, and water or an acidic aqueous solution as mobile phase B. An optimized gradient elution program is used to obtain nine common characteristic peaks: peak 2 (vanillic acid); peak 6 (hymenoside); peak 7 (S peak) (isohymenoside); peak 8 (viticin); and peak 9 (rutin). This method effectively separates multiple common characteristic peaks, resulting in uniform peak height or area. However, the characteristic chromatograms obtained using this method mainly show moderately polar components (5%–30% organic phase), and the resulting chromatograms have unstable baselines and numerous extraneous peaks, often making accurate identification of characteristic peaks difficult. Furthermore, the information presented is insufficient, and the elution gradient only shows the nine ultimately identified characteristic peaks.

[0005] Therefore, establishing HPLC characteristic chromatograms of Dayflower herb and its pharmaceutical preparations as a preliminary method for quality control of Dayflower herb and downstream products is an effective quality control approach. Summary of the Invention

[0006] Therefore, the technical problem to be solved by the present invention is to provide a method for constructing a characteristic spectrum of Commelina communis herb and pharmaceutical preparations, so as to comprehensively reflect the intrinsic quality and medication safety of Commelina communis and its pharmaceutical preparations.

[0007] The second technical problem to be solved by the present invention is to provide a quality testing method for Dayflower herb and pharmaceutical preparations.

[0008] To solve the above-mentioned technical problems, the present invention provides a method for constructing characteristic chromatograms of Commelina communis medicinal material and pharmaceutical preparations, including the step of performing high performance liquid chromatography detection on the test solution of Commelina communis medicinal material and / or Commelina communis pharmaceutical preparations.

[0009] Chromatographic conditions included: using octadecylsilane-bonded silica gel as the stationary phase, acetonitrile as mobile phase A, and 0.08%–0.12% formic acid solution as mobile phase B, with gradient elution performed according to the following procedure:

[0010] 0-5 min, A:B ratio was 2%:98%;

[0011] 5-15 min, A:B changed from 2% to 7%; 98% to 93%;

[0012] 15-25 min, A:B changed from 7% to 12%; 93%:B changed from 88%;

[0013] 25-35 min, A:B ratio is 12%:88%;

[0014] 35-40 min, A:B ratio changed from 12% to 14%; 88%:B ratio changed from 88% to 86%.

[0015] 40-60 min, A:B ratio was 14%:86%.

[0016] Preferably, in the chromatographic conditions, acetonitrile is used as mobile phase A and 0.10% formic acid solution is used as mobile phase B.

[0017] Specifically, in the method for constructing the characteristic chromatograms of the herbal medicine and pharmaceutical preparation of *Commelina communis*, the chromatographic conditions in the high-performance liquid chromatography detection step further include: a flow rate of 0.9–1.1 ml / min, a column temperature of 23–27 °C, and a detection wavelength of 260 nm.

[0018] Preferably, the chromatographic conditions are as follows: flow rate 1.0 ml / min, column temperature 25°C, and detection wavelength 260 nm.

[0019] Specifically, the method for constructing the characteristic spectrum of the herbal medicine and pharmaceutical preparation of *Commelina communis* includes the following method for preparing the test sample solution: accurately adding the test sample to a first solvent for mixing, adding a second solvent for extraction, evaporating the combined extracts to dryness, adding a third solvent to redissolve the residue, and filtering after ultrasonic treatment to obtain the final product.

[0020] Specifically, the method for constructing the characteristic spectrum of the herbal medicine and pharmaceutical preparation of Commelina communis includes the step of preparing a reference solution of the control herbal medicine, which specifically includes: taking Commelina communis reference herbal medicine, precisely adding it to the first solvent and mixing it, heating and refluxing it, collecting the filtrate and evaporating it to dryness, adding the residue to the first solvent and mixing it, adding the second solvent and extracting it, combining the extracts and evaporating them to dryness, adding the residue to the third solvent to redissolve it, and filtering it after ultrasonic treatment to obtain the final product.

[0021] Specifically, the method for constructing the characteristic chromatogram of the herbal medicine and pharmaceutical preparation of *Commelina communis* further includes the step of preparing a reference solution and the step of constructing the characteristic chromatogram of the reference solution based on the high performance liquid chromatography method;

[0022] The reference standards include hypoxanthine, adenosine, guanosine, L-tryptophan and / or isohypoglycine;

[0023] The preparation method of the reference solution includes: precisely adding the reference standard to a third solvent and mixing.

[0024] Specifically, the method for constructing the characteristic spectrum of the herbal medicine *Commelina communis* and its pharmaceutical preparations is as follows:

[0025] The first solvent includes water; and / or,

[0026] The second solvent includes water-saturated n-butanol; and / or,

[0027] The third solvent includes methanol; and / or,

[0028] The ultrasonic treatment step has a power of 450-550W and a frequency of 35-40kHz.

[0029] Specifically, the method for constructing the characteristic spectrum of the herbal medicine and pharmaceutical preparation of Commelina communis includes Commelina communis formula granules, Commelina communis slices, or Commelina communis standard decoction.

[0030] This invention also discloses a characteristic spectrum and / or a control characteristic spectrum of the herbal medicine and pharmaceutical preparation of *Commelina communis*, wherein the characteristic spectrum or control characteristic spectrum of the herbal medicine and pharmaceutical preparation is constructed by the method described above;

[0031] Preferably, the characteristic spectrum has 14 characteristic peaks, wherein peaks 1, 3, 4, 8, and 14 correspond to the retention times of the corresponding reference peaks; the peak corresponding to the guanosine reference peak is peak S1, and the relative retention times of peaks 1-10 with peak S1 are calculated; the peak corresponding to the isopropanol reference peak is peak S2, and the relative retention times of peaks 11-13 with peak S2 are calculated. The relative retention times should be within ±10% of the specified values. The specified values ​​for each characteristic peak are: peak 1: 0.48, peak 2: 0.80, peak 3: 0.86, peak 5: 1.08, peak 6: 1.29, peak 7: 1.61, peak 8: 1.74, peak 9: 2.15, peak 10: 2.46, peak 11: 0.79, peak 12: 0.84, and peak 13: 0.93.

[0032] The present invention also discloses the method for constructing the characteristic spectrum of the said duckweed herb and pharmaceutical preparation and / or the application of the characteristic spectrum and / or control characteristic spectrum of the said duckweed herb and pharmaceutical preparation in the field of quality testing of duckweed herb and pharmaceutical preparation.

[0033] The present invention also discloses a quality detection method for the herbal medicine of Dayflower and its pharmaceutical preparation, including the steps of constructing the characteristic spectrum and the control characteristic spectrum according to the method, and the step of comparing the characteristic spectrum with the control characteristic spectrum.

[0034] This invention focuses on the medicinal material *Commelina communis* and its standard decoction. Fourteen common characteristic peaks were identified, and five components were identified: Peak 1: hypoxanthine; Peak 3: adenosine; Peak 4: guanosine; Peak 8: L-tryptophan; Peak 14: isopropanol. Under these chromatographic conditions, the established characteristic chromatograms for the detection of *Commelina communis* and its related preparations show significant characteristic components, good separation effect, and are simple to use. Furthermore, it allows for a simple and rapid identification of the types and quantities of chemical components contained in *Commelina communis* and its related preparations (standard decoction freeze-dried powder, formula granules). This provides a more comprehensive, objective, and rapid quality evaluation method for *Commelina communis* and its pharmaceutical preparations, and is of great significance for comprehensive quality testing and overall quality control.

[0035] This invention employs high-performance liquid chromatography (HPLC) to establish characteristic chromatograms of Commelina communis herb and related preparations. By using a low-proportion organic phase (2%–14% organic phase) for elution and setting gradient times, while simultaneously purifying the sample using liquid-liquid extraction, the resulting characteristic chromatograms exhibit stable baselines, significant characteristic components, and good separation. Fourteen common characteristic peaks were identified, along with five highly polar characteristic components: hypoxanthine, adenosine, guanosine, L-tryptophan, and isopropanol. This increases the variety and quantity of chemical components in Commelina communis and related preparations, making the characteristic peaks in the chromatograms easier to identify and providing more accurate and reliable results.

[0036] This invention provides a process for optimizing chromatographic conditions for characteristic chromatograms, a process for methodological validation, and test results from multiple batches of medicinal slices and lyophilized powders of standard decoctions. It confirms 14 common characteristic peaks and identifies 5 components: peak 1: hypoxanthine; peak 3: adenosine; peak 4: guanosine; peak 8: L-tryptophan; peak 14: isopropanol. These 5 characteristic components provide a rapid and reliable detection method for the characteristic chromatographic identification of Commelina communis herb and related preparations.

[0037] The invention establishes a characteristic spectrum, taking processed medicinal materials and related preparations as the research object, confirming 15 common characteristic peaks and identifying 5 characteristic components. The results shown by the spectrum are easy to judge, providing a rapid and reliable detection method for the characteristic spectrum identification of duckweed medicinal materials and their pharmaceutical preparations. Attached Figure Description

[0038] To make the content of this invention easier to understand, the invention will be further described in detail below with reference to specific embodiments and accompanying drawings, wherein...

[0039] Figure 1 The results of the full gradient investigation with different solvents in Example 1;

[0040] Figure 2 The results of the elution gradient optimization investigation in Example 1;

[0041] Figure 3 The results are from the investigation under different mobile phase systems in Example 1;

[0042] Figure 4 The results are from different detection wavelengths in Example 1;

[0043] Figure 5 The results are from different chromatographic columns used in Example 1.

[0044] Figure 6 The results of the investigation under different extraction solvents in Example 2;

[0045] Figure 7 The results of the investigation under different purification methods - solid-phase column packing in Example 2;

[0046] Figure 8 The results of the investigation of different liquid phase extraction solvents in Example 2;

[0047] Figure 9 The chromatograms are of the following: the test sample of the standard decoction of Commelina communis slices in Example 3; the chromatograms of hypoxanthine reference standard; the chromatograms of adenosine reference standard; the chromatograms of guanosine reference standard; the chromatograms of L-tryptophan reference standard; and the chromatograms of isopropanol reference standard.

[0048] Figure 10 This is the verification result of the chromatographic conditions and system suitability in Example 4;

[0049] Figure 11 The results of the precision test in Example 4;

[0050] Figure 12 The results are from the repeatability test in Example 4;

[0051] Figure 13 The results of the intermediate precision (different operators) assessment for Example 4;

[0052] Figure 14 The results of the stability test in Example 4;

[0053] Figure 15 The results of durability testing at different flow rates in Example 4;

[0054] Figure 16 The results of the durability test at different column temperatures in Example 4;

[0055] Figure 17 This is the comparative feature map in Example 4;

[0056] Figure 18 The characteristic chromatograms of the three batches of Commelina communis standard decoction in Example 5 are shown.

[0057] Figure 19 This is the verification result of the chromatographic conditions and system suitability in Example 6.

[0058] Figure 20 This is a comparative feature map in Example 6. Detailed Implementation

[0059] The instruments, equipment, and reagents involved in the following embodiments of the present invention are shown in Tables 1-2 below.

[0060] Table 1. Statistics on Instruments and Equipment

[0061] Instrument Name model factory One ten-thousandth balance ME104 Mettler Toledo International Trading (Shanghai) Co., Ltd. One-millionth balance XPR2 Mettler Toledo International Trading (Shanghai) Co., Ltd. CNC ultrasonic instrument KQ-500DE Kunshan Ultrasonic Instruments Co., Ltd. High Performance Liquid Chromatography e2695 Waters Corporation High Performance Liquid Chromatography Thermo U3000 Thermo Fisher Scientific Chromatographic column (2.5x250mm, 5μm) JADE-PAK ODS-AQ Taiwei Technology Company Chromatographic column (2.5x250mm, 5μm) Welch Ultimate AQ-C18 Moonrise Technology Company Chromatographic column (2.5x250mm, 5μm) Atlantis T3 Waters Corporation

[0062] Table 2. Statistics of Reagents and Test Items

[0063] reagents and reagents Grade / Purity batch number factory hypoxanthine 99.4％ 140661-202005 China National Institutes for Food and Drug Control adenosine 99.4％ 110879-202204 China National Institutes for Food and Drug Control Guanosine 88.6％ 111977-202202 China National Institutes for Food and Drug Control L-Tryptophan 100.0％ 140686-202205 China National Institutes for Food and Drug Control Isorhizonin 94.0％ 111974-201401 China National Institutes for Food and Drug Control methanol Chromatographic purity F24013202 Fisher Acetonitrile Chromatographic purity F23N5G202 Fisher methanol Analytical Pure 20231201-11 Guangzhou Chemical Reagent Factory Formic acid Analytical Pure 20230429 Guangdong Guanghua Technology Co., Ltd. Ethyl acetate Analytical Pure 20230823 Guangdong Guanghua Technology Co., Ltd. n-Butanol Analytical Pure 20230519 Guangdong Guanghua Technology Co., Ltd.

[0064] Example 1

[0065] This embodiment investigates the optimization of chromatographic conditions.

[0066] (1) Investigation of the full gradient of the mobile phase

[0067] Mobile phase conditions 1: Using acetonitrile as mobile phase A and water as mobile phase B, gradient elution was performed according to the specifications in Table 3. The chromatogram is shown in the appendix. Figure 1 (a)

[0068] Mobile phase condition 2: Using methanol as mobile phase A and water as mobile phase B, gradient elution was performed according to the specifications in Table 3. The chromatogram is attached. Figure 1 (b)

[0069] Table 3 Gradient Procedure

[0070] program Time (minutes) Mobile phase A (%) Mobile phase B (%) Mobile phase 1 0～60 2→95 98→5 Mobile phase 2 0～60 2→95 98→5

[0071] The results showed that in the full gradient investigation of acetonitrile and methanol, the chromatographic peaks were numerous and disordered. When eluted with acetonitrile, the chromatographic peaks were more concentrated. Therefore, we tried to use acetonitrile for further optimization to improve the separation between them and make their elution time more uniform.

[0072] (2) Elution gradient optimization

[0073] As can be seen from the above acetonitrile full gradient investigation, the chromatographic peaks are mainly distributed in the first 25 minutes, concentrated around the retention times of 5 minutes and 15 minutes. The corresponding elution gradient ratios were then developed.

[0074] Mobile phase conditions: Acetonitrile was used as mobile phase A, and water was used as mobile phase B. Gradient elution was performed according to the procedure specified in Table 4. The chromatograms obtained under gradient optimization procedures 1-5 are shown in the appendix. Figure 2 As shown in (a)-(e).

[0075] Table 4 Gradient elution program

[0076]

[0077] (3) Optimization of mobile phase screening

[0078] Under the current mobile phase conditions, there are a large number of peaks in the first 15 minutes. The retention time of the reference substance isopropargyl is about 30 minutes. After 35 minutes, the baseline is not smooth and there are many impurity peaks. Therefore, the chromatographic conditions are further optimized. The elution gradient organic phase in the first 15 minutes can be slowed down to increase the peak resolution. The gradient increase in the second half can also be slowed down slightly to improve the baseline smoothness. The effect of the water / formic acid / acetic acid / phosphoric acid aqueous phase system on the chromatographic peak separation is also investigated.

[0079] In this embodiment, the chromatograms of mobile phase 1 (aqueous phase without acid), mobile phase 2 (aqueous phase with 0.1% acetic acid), mobile phase 3 (aqueous phase with 0.1% phosphoric acid), and mobile phase 4 (aqueous phase with 0.1% formic acid) are shown in the appendix. Figure 3 As shown in (a)-(d).

[0080] (4) Detection wavelength selection

[0081] This embodiment examines the chromatographic peak performance at different wavelengths, selecting the wavelength with the strongest absorption. Absorption spectra at 220, 230, 240, 250, 260, 270, 280, 290, and 300 nm are shown in the attached figures. Figure 4 As shown in (a)-(i).

[0082] The results show that the absorption of each chromatographic peak is the largest at a wavelength of 260 nm, so the absorption wavelength is tentatively set at 260 nm.

[0083] (5) Chromatographic column investigation

[0084] Under the aforementioned mobile phase gradient conditions, the effects of JADE-PAK ODS-AQ, Welch Ultimate AQ-C18, and Atlantis T3 columns on this method were investigated, and the results are shown in the appendix. Figure 5 As shown in (a)-(c).

[0085] The results showed that the Atlantis T3 column produced more separated chromatographic peaks with a more uniform time distribution, so this method is tentatively recommended for this column model.

[0086] Example 2

[0087] This embodiment investigates the optimization of the preparation method of the test solution.

[0088] (1) Investigation of different extraction solvents

[0089] Take approximately 0.5g of freeze-dried Commelina communis powder, accurately weigh it, and place it in a stoppered conical flask. Accurately add 10% methanol, 20% methanol, 40% methanol, 50% methanol, 60% methanol, 80% methanol, and 20mL of methanol, respectively. Seal the flask tightly, weigh it, and sonicate it (power 250W, frequency 40kHz) for 30 minutes. Let it cool, weigh it again, and make up the lost weight with methanol. Shake well, filter it, and collect the filtrate to obtain the final product.

[0090] Chromatograms for the above extraction solvent systems were examined separately, and are shown in the appendix. Figure 6 As shown in (a)-(g), the chromatogram of isopropargyl glycoside reference standard is attached. Figure 6 As shown in (h).

[0091] The results showed that when 10% methanol and methanol were used as extraction solvents, there was almost no chromatographic peak response, indicating that the components could not be extracted. When 20% methanol, 40% methanol, 50% methanol, 60% methanol and 80% methanol were used as extraction solvents, there were chromatographic peak responses, but the baseline noise was large and there were many impurity peaks. Therefore, the test sample needs to be further purified.

[0092] (2) Investigation of different purification methods and solid-phase column packing

[0093] Sample Preparation 1: Accurately weigh 0.5g of freeze-dried Commelina communis powder, add 5mL of water, sonicate to dissolve, and pass through a neutral alumina solid-phase extraction column. The solution was eluted sequentially with ethyl acetate, ethyl acetate-methanol (2:1), ethyl acetate-methanol (1:1), ethyl acetate-methanol (1:2), methanol, 50% methanol, and 25% methanol (30 ml each). The eluates were collected, filtered, and used as the test solution. Chromatograms of the above solid-phase column packing systems were examined, and are shown in the appendices. Figure 7 As shown in (a)-(g).

[0094] Sample Preparation 2: Accurately weigh 0.5 g of lyophilized Commelina communis powder, add 5 mL of water, weigh again, and sonicate to dissolve. Pass the solution through a D101 type macroporous adsorption resin (1 cm inner diameter, 20 cm height; wet packing). Elute sequentially with water, 25% methanol, 50% methanol, 70% methanol, and 30 mL of methanol. Filter and collect the eluents separately, filtering each eluent to obtain the test solution. Chromatograms of the above solid-phase column packing systems are shown in the appendix. Figure 7 As shown in (h)-(l).

[0095] The chromatogram of isoharmonin reference standard is attached. Figure 7 As shown in the middle (m).

[0096] The results showed that the chromatograms of the test samples prepared by neutral alumina and macroporous resin showed increased separation and response of chromatographic peaks compared with those obtained by direct methanol-water extraction. However, the chromatographic peaks were mainly concentrated in the first 20 minutes, and isopropargyl glycoside, the main representative component of Commelina communis, was severely lost. Therefore, the liquid-liquid extraction method was further investigated.

[0097] (3) Investigation of liquid-liquid extraction solvents

[0098] Preparation of the test solution: Take about 0.5g of freeze-dried Commelina communis powder, add 10mL of water to dissolve it, extract with 20mL of water-saturated n-butanol and ethyl acetate respectively, extract three times, combine the extracts, evaporate to dryness, add 5mL of 50% methanol and methanol to redissolve the residue respectively, filter, and take the filtrate to obtain the test solution.

[0099] In this embodiment, the results of the liquid-liquid extraction systems 1 (ethyl acetate extraction-methanol redissolution), 2 (ethyl acetate extraction-50% methanol redissolution), 3 (water-saturated n-butanol extraction-methanol redissolution), and 4 (water-saturated n-butanol extraction-50% methanol redissolution) are shown in the appendix. Figure 8 As shown in (a)-(d).

[0100] The results showed that the number of peaks in the samples extracted with water-saturated n-butanol was significantly greater than that in the samples extracted with ethyl acetate, and both were superior to those purified by neutral alumina and macroporous resin. Furthermore, the reconstitution solvent had little effect on the chromatographic peaks. Therefore, it is tentatively decided to use water-saturated n-butanol extraction and methanol reconstitution to prepare the samples, further optimize the elution gradient, and increase the resolution between the peaks.

[0101] Example 3

[0102] Based on the investigation in Examples 1-2 above, the method for determining the characteristic spectrum of Commelina communis in this example is tentatively set as follows: using octadecylsilane-bonded silica gel as the packing material (column length 250 mm, inner diameter 4.6 mm, particle size 5 μm), acetonitrile as mobile phase A, and 0.1% formic acid as mobile phase B, gradient elution is performed according to the specifications in Table 5; the flow rate is 1.0 ml per minute, the column temperature is 25 °C, and the detection wavelength is 260 nm.

[0103] Table 5 Elution Procedure

[0104] Time (min) Mobile phase A (%) Mobile phase B (%) 0～5 2 98 5～15 2～7 98～93 15～25 7～12 93～88 25～35 12 88 35～40 12～14 88～86 40～60 14 86

[0105] Preparation of reference solutions: Accurately weigh 1g of Commelina communis reference material, place it in a stoppered conical flask, add 40mL of water, heat under reflux for 45 minutes, filter, evaporate the filtrate to dryness, dissolve the residue in 10mL of water, extract with 20mL of water-saturated n-butanol, extract three times, combine the n-butanol extracts, evaporate to dryness, add 5mL of methanol to the residue, sonicate (500W power, 37kHz frequency) for 10 minutes, filter, and use the filtrate as the reference solution. Separately, accurately weigh appropriate amounts of hypoxanthine, adenosine, guanosine, L-tryptophan, and isohypogonin reference standards, and dissolve them in methanol to prepare solutions containing 50μg per mL, as reference solutions.

[0106] Preparation of the test solution: Take 0.5g of the powder, add 10mL of water to dissolve it, extract with 20mL of water-saturated n-butanol, extract 3 times, combine the n-butanol extracts, evaporate to dryness, add 5mL of methanol to the residue, sonicate (power 500W, frequency 37kHz) for 10 minutes, filter, and take the filtrate to obtain the test solution.

[0107] Determination method: Accurately pipette 10 μl of the reference solution and the test solution into the liquid chromatograph and determine the result.

[0108] In this embodiment, the chromatogram of the standard decoction of Commelina communis slices is attached. Figure 9 As shown in (a), the chromatograms of hypoxanthine reference standard, adenosine reference standard, guanosine reference standard, L-tryptophan reference standard, and isopropanol reference standard are respectively attached. Figure 9 As shown in (b)-(f).

[0109] The system suitability parameters for adenosine, guanosine, L-tryptophan, and isopropanol reference standards are shown in Table 6 below, and the system suitability parameters for the test samples are shown in Table 7 below.

[0110] Table 6 System Applicability Parameters

[0111] name Retention time area high width Resolution Symmetry factor USP Theoretical Plates adenosine 11.983 993907 68596 48.3 - 0.861 15320.02 Guanosine 14.215 664787 60596 29.4 - 0.79 36239.60 L-Tryptophan 24.187 433859 37919 41.6 - 1.17 99979.32 Isorhizonin 57.571 619625 19714 86.6 - 0.945 76864.06

[0112] Table 7 System Suitability Parameters for Test Samples

[0113]

[0114] Selection of S peak in characteristic spectrum

[0115] The selection criteria for reference peak S: Based on the peak identification and selection results in the characteristic peaks of the standard decoction (lyophilized powder) of Commelina communis, the readily available chromatographic peaks of the reference standard are hypoxanthine, adenosine, guanosine, L-tryptophan, and isoharonidine. Among them, peak 4 is guanosine and peak 14 is isoharonidine. The elution time of the entire chromatogram is relatively long, so peak 4 is taken as peak S1, and the relative retention times of characteristic peaks 1-10 are calculated. Peak 14 is taken as peak S2, and the relative retention times of characteristic peaks 11-13 are calculated.

[0116] The chromatogram of the test sample should show 14 characteristic peaks, which should correspond to the retention times of the 14 characteristic peaks in the chromatogram of the reference medicinal material. Among them, peaks 1, 3, 4, 8, and 14 should correspond to the retention times of the corresponding reference material peaks. The peak corresponding to the guanosine reference peak is peak S1, and the relative retention times of peaks 1 to 10 with peak S1 should be calculated. The peak corresponding to the isopropanol reference peak is peak S2, and the relative retention times of characteristic peaks 11 to 13 should be calculated. The relative retention times should be within ±10% of the specified values, which are: 0.48 (peak 1), 0.80 (peak 2), 0.86 (peak 3), 1.08 (peak 5), 1.29 (peak 6), 1.61 (peak 7), 1.74 (peak 8), 2.15 (peak 9), 2.46 (peak 10), 0.79 (peak 11), 0.84 (peak 12), and 0.93 (peak 13).

[0117] Chromatographic peak identification and reference standard localization of standard decoction of Commelina communis slices

[0118] The *Commelina communis* sample was analyzed using ultra-high performance liquid chromatography-high resolution mass spectrometry (UPLC-Q-TOF / MS). Based on the multi-stage mass spectrometry information of the sample and in conjunction with relevant literature, the target peaks were identified. Peak 1 was consistent with the elution position of hypoxanthine, peak 3 with the elution position of adenosine, peak 5 with the elution position of guanosine, and peak 10 with the elution position of L-tryptophan. The results are shown in Table 8 below.

[0119] Table 8. Identification results of target components in Commelina communis samples.

[0120]

[0121] Example 4

[0122] This embodiment investigates the methodology for preparing standard decoctions of Commelina communis slices.

[0123] (1) Validation of chromatographic conditions and system suitability

[0124] The test solution and reference solution were prepared according to the optimized test solution preparation method described above. Analysis was performed under the chromatographic conditions described above to verify the chromatographic conditions and system suitability of the *Commelina communis* decoction standard, and to investigate whether the blank solvent would cause interference. The chromatograms of the blank solvent and the test solution of the *Commelina communis* decoction standard are shown in the appendix. Figure 10 As shown in (a)-(b).

[0125] As shown in the figure, the blank solvent does not interfere with the characteristic chromatogram of the test sample. The chromatographic method system has good applicability and specificity, and can be used as a detection method for the standard decoction of Commelina communis.

[0126] (2) Precision

[0127] Take the same sample solution and inject it six times under the chromatographic conditions described above. Measure the relative retention time and relative peak area of ​​the 14 common peaks. Use peak number 4 as the reference peak for the first 10 characteristic peaks and peak number 14 as the reference peak for the last 4 characteristic peaks. The RSD of the relative retention time is less than 2.0%, indicating good instrument precision. See the attached precision chromatogram. Figure 11 The detailed results are shown in Table 9-10 below.

[0128] Table 9. Results of Instrument Precision and Relative Retention Time Tests

[0129]

[0130] Table 10 Results of Instrument Precision Relative Peak Area Test

[0131]

[0132]

[0133] (3) Method repeatability

[0134] Six samples from the same batch were taken, and the relative retention times and relative peak areas of the 14 common peaks were determined under the chromatographic conditions described above. The results showed that the RSD values ​​of the relative retention times of each characteristic peak were all less than 2.0%, indicating that the method has good repeatability. See the attached chromatogram for the specific repeatability test. Figure 12The detailed results are shown in Tables 11-12 below.

[0135] Table 11 Results of the method repeatability relative retention time test

[0136]

[0137]

[0138] Table 12 Results of the method repeatability relative peak area test

[0139]

[0140]

[0141] (4) Intermediate precision (different operators)

[0142] Three inspectors, at different times, collected the same sample of *Commelina communis* decoction and prepared the sample according to the chromatographic conditions described above. The relative retention time and relative peak area of ​​each common peak were measured using the same instrument. The results showed that the RSD values ​​of the relative retention times of each characteristic peak were all less than 2.0%, indicating good intermediate precision of the method. The chromatograms for the intermediate precision test are attached. Figure 13 The detailed results are shown in Tables 13-14 below.

[0143] Table 13 Intermediate Precision Relative Retention Time Test Results (Different Operators)

[0144]

[0145] Table 14. Results of intermediate precision relative peak area tests (by different operators)

[0146]

[0147]

[0148] (5) Stability test

[0149] Using the same batch of test samples, after preparation, chromatographically analyzed under the conditions described above, the samples were injected at 0, 4, 8, 12, 16, and 24 hours. The relative retention times and relative peak areas of the 14 common peaks were determined. The relative retention times of each characteristic peak and the reference peak (S peak) were less than 2%. The results indicate that the test solution was stable within 24 hours, meeting the determination requirements. See the attached chromatogram for the specific stability test. Figure 14 The detailed results are shown in Tables 15-16 below.

[0150] 15. Results of the stability test relative to retention time

[0151]

[0152]

[0153] Table 16 Results of Stability Relative Peak Area Test

[0154]

[0155]

[0156] (6) Durability test at different flow rates

[0157] The same sample solution was tested under the above chromatographic conditions at different flow rates (0.9 ml / min, 1.0 ml / min, and 1.1 ml / min). The relative retention times and relative peak areas of each characteristic peak were measured. The results showed that the RSD values ​​of the relative retention times of each characteristic peak and the reference peak S were all less than 4.0%. However, the peak shapes of some chromatographic peaks deteriorated, indicating that even small changes in flow rate had a certain impact on each characteristic peak. Therefore, this method is considered to have poor flow rate robustness, and a fixed flow rate of 1.0 ml / min is recommended. The chromatograms for different flow rates (0.9 ml / min, 1.0 ml / min, and 1.1 ml / min) are attached. Figure 15 (a)-(c), detailed results are shown in Tables 17-18 below.

[0158] Table 17 Results of relative retention times for different flow velocities

[0159]

[0160]

[0161] Table 18 Results of relative peak area at different flow velocities

[0162]

[0163] (7) Durability test at different column temperatures

[0164] The same sample solution was analyzed under the above chromatographic conditions at different column temperatures (23℃, 25℃, and 27℃). The relative retention times and relative peak areas of each characteristic peak were measured. The results showed that the RSD values ​​of the relative retention times of each characteristic peak and the reference peak S were all less than 2.0%. However, the peak shapes of some peaks deteriorated, indicating that even small changes in column temperature had a certain impact on each characteristic peak. Therefore, this method is considered to have poor column temperature robustness, and it is recommended to fix the column temperature at 25℃. The chromatograms of the experiments at different column temperatures (23℃, 25℃, and 27℃) are attached. Figure 16 (a)-(c), detailed results are shown in Tables 19-20 below.

[0165] Table 19 Comparison of relative retention times at different column temperatures

[0166]

[0167]

[0168] Table 20 Comparison of relative peak areas at different column temperatures

[0169]

[0170] Based on the above methodological investigation results, among the 14 common peaks in the established standard decoction of Commelina communis slices [characteristic chromatogram], each chromatographic peak is affected to some extent by column temperature and mobile phase flow rate, while the other chromatographic conditions have little effect. The relative retention time values ​​are within ±10%. To improve its robustness, it is recommended to control the specified value range within ±10%.

[0171] In summary, the method for determining the characteristic chromatogram of Commelina communis formula particles is as follows: using octadecylsilane-bonded silica gel as the packing material (column length 250 mm, inner diameter 4.6 mm, particle size 5 μm), acetonitrile as mobile phase A, and 0.1% formic acid as mobile phase B, gradient elution is performed according to the specifications in Table 5 of Example 3 above; the flow rate is 1.0 ml per minute, the column temperature is 25 °C, and the detection wavelength is 260 nm.

[0172] Preparation of reference solutions: Accurately weigh 1g of Commelina communis reference material, place it in a stoppered conical flask, add 40mL of water, heat under reflux for 45 minutes, filter, evaporate the filtrate to dryness, dissolve the residue in 10mL of water, extract with 20mL of water-saturated n-butanol, extract three times, combine the n-butanol extracts, evaporate to dryness, add 5mL of methanol to the residue, sonicate (500W power, 37kHz frequency) for 10 minutes, filter, and use the filtrate as the reference solution. Separately, accurately weigh appropriate amounts of hypoxanthine, adenosine, guanosine, L-tryptophan, and isohypogonin reference standards, and dissolve them in methanol to prepare solutions containing 50μg per mL, as reference solutions.

[0173] Preparation of the test solution: Take 0.5g of the powder, add 10mL of water to dissolve it, extract with 20mL of water-saturated n-butanol, extract 3 times, combine the n-butanol extracts, evaporate to dryness, add 5mL of methanol to the residue, sonicate (power 500W, frequency 37kHz) for 10 minutes, filter, and take the filtrate to obtain the test solution.

[0174] Assay: Accurately pipette 10 μl each of the reference solution and the test solution into the liquid chromatograph and determine the result. See the attached characteristic chromatogram for the reference solution. Figure 17 As shown.

[0175] The chromatogram of the test sample should show 14 characteristic peaks, and the retention times should correspond to the 14 characteristic peaks in the chromatogram of the reference medicinal material. Peaks 1, 3, 4, 8, and 14 should correspond to the retention times of the corresponding reference peaks. The peak corresponding to the guanosine reference peak is designated as peak S1, and the relative retention times of peaks 1-10 with peak S1 should be calculated. The peak corresponding to the isopropanol reference peak is designated as peak S2, and the relative retention times of peaks 11-13 should be calculated. The relative retention time of peak S2 should be within ±10% of the specified value, which is: 0.48 (peak 1), 0.80 (peak 2), 0.86 (peak 3), 1.08 (peak 5), 1.29 (peak 6), 1.61 (peak 7), 1.74 (peak 8), 2.15 (peak 9), 2.46 (peak 10), 0.79 (peak 11), 0.84 (peak 12), and 0.93 (peak 13).

[0176] After comparison, the peaks were determined to be: peak 1: hypoxanthine; peak 3: adenosine; peak 4: guanosine; peak 8: L-tryptophan; peak 14: isopropanol.

[0177] Example 5

[0178] This embodiment involves the determination of characteristic chromatograms for three batches of *Commelina communis* (Duckweed) standard decoctions. Following the established *Commelina communis* characteristic chromatogram method, the characteristic chromatograms of the three batches of *Commelina communis* standard decoctions were determined, and the results are as follows. The results show that the relative retention times of each characteristic peak in the three batches of *Commelina communis* extracts are all within ±10% of the specified values. The characteristic chromatograms of the three batches of *Commelina communis* standard decoctions are attached. Figure 18 The detailed results are shown in Tables 21-22 below.

[0179] Table 21. Relative retention time results of characteristic chromatogram determination of three batches of Commelina communis standard decoction.

[0180]

[0181]

[0182] Table 22. Results of relative peak area determination of characteristic chromatograms of three batches of Commelina communis standard decoction.

[0183]

[0184] Example 6

[0185] This embodiment is based on the method of using the medicinal material of Commelina communis.

[0186] (1) Validation of chromatographic conditions and system suitability

[0187] The test solution and reference solution were prepared according to the method for preparing the test solution, and analyzed under the chromatographic conditions described above to verify the chromatographic conditions and system suitability of the *Commelina communis* herb, and to investigate whether the blank solvent would cause interference. The chromatograms are recorded as attached. Figure 19 It is evident that the blank solvent does not interfere with the characteristic chromatogram of the test sample, and the chromatographic method system exhibits good applicability and specificity, making it suitable as a detection method for Commelina communis herb.

[0188] (2) Precision

[0189] The same sample solution was injected six times under the chromatographic conditions described above. The relative retention times and relative peak areas of the 14 common peaks were determined. Peak 4 was used as the reference peak for the first 10 characteristic peaks, and peak 14 was used as the reference peak for the last four characteristic peaks. The RSD of the relative retention times was less than 2.0%, indicating good instrument precision. Detailed results are attached. Figure 20 And see Tables 23-24 below.

[0190] Table 23 Results of Instrument Precision and Relative Retention Time Tests

[0191]

[0192]

[0193] Table 24 Results of Instrument Precision Relative Peak Area Test

[0194]

[0195] (3) Method repeatability

[0196] Six samples from the same batch were taken, and the relative retention times and relative peak areas of the 14 common peaks were determined under the chromatographic conditions described above. The results showed that the RSD values ​​of the relative retention times of each characteristic peak were all less than 2.0%, indicating that the method had good repeatability. Detailed results are shown in Tables 25-26 below.

[0197] Table 25 Results of the method repeatability relative retention time test

[0198]

[0199] Table 26 Results of Method Repeatability Relative Peak Area Test

[0200]

[0201]

[0202] (4) Intermediate precision (different operators)

[0203] Three inspectors, at different times, took the same sample of Commelina communis decoction and prepared the sample according to the test sample preparation method under the above chromatographic conditions. The relative retention time and relative peak area of ​​each common peak were measured using the same equipment. The results showed that the RSD values ​​of the relative retention times of each characteristic peak were all less than 2.0%, indicating good intermediate precision of this method. Detailed results are shown in Tables 27-28 below.

[0204] Table 27 Intermediate Precision Relative Retention Time Test Results (Different Operators)

[0205]

[0206]

[0207] Table 28 Intermediate Precision Relative Peak Area Test Results (Different Operators)

[0208]

[0209] (5) Stability test

[0210] Samples from the same batch were injected at 0, 4, 8, 12, 16, and 24 hours after preparation, under the chromatographic conditions described above. The relative retention times and relative peak areas of the 14 common peaks were determined. The relative retention times of each characteristic peak and the reference peak S were less than 2%. The results indicate that the sample solution is stable within 24 hours and meets the determination requirements. Detailed results are shown in Tables 29-30 below.

[0211] Table 29 Results of the relative retention time test for stability

[0212]

[0213] Table 30 Results of Stability Relative Peak Area Test

[0214]

[0215]

[0216] (6) Durability test at different flow rates

[0217] Take the same sample solution and, under the above chromatographic conditions, detect the same sample solution at different flow rates (0.9 ml / min, 1.0 ml / min, and 1.1 ml / min). Measure the relative retention time and relative peak area of ​​each characteristic peak. The results show that the RSD values ​​of the relative retention times of each characteristic peak and the reference peak S are all less than 4.0%. However, the peak shapes of some chromatographic peaks deteriorated, indicating that small changes in flow rate have a certain impact on each characteristic peak. It is considered that this method has poor robustness to flow rate, and it is recommended to fix the flow rate at 1.0 ml / min. See Tables 31-32 below for detailed results.

[0218] Table 31 Results of relative retention time for different flow velocities

[0219]

[0220] Table 32 Results of relative peak area at different flow velocities

[0221]

[0222] (7) Durability test at different column temperatures

[0223] Take the same sample solution and, under the above chromatographic conditions, use different column temperatures (23℃, 25℃, and 27℃) to detect the same sample solution. Measure the relative retention time and relative peak area of ​​each characteristic peak. The results show that the RSD values ​​of the relative retention times of each characteristic peak and the reference peak S are all less than 2.0%. However, the peak shape of some chromatographic peaks deteriorated, indicating that small changes in column temperature have a certain impact on each characteristic peak. It is believed that this method has poor column temperature robustness, and it is recommended to fix the column temperature at 25℃. For detailed results, please refer to Tables 33-34 below.

[0224] Table 33 Comparison of relative retention times at different column temperatures

[0225]

[0226] Table 34 Comparison of relative peak areas at different column temperatures

[0227]

[0228]

[0229] Based on the methodological findings described above, among the 14 common peaks in the characteristic chromatogram of Commelina communis herb, each peak was affected to some extent by column temperature and mobile phase flow rate, while the other chromatographic conditions had little effect. The relative retention times were within ±10%. To improve robustness, it is recommended that the specified value range be controlled within ±10%.

[0230] In summary, the method for determining the characteristic spectrum of the herbal medicine *Duckweed* in this invention is as follows: using octadecylsilane-bonded silica gel as the packing material (column length 250 mm, inner diameter 4.6 mm, particle size 5 μm), acetonitrile as mobile phase A, and 0.1% formic acid as mobile phase B, gradient elution is performed according to the specifications in Table 5 of the aforementioned examples; the flow rate is 1.0 ml per minute, the column temperature is 25 °C, and the detection wavelength is 260 nm.

[0231] Preparation of the reference solution: Accurately weigh 1 g of Commelina communis reference material and place it in a stoppered conical flask. Add 40 mL of water, heat under reflux for 45 minutes, filter, evaporate the filtrate to dryness, dissolve the residue in 10 mL of water, and extract with 20 mL of water-saturated n-butanol three times. Combine the n-butanol extracts, evaporate to dryness, add 5 mL of methanol to the residue, and sonicate (500 W, 37 kHz) for 10 minutes. Filter and use the filtrate as the reference solution. Separately, accurately weigh appropriate amounts of hypoxanthine, adenosine, guanosine, L-tryptophan, and isohypogonin reference standards, and dissolve them in methanol to prepare solutions containing 50 μg per mL, as reference solutions.

[0232] Preparation of the test solution: Take 0.5 g of the powder, add 10 mL of water to dissolve it, extract with 20 mL of water-saturated n-butanol, extract 3 times, combine the n-butanol extracts, evaporate to dryness, add 5 mL of methanol to the residue, sonicate (power 500 W, frequency 37 kHz) for 10 minutes, filter, and take the filtrate to obtain the test solution.

[0233] The assay involves accurately injecting 10 μl each of the reference solution and the test solution into the liquid chromatograph and measuring the concentration. The characteristic chromatograms of the reference solution are attached. Figure 20 As shown.

[0234] It is evident that the characteristic chromatogram of the test sample should exhibit 14 characteristic peaks, which should correspond to the retention times of the 14 characteristic peaks in the chromatogram of the reference medicinal material. Peaks 1, 3, 4, 8, and 14 should correspond to the retention times of the corresponding reference peaks. The peak corresponding to the guanosine reference peak is peak S1; calculate the relative retention times of peaks 1-10 with peak S1. The peak corresponding to the isopropanol reference peak is peak S2; calculate the relative retention times of peaks 11-10. The relative retention times of peak 13 and peak S2 should be within ±10% of the specified values, which are: 0.48 (peak 1), 0.80 (peak 2), 0.86 (peak 3), 1.08 (peak 5), 1.29 (peak 6), 1.61 (peak 7), 1.74 (peak 8), 2.15 (peak 9), 2.46 (peak 10), 0.79 (peak 11), 0.84 (peak 12), and 0.93 (peak 13).

[0235] After comparison, the peaks were identified as follows: Peak 1: hypoxanthine; Peak 3: adenosine; Peak 4: guanosine; Peak 8: L-tryptophan; Peak 14: isopropanol.

[0236] Example 7

[0237] This embodiment is based on the method determined in the aforementioned embodiment 6, and performs characteristic spectrum determination on multiple batches of medicinal materials.

[0238] The characteristic chromatograms of 20 batches of *Commelina communis* medicinal materials were determined according to the established method for [characteristic chromatogram] analysis. The results are shown in Tables 35-36 below.

[0239] Table 35. Relative retention time results of characteristic chromatogram determination of 20 batches of Commelina communis medicinal materials.

[0240]

[0241]

[0242]

[0243] Table 36. Results of relative peak area determination of characteristic spectra of 20 batches of Commelina communis medicinal materials

[0244]

[0245]

[0246]

[0247] The results showed that the relative retention times of each characteristic peak in the 20 batches of *Commelina communis* medicinal materials were all within ±10% of the specified values. Combining the methodological validation results of the *Commelina communis* medicinal material characteristic chromatogram and the common characteristic peaks of the 20 batches, analysis revealed 14 characteristic peaks in total.

[0248] Obviously, the above embodiments are merely illustrative examples for clear explanation and are not intended to limit the implementation. Those skilled in the art will recognize that other variations or modifications can be made based on the above description. It is neither necessary nor possible to exhaustively list all possible implementations here. However, obvious variations or modifications derived therefrom are still within the scope of protection of this invention.

Claims

1. A method for constructing a characteristic spectrum of the herbal medicine *Commelina communis* and its pharmaceutical preparations, characterized in that, The procedure includes high-performance liquid chromatography (HPLC) detection of test solutions of Commelina communis herb and / or Commelina communis herb preparations; The preparation method of the test sample solution includes: taking the test sample and accurately adding it to the first solvent for mixing, adding the second solvent for extraction, evaporating the combined extracts to dryness, adding the residue to the third solvent for redissolution, and filtering after ultrasonic treatment to obtain the final product; The first solvent is water; The second solvent is water-saturated n-butanol; The third solvent is methanol; The method further includes the steps of preparing a reference solution and constructing a characteristic chromatogram of the reference solution based on the high performance liquid chromatography. The reference standards include hypoxanthine, adenosine, guanosine, L-tryptophan, and isochorin; The method for preparing the reference solution includes: precisely adding the reference standard to a third solvent and mixing to obtain the solution; Chromatographic conditions included: using an Atlantis T3 column, acetonitrile as mobile phase A, 0.08%–0.12% formic acid solution as mobile phase B, and a detection wavelength of 260 nm, with gradient elution performed according to the following procedure: 0-5 min, A:B ratio is 2%:98%; 5-15 min, A:B ratio changes from 2% to 7%; 98%:B ratio changes from 93% to 93%. 15-25 min, A:B ratio changes from 7% to 12%; A:B ratio changes from 93% to 88%. 25-35 min, A:B ratio 12%:88%; 35-40 minutes, A:B ratio changed from 12% to 14%; 88%:B ratio changed from 88% to 86%. 40-60 min, A:B ratio is 14%:86%.

2. The method for constructing the characteristic spectrum of the herbal medicine and pharmaceutical preparation of *Commelina communis* according to claim 1, characterized in that, The chromatographic conditions in the high-performance liquid chromatography detection step also include: a flow rate of 0.9–1.1 ml / min and a column temperature of 23–27 °C.

3. The method for constructing the characteristic spectrum of *Commelina communis* herbal medicine and pharmaceutical preparation according to claim 1 or 2, characterized in that, The method further includes the step of preparing a reference solution of the control medicinal material, specifically including: taking the reference medicinal material of Commelina communis and accurately adding it to the first solvent and mixing it, heating and refluxing it, collecting the filtrate and evaporating it to dryness, adding the residue to the first solvent and mixing it, adding the second solvent and extracting it, combining the extracts and evaporating them to dryness, adding the residue to the third solvent to redissolve it, and filtering it after ultrasonic treatment to obtain the final product.

4. The method for constructing the characteristic spectrum of the herbal medicine and pharmaceutical preparation of *Commelina communis* according to claim 1 or 2, characterized in that, The ultrasonic treatment step has a power of 450-550W and a frequency of 35-40kHz.

5. The method for constructing the characteristic spectrum of the herbal medicine and pharmaceutical preparation of *Commelina communis* according to claim 1 or 2, characterized in that, The medicinal preparations of Commelina communis include Commelina communis formula granules, Commelina communis slices, or Commelina communis standard decoction.

6. The method for constructing the characteristic spectrum of *Commelina communis* herbal medicine and pharmaceutical preparation according to claim 1 or 2, characterized in that, The characteristic spectrum has 14 characteristic peaks. Peaks 1, 3, 4, 8, and 14 correspond to the retention times of the corresponding reference peaks. The peak corresponding to the guanosine reference peak is peak S1, and the relative retention times of peaks 1-10 with peak S1 are calculated. The peak corresponding to the isopropanol reference peak is peak S2, and the relative retention times of peaks 11-13 with peak S2 are calculated. The relative retention times should be within ±10% of the specified values. The specified values ​​for each characteristic peak are: peak 1: 0.48, peak 2: 0.80, peak 3: 0.86, peak 5: 1.08, peak 6: 1.29, peak 7: 1.61, peak 8: 1.74, peak 9: 2.15, peak 10: 2.46, peak 11: 0.79, peak 12: 0.84, and peak 13: 0.

93.

7. The application of the method for constructing characteristic spectra of *Commelina communis* medicinal materials and pharmaceutical preparations according to any one of claims 1-6 in the field of quality testing of *Commelina communis* medicinal materials and pharmaceutical preparations.

8. A method for quality testing of *Commelina communis* herbal medicine and pharmaceutical preparations, characterized in that, The method includes the steps of constructing the feature map and the reference feature map according to any one of claims 1-6, and the step of comparing the feature map with the reference feature map.

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