CN104730193A - Detection method of volatile component in heart stabilizing particles - Google Patents
Detection method of volatile component in heart stabilizing particles Download PDFInfo
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Abstract
The invention provides a detection method of a volatile component in heart stabilizing particles. By adopting a head space solid phase microextraction technology, the volatile component in the heart stabilizing particles can be extracted. By utilizing a gas chromatography and mass spectrum combined technology, the volatile component 74 can be isolated and identified together; 10 batches of heart stabilizing particles are analyzed, the relative content range of the main volatile component can be acquired. The head space solid phase microextraction technology is combined with the gas chromatography and mass spectrum combined technology for the first time, and the volatile components in the heart stabilizing particles are extracted and analyzed, so that the defects of analysis on the volatile components of the heart stabilizing particles are overcome, and a comprehensive detection method is provided for quality control of medicines.
Description
Technical field
The present invention relates to Chinese patent drug detection method field, especially relate to the detection method of volatile ingredient in a kind of WENXIN KELI.
Background technology
WENXIN KELI, as conventional anti-arrhythmia Chinese patent drug, primarily of medicinal material compositions such as Radix Codonopsis, sealwort, pseudo-ginseng and rhizoma nardostachyos, has supplementing qi and nourishing yin, multiple arteries and veins of surely throbbing with fear, effect promoting blood circulation and removing blood stasis.State approval code is: Z10950026.Cure mainly: deficiency of both qi and yin is held concurrently the palpitation and uneasiness caused by heart arteries and veins stasis blocking, shortness of breath and fatigue, dizzy vexed, feeling of oppression and pain in the chest.Be applicable to the arrhythmia cordis such as premature beat, atrial fibrillation, nodal tachycardia that a variety of causes causes.Clinical practice is evident in efficacy.Except auxiliary material, in side, principal ingredient is polysaccharide, saponin(e and volatile ingredient (Spikenard volatile oil), under 2010 editions " Chinese Pharmacopoeias " record WENXIN KELI item, specifies to differentiate with Radix Codonopsis, pseudo-ginseng and sealwort, with Panax Notoginseng saponin R
1, ginsenoside Rg
1and Rb
1total content carry out assay, the specification of the volatile ingredient being mainly derived from rhizoma nardostachyos is not related to.Rhizoma nardostachyos " makes medicine " at Fang Zhongwei, and clinical effect in recent years in anti-arrhythmia and pharmacological mechanism report more, more and more receive publicity.The present invention " status " of clear and definite rhizoma nardostachyos in side, solve the resource scarcity problem that faces and WENXIN KELI side is optimized to simplify etc. in tool be of great significance.
Headspace solid-phase microextraction (head space-solid phase microextraction, HS-SPME) be a novel sample pre-treatments of rising the nineties in 20th century and beneficiation technologies, it is a kind of extraction and separation technology with features such as rate of extraction are fast, simple to operate, bioaccumulation efficiency is high, required sample is few grown up on Solid-Phase Extraction (solid phaseextraction, SPE) basis.
Summary of the invention
The invention provides the detection method of volatile ingredient in a kind of WENXIN KELI, object is, for the quality assessment of WENXIN KELI, and is the volatile ingredient quality control approach effectively in WENXIN KELI.
For solving the problems of the technologies described above, the detection method of volatile ingredient in a kind of WENXIN KELI provided by the invention, the present invention adopts the volatile ingredient in headspace solid-phase microextraction technology extraction WENXIN KELI, and utilizes gas chromatography-mass spectrography Analysis and Identification.
The detection method of the volatile ingredient in WENXIN KELI of the present invention, wherein said headspace solid-phase microextraction technical step is get WENXIN KELI, saturated sodium-chloride water solution, stir, insert extracting head, extraction time 25 ~ 35min, take out, by the volatile ingredient of extraction gained, inject gas chromatograph injection port, injector temperature 250 DEG C, desorption time 4 ~ 6min, sample introduction pattern is Splitless injecting samples, and utilizes gas chromatography-mass spectrography Analysis and Identification.
Preferably, the model of described extracting head is 100 μm of PDMS.
Preferably, described extraction time 30min.
Preferably, described desorption time 5min.
Preferably, described gas chromatography-mass spectrography Analysis and Identification condition is: chromatographic condition: HP-5MS, 60m × 320 μm × 0.25 μm; Carrier gas: He, flow 0.8mL/min; Vapourizing temperature: 250 DEG C; Interface temperature: 280 DEG C; Chromatographic column initial temperature 50 DEG C, keeps 1min, rises to 130 DEG C with 10 DEG C/min heating rate, keeps 6min, rise to 145 DEG C with 1 DEG C/min heating rate, keep 5min, rise to 160 DEG C with 1 DEG C/min heating rate, keep 3min, rise to 220 DEG C with 3 DEG C/min heating rate, keep 1min;
Mass Spectrometry Conditions: ionization mode: EI source, energy 70eV; Ion source temperature: 230 DEG C; Quadrupole rod temperature: 150 DEG C; Mass scan range: 35 ~ 800amu; Electron-multiplier voltage: 1400V.
Further, according to above-mentioned set up assay method, measure the finger-print that in WENXIN KELI, volatile ingredient obtains, have 19 common characteristic fingerprint peakses in the finger-print generated according to the method, the retention time obtained successively is: No. 1 peak is 10.78 ± 0.006min, No. 2 peaks are: 20.42 ± 0.007min, No. 3 peaks are: 21.03 ± 0.007min, No. 4 peaks are 21.39 ± 0.007min, No. 5 peaks are: 22.56 ± 0.008min, No. 6 peaks are 23.36 ± 0.009min, No. 7 peaks are: 24.71 ± 0.014min, No. 8 peaks are: 25.23 ± 0.012min, No. 9 peaks are: 26.11 ± 0.037min, No. 10 peaks are: 27.42 ± 0.022min, No. 11 peaks are: 34.80 ± 0.014min, No. 12 peaks are: 36.09 ± 0.018min, No. 13 peaks are: 37.25 ± 0.035min, No. 14 peaks are: 37.70 ± 0.048min, No. 15 peaks are: 40.85 ± 0.262min, No. 16 peaks are: 41.19 ± 0.053min, No. 17 peaks are: 44.29 ± 0.012min, No. 18 peaks are: 44.64 ± 0.014min, No. 19 peaks are: 60.81 ± 0.008min.
Further, according to above-mentioned set up assay method, measure the finger-print that in WENXIN KELI, volatile ingredient obtains, 19 common characteristic fingerprint peakses are had in the finger-print generated according to the method, wherein No. 1 peak is: Eucalyptol Cineole, No. 2 peaks are: δ-EIemene δ-elemene, No. 3 peaks are: α-Cubebene α-cubebene, No. 4 peaks are: Tetracyclo [4.4.1.1 (7, 10) .0 (2, 5)] dodec-3-en-11-ol, No. 5 peaks are: α-Copaene α-pepper alkene, No. 6 peaks are: β-Patchoulene β-patchoulene, No. 7 peaks are: Cadinene cadinene, No. 8 peaks are: (+)-Calarene Calamus alkene, No. 9 peaks are: (-)-Aristolene aristolene, No. 10 peaks are: Caryophyllene cloves alkene, No. 11 peaks are: Spathulenol spainulenol, No. 12 peaks are: Globulol globulol, No. 13 peaks are: (-)-Spathulenol (-)-spainulenol, No. 14 peaks are: the different aromadendrene epoxide of Isoaromadendrene epoxide, No. 15 peaks are: Humulene epoxide2 humulene epoxide 2, No. 16 peaks are: 1, 3a-Ethano (1H) inden-4-ol, octahydro-2, 2, 4, 7a-tetramethyl-, No. 17 peaks are: Widdrol widdrol, No. 18 peaks are: Patchoulialcohol Patchoulicalcohol, No. 19 peaks are: 4-(3, 3-Dimethyl-but-1-ynyl)-4-hydroxy-2, 6, 6-trimethylcyclohex-2-enone.
The present invention adopts the qualitative and quantitative analysis of volatile ingredient, from total ion current figure, each component percentages is calculated through chem workstation process and by area normalization method, and its KI value is calculated to chromatographic peak each in total ion current figure, utilize mass spectrogram corresponding to each chromatographic peak to carry out the inspection of NIST08.L standard spectrum storehouse and determine each component.Measure through multiple batches of volatile component content, adopt preferred gas chromatography-mass spectrography analysis condition, the 10 batches of WENXIN KELIs randomly drawed are analyzed, choose and represent peak and carry out statistical treatment.
Headspace solid-phase microextraction (head space-solid phase microextraction, HS-SPME) has the advantages that rate of extraction is fast, simple to operate, bioaccumulation efficiency is high.
Technical solution of the present invention adopts headspace solid-phase microextraction and Gas chromatographyMass spectrometry (HS-SPME-GC-MS) extract WENXIN KELI and analyze, and being divided into from determining 74 components, accounting for 80.90% of total peak area.
The present invention adopts headspace solid-phase microextraction and Gas chromatographyMass spectrometry, extracts 10 batches of WENXIN KELIs and analyzes, and have chosen 19 and represent peak and carry out relative content statistics, 19 relative area sums representing peak are not less than 40% of the total area.
Beneficial effect of the present invention:
The present invention adopts headspace solid-phase microextraction to carry out extracting and analyze to volatile ingredient in WENXIN KELI in conjunction with GC-MS(gas chromatography-mass spectrography) first, by a large amount of experimental studies, optimizes GC conditions-Mass Spectrometry Conditions.And Content inspect has been carried out to the sample of 10 batches, compensate in the past to the deficiency that WENXIN KELI volatile matter is analyzed, also construct the GC/MS total ion current figure of WENXIN KELI volatile ingredient.The present invention adopts headspace solid-phase microextraction and Gas chromatographyMass spectrometry extract WENXIN KELI and analyze, and being divided into from identifying volatile ingredient 74, accounting for 80.90% of general volatile composition.The chemical composition and content thereof that objectively respond WENXIN KELI main volatile component can be compared, effectively characterize the quality of WENXIN KELI volatile ingredient.Further, analyze the 10 batches of WENXIN KELIs randomly drawed, 19 that choose that degree of separation is good, purity is high represent peak and carry out statistical treatment, and found that, the retention time at each peak and peak area are all more stable.This also illustrates the detection method that the present invention sets up, there is good reappearance and reliability, in WENXIN KELI, the quality control of volatile ingredient provides reliable approach.
Accompanying drawing explanation
Fig. 1 is the GC-MS total ion current figure of volatile ingredient in WENXIN KELI.
Fig. 2 is that the GC-MS total ion current of volatile ingredient in 10 batches of WENXIN KELIs stacks figure.
Fig. 3 is that 10 batches of WENXIN KELIs represent chromatographic peak relative content result.Compound title wherein in Fig. 3 representated by 19 chromatographic peaks is respectively 1-Eucalyptol Cineole, 2-δ-EIemene δ-elemene, 3-α-Cubebene α-cubebene, 4-Tetracyclo [4.4.1.1 (7, 10) .0 (2, 5)] dodec-3-en-11-ol, 5-α-Copaene α-pepper alkene, 6-β-Patchoulene β-patchoulene, 7-Cadinene cadinene, 8-(+)-Calarene Calamus alkene, 9-(-)-Aristolene aristolene, 10-Caryophyllene cloves alkene, 11-Spathulenol spainulenol, 12-Globulol globulol, 13-(-)-Spathulenol (-)-spainulenol, the different aromadendrene epoxide of 14-Isoaromadendrene epoxide, 15-Humulene epoxide 2 humulene epoxide 2, 16-1, 3a-Ethano (1H) inden-4-ol, octahydro-2, 2, 4, 7a-tetramethyl-, 17-Widdrol widdrol, 18-Patchouli alcohol Patchoulicalcohol, 19-4-(3, 3-Dimethyl-but-1-ynyl)-4-hydroxy-2, 6, 6-trimethylcyclohex-2-enone.
Embodiment
In order to make those skilled in the art better understand technical scheme of the present invention, below in conjunction with the drawings and the specific embodiments, technical scheme of the present invention is described in further detail.
1, the detection method of volatile ingredient in WENXIN KELI
A () headspace solid-phase microextraction extracts volatile ingredient:
By WENXIN KELI (Shandong Buchang Pharmaceutical Co., Ltd., 5g/ bag, product batch number: 1405023) 0.51g, analysis pure sodium chloride 3.6g and water 12mL put into 150ml ml headspace bottle, add stirrer, seal pad (containing transparent blue silica gel/PTFE pad) and aluminium cap, 70 DEG C of magnetic agitation, insert 100 μm of PDMS extracting head (Supelco, Bellefonte, Pa).Make extracting head stretch out 1cm, extraction 30min, take out, insert gas chromatograph injection port immediately, injector temperature 250 DEG C, desorb 5min, sample introduction pattern is Splitless injecting samples.
(b) gas chromatography-mass spectrography analysis condition:
Agilent 7890A-5975C GC-MS gas chromatography mass spectrometry mass spectrometer (Agilent scientific & technical corporation of the U.S.), Chromatographic data system (MSD ChemStation E.02.01, Agilent scientific & technical corporation of the U.S.); Chromatographic column: HP-5MS (60m × 320 μm × 0.25 μm); Carrier gas: He, flow 0.8mL/min; Vapourizing temperature: 250 DEG C; Interface temperature: 280 DEG C; Chromatographic column initial temperature 50 DEG C (keeping 1min), rise to 130 DEG C (keeping 6min) with 10 DEG C/min heating rate, rise to 145 DEG C (keeping 5min) with 1 DEG C/min heating rate, rise to 160 DEG C (keeping 3min) with 1 DEG C/min heating rate, rise to 220 DEG C with 3 DEG C/min heating rate, finally keep 1min.Ionization mode: EI source, energy 70eV; Ion source temperature: 230 DEG C; Quadrupole rod temperature: 150 DEG C; Mass range: (35 ~ 800) amu; Electron-multiplier voltage: 1400V.
(c) KI pH-value determination pH:
Get n-alkane mixing reference substance (C
8-C
20, FLUKA company of the U.S., numbering: 04070,40mg/Lin hexane) 0.8 μ L, by gas chromatography-mass spectrography analysis condition under above-mentioned steps (b) item, record each n-alkane retention time, adopt the KI value of each component of linear temperature increase formulae discovery
In formula, tx, tn and tn+1 are respectively analyzed component and carbon number and are in n-alkane (tn<tx<tn+1) eluting peak retention time (min) between n and n+1, and measurement result is in table 1.
Table 1 n-alkane retention time
The qualitative and quantitative analysis of (d) volatile ingredient:
The GC-MS total ion current figure of WENXIN KELI volatile ingredient is shown in Fig. 1.
From total ion current figure, each component percentages is calculated through chem workstation process and by area normalization method, its KI value is calculated to chromatographic peak each in total ion current figure, utilize mass spectrogram corresponding to each chromatographic peak to carry out the inspection of NIST08.L standard spectrum storehouse and determine each component, detailed results is in table 2.
Volatile chemical component in table 2 WENXIN KELI
Note :-: exceed n-alkane maximum retention time
The present invention adopts headspace solid-phase microextraction and Gas chromatographyMass spectrometry, and (i.e. HS-SPME-GC-MS) extracts WENXIN KELI and analyze, and being divided into from identifying volatile ingredient 74, accounting for 80.90% of general volatile composition.
As seen from Table 2, in WENXIN KELI, volatile ingredient is mainly different aromadendrene epoxide (11.71%), (-)-spainulenol (6.01%), 1,3a-Ethano (1H) inden-4-ol, octahydro-2,2, the sesquiterpenoidss such as 4,7a-tetramethyl-(5.08%), humulene epoxide 2 (4.14%).
E () multiple batches of volatile component content measures:
The present invention adopts headspace solid-phase microextraction and Gas chromatographyMass spectrometry, analyzes the 10 batches of WENXIN KELIs randomly drawed, and 19 that choose that degree of separation is good, purity is high represent peak and carry out statistical treatment, and the relative content often criticized the results are shown in Table 3.Experimental result shows: 19 peak areas of the volatile ingredient in 10 batches of WENXIN KELIs and retention time more stable.After rejecting abnormalities value, carry out statistical study, show that WENXIN KELI represents retention time and the relative content scope at peak, the results are shown in Table 4.
GraphPad Prism 5 software analysis WENXIN KELI represents peak relative content and the results are shown in Figure 3.
Table 4 WENXIN KELI represents peak relative content scope
Claims (8)
1. the detection method of volatile ingredient in WENXIN KELI, is characterized in that: adopt headspace solid-phase microextraction technology to extract volatile ingredient in WENXIN KELI, and utilize gas chromatography-mass spectrography Analysis and Identification.
2. detection method as claimed in claim 1, is characterized in that: described headspace solid-phase microextraction technical step is get WENXIN KELI, saturated sodium-chloride water solution, stir, insert extracting head, extraction time 25 ~ 35min, take out, by the volatile ingredient of extraction gained, inject gas chromatograph injection port, injector temperature 250 DEG C, desorption time 4 ~ 6min, sample introduction pattern is Splitless injecting samples, and utilizes gas chromatography-mass spectrography Analysis and Identification.
3. detection method as claimed in claim 2, is characterized in that: the model of described extracting head is 100 μm of PDMS.
4. detection method as claimed in claim 2, is characterized in that: described extraction time 30min.
5. detection method as claimed in claim 2, is characterized in that: described desorption time 5min.
6. the detection method as described in as arbitrary in claim 1-5, is characterized in that: described gas chromatography-mass spectrography Analysis and Identification condition is:
Chromatographic condition: HP-5MS, 60m × 320 μm × 0.25 μm; Carrier gas: He, flow 0.8mL/min; Vapourizing temperature: 250 DEG C; Interface temperature: 280 DEG C; Chromatographic column initial temperature 50 DEG C, keeps 1min, rises to 130 DEG C with 10 DEG C/min heating rate, keeps 6min, rise to 145 DEG C with 1 DEG C/min heating rate, keep 5min, rise to 160 DEG C with 1 DEG C/min heating rate, keep 3min, rise to 220 DEG C with 3 DEG C/min heating rate, keep 1min;
Mass Spectrometry Conditions: ionization mode: EI source, energy 70eV; Ion source temperature: 230 DEG C; Quadrupole rod temperature: 150 DEG C; Mass scan range: 35 ~ 800amu; Electron-multiplier voltage: 1400V.
7. detection method as claimed in claim 6, is characterized in that, have 19 common characteristic fingerprint peakses in the finger-print generated according to the method, the retention time obtained successively is: No. 1 peak is 10.78 ± 0.006min, No. 2 peaks are: 20.42 ± 0.007min, No. 3 peaks are: 21.03 ± 0.007min, No. 4 peaks are 21.39 ± 0.007min, No. 5 peaks are: 22.56 ± 0.008min, No. 6 peaks are 23.36 ± 0.009min, No. 7 peaks are: 24.71 ± 0.014min, No. 8 peaks are: 25.23 ± 0.012min, No. 9 peaks are: 26.11 ± 0.037min, No. 10 peaks are: 27.42 ± 0.022min, No. 11 peaks are: 34.80 ± 0.014min, No. 12 peaks are: 36.09 ± 0.018min, No. 13 peaks are: 37.25 ± 0.035min, No. 14 peaks are: 37.70 ± 0.048min, No. 15 peaks are: 40.85 ± 0.262min, No. 16 peaks are: 41.19 ± 0.053min, No. 17 peaks are: 44.29 ± 0.012min, No. 18 peaks are: 44.64 ± 0.014min, No. 19 peaks are: 60.81 ± 0.008min.
8. detection method as claimed in claim 6, it is characterized in that: in the finger-print generated according to the method, have 19 common characteristic fingerprint peakses, wherein No. 1 peak is: Eucalyptol Cineole, No. 2 peaks are: δ-EIemene δ-elemene, No. 3 peaks are: α-Cubebene α-cubebene, and No. 4 peaks are:
Tetracyclo [4.4.1.1 (7,10) .0 (2,5)] dodec-3-en-11-ol, No. 5 peaks are: α-Copaene α-pepper alkene, No. 6 peaks are: β-Patchoulene β-patchoulene, and No. 7 peaks are: Cadinene cadinene, and No. 8 peaks are:
(+)-Calarene Calamus alkene, No. 9 peaks are: (-)-Aristolene aristolene, and No. 10 peaks are:
Caryophyllene cloves alkene, No. 11 peaks are: Spathulenol spainulenol, and No. 12 peaks are:
Globulol globulol, No. 13 peaks are: (-)-Spathulenol (-)-spainulenol, and No. 14 peaks are:
The different aromadendrene epoxide of Isoaromadendrene epoxide, No. 15 peaks are: Humulene epoxide 2 humulene epoxide 2, No. 16 peaks are: 1,3a-Ethano (1H) inden-4-ol,
Octahydro-2,2,4,7a-tetramethyl-, No. 17 peaks are: Widdrol widdrol, and No. 18 peaks are: Patchoulialcohol Patchoulicalcohol, and No. 19 peaks are:
4-(3,3-Dimethyl-but-1-ynyl)-4-hydroxy-2,6,6-trimethylcyclohex-2-enone。
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN105548425A (en) * | 2016-01-11 | 2016-05-04 | 山东步长制药股份有限公司 | High-performance liquid phase detection method for heart-calming granules |
| CN113125598A (en) * | 2021-04-09 | 2021-07-16 | 广州白云山光华制药股份有限公司 | Quality control method of volatile components of Xiaochaihu granules |
| CN113125598B (en) * | 2021-04-09 | 2023-02-24 | 广州白云山光华制药股份有限公司 | Quality control method for volatile components of Xiaochaihu granules |
| CN114894917A (en) * | 2022-04-06 | 2022-08-12 | 山东步长制药股份有限公司 | Detection and fingerprint spectrum construction method for volatile components of nardostachys chinensis bunge |
| CN114894917B (en) * | 2022-04-06 | 2023-10-20 | 山东步长制药股份有限公司 | Method for detecting volatile components of rhizoma nardostachyos and constructing fingerprint |
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