CN114755350A - Method for identifying astragalus mongholicus honey - Google Patents

Method for identifying astragalus mongholicus honey Download PDF

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CN114755350A
CN114755350A CN202210664107.0A CN202210664107A CN114755350A CN 114755350 A CN114755350 A CN 114755350A CN 202210664107 A CN202210664107 A CN 202210664107A CN 114755350 A CN114755350 A CN 114755350A
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honey
astragalus
calycosin
formononetin
sample
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李强强
赵彤
赵柳微
吴黎明
薛晓锋
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Institute of Apicultural Research of Chinese Academy of Agricultural Sciences
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    • G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
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    • G—PHYSICS
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    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
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    • G01N30/06—Preparation
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    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
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    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
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    • G01N30/26—Conditioning of the fluid carrier; Flow patterns
    • G01N30/28—Control of physical parameters of the fluid carrier
    • G01N30/32—Control of physical parameters of the fluid carrier of pressure or speed
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
    • G01N30/02—Column chromatography
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    • G01N30/28—Control of physical parameters of the fluid carrier
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    • G01N30/02—Column chromatography
    • G01N30/26—Conditioning of the fluid carrier; Flow patterns
    • G01N30/28—Control of physical parameters of the fluid carrier
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Abstract

The invention relates to the technical field of food detection, in particular to an identification method of astragalus mongholicus honey. The invention discovers the difference characteristic of the content of calycosin and formononetin in astragalus honey and other honey, the characteristic stably exists in astragalus honey and other honey from different years and production areas, so the calycosin and formononetin can be used as characteristic markers for identifying the astragalus honey. The method for identifying the astragalus honey provided by the invention has the characteristics of high accuracy, simplicity in operation, high efficiency and the like, and is suitable for identification and quality control of the astragalus honey in practice.

Description

Method for identifying astragalus mongholicus honey
Technical Field
The invention relates to the technical field of food detection, in particular to an identification method of astragalus mongholicus honey.
Background
Radix astragali is a Chinese medicinal plant, has effects of invigorating qi, consolidating superficial resistance, removing toxic substance, and inducing diuresis, and can be used for preventing and treating diseases such as qi deficiency, edema, chronic nephritis, and diabetes. The astragalus honey is a natural sweet substance which is prepared by fully brewing the honey collected by bees and mixed with self secretion. The astragalus honey is light amber, sweet and not greasy, has fragrant smell, is easy to crystallize, has natural slightly sour taste, and has the effects of enhancing the specific and non-specific immunologic functions of a human body, improving the physique, treating qi deficiency and the like.
The production place of astragalus honey is the centralized distribution place of astragalus, which is a honey source plant, and the astragalus prefers long sunshine and cool climate and is widely distributed in northeast, northwest and southwest areas. The flower period of the astragalus root is different due to different climates of different regions, the flower period is mostly concentrated in 6-8 months, the flower period of the astragalus root can be continued for 30-40 days, the honey flow period is about 25-35 days, each production group (apis mellifera) in one flower period can produce about 20-40 jin of astragalus root honey, and the honey yield of each production group in high yield can reach more than 60 jin. The astragalus honey belongs to rare honey species, and has relatively high nutritive value and functional activity, so that the development of an efficient astragalus honey identification method has important significance for quality control of the astragalus honey.
Disclosure of Invention
The invention aims to provide an identification method of astragalus honey, which takes calycosin and/or formononetin as characteristic markers.
In order to achieve the purpose, the invention respectively utilizes the ultra-high performance liquid phase tandem quadrupole-time of flight mass spectrum (UPLC-Q-TOF/MS) and the ultra-high performance liquid phase tandem triple quadrupole mass spectrum (UPLC-QQQ/MS) to carry out component detection on a large amount of astragalus honey and other honey samples such as acacia honey, date honey, wattle honey, linden honey and the like in different years and different producing areas, finds two characteristic peaks which are specific to the astragalus honey, and the two characteristic peaks stably exist in the astragalus honey which is sourced in different years and different producing areas, but cannot be detected in other honey. Further comparison and analysis prove that the compounds corresponding to the two characteristic peaks are calycosin and formononetin respectively.
Specifically, the invention provides the following technical scheme:
in a first aspect, the present invention provides the use of calycosin and/or formononetin as a marker characteristic of Astragalus honey.
In some embodiments of the invention, the invention provides the use of calycosin as a marker characteristic of Astragalus honey.
In some embodiments of the present invention, the present invention provides the use of formononetin as a marker characteristic of astragalus honey.
In some embodiments of the invention, the invention provides the use of calycosin and formononetin as markers characteristic of Astragalus honey.
In a second aspect, the invention provides the use of calycosin and/or formononetin in the identification of astragalus honey.
In some embodiments of the present invention, the present invention provides the use of calycosin for identifying Astragalus honey.
In some embodiments of the invention, the invention provides application of formononetin in identification of astragalus honey.
In some embodiments of the invention, the invention provides the use of calycosin and formononetin in the identification of Astragalus honey.
In a third aspect, the invention provides the use of calycosin and/or formononetin to detect the authenticity of astragalus honey.
In some embodiments of the invention, the invention provides the use of calycosin in detecting the authenticity of Astragalus honey.
In some embodiments of the present invention, the present invention provides the use of formononetin for detecting the authenticity of astragalus honey.
In some embodiments of the invention, the invention provides the use of calycosin and formononetin for detecting the authenticity of astragalus honey.
The authenticity detection is to distinguish real astragalus honey and adulterated astragalus honey, wherein the adulterated astragalus honey is prepared by adding other substances except astragalus honey, calycosin and formononetin into the astragalus honey; the content of calycosin and formononetin in the adulterated astragalus honey is obviously reduced, so that whether the astragalus honey is adulterated or not can be judged by detecting the content of calycosin and formononetin, and the authenticity detection of the astragalus honey is carried out.
In a fourth aspect, the invention provides an astragalus honey identification method, which detects a honey sample by taking calycosin and/or formononetin as characteristic markers, and judges that the honey sample is astragalus honey if the content of calycosin in the honey sample is not lower than 20 μ g/kg and/or the content of formononetin is not lower than 20 μ g/kg; if the content of calycosin in the honey sample is less than 20 mug/kg and/or the content of formononetin in the honey sample is less than 20 mug/kg, the honey sample is judged to be honey other than astragalus honey or adulterated astragalus honey.
In some embodiments of the invention, the method detects the honey sample by taking calycosin as a characteristic marker, and if the content of the calycosin in the honey sample is not lower than 20 mug/kg, the honey sample is judged to be astragalus honey; if the content of calycosin in the honey sample is less than 20 mug/kg, judging that the honey sample is honey other than astragalus honey or adulterated astragalus honey.
In some embodiments of the invention, the method detects the honey sample by taking formononetin as a characteristic marker, and if the content of formononetin in the honey sample is not lower than 20 mug/kg, the honey sample is judged to be astragalus honey; if the content of formononetin in the honey sample is less than 20 mug/kg, judging that the honey sample is other honey except astragalus honey or adulterated astragalus honey.
In some embodiments of the invention, the method detects the honey sample by taking calycosin and formononetin as characteristic markers, and if the content of calycosin and formononetin in the honey sample is not lower than 20 mug/kg, the honey sample is judged to be astragalus honey; if the content of calycosin and formononetin in the honey sample is less than 20 mug/kg, judging that the honey sample is honey other than astragalus honey or adulterated astragalus honey.
Preferably, when calycosin is taken as the characteristic marker, if the content of calycosin in the honey sample is 20-170 μ g/kg, the honey sample is judged to be astragalus honey, and if the content of calycosin in the honey sample is less than 20 μ g/kg, the honey sample is judged to be honey other than astragalus honey or adulterated astragalus honey.
Further preferably, if the content of calycosin in the honey sample is less than the detection Limit (LOD), the honey sample is judged to be honey other than astragalus honey.
When the formononetin is used as a characteristic marker, if the content of the formononetin in the honey sample is 20-40 mug/kg, the honey sample is judged to be astragalus honey, and if the content of the formononetin in the honey sample is less than 20 mug/kg, the honey sample is judged to be other honey except the astragalus honey or adulterated astragalus honey.
Further preferably, if the content of formononetin in the honey sample is less than a detection Limit (LOD), the honey sample is judged to be honey other than astragalus honey.
The honey other than the astragalus honey as described above includes, but is not limited to, acacia honey, date flower honey, wattle honey, linden honey.
In the above identification method, preferably, the honey sample is detected by ultra-high performance liquid tandem triple quadrupole mass spectrometry.
Preferably, the liquid phase conditions of the ultra-high performance liquid phase tandem triple quadrupole mass spectrometry are as follows: adopting a C18 chromatographic column, taking 0.1% formic acid water solution as a mobile phase A, and taking acetonitrile as a mobile phase B; separation was performed using a gradient elution procedure: 0min, 95% mobile phase a; 1min, 95% mobile phase a; 2min, 65% mobile phase a; 3min, 50% mobile phase A; 5min, 25% mobile phase A; 5.1min, 0% mobile phase A; 7min, 0% mobile phase A, 3min post run, 95% mobile phase A.
Further preferably, the column is ZORBAX Eclipse Plus C18 column (2.1X 100 mm, 1.8 μm).
In the liquid phase condition of the ultra-high performance liquid phase tandem triple quadrupole mass spectrometry detection, the column temperature is 39-41 ℃ (preferably 40 ℃); the flow rate of the mobile phase is 0.25 to 0.4 mL/min (preferably 0.3 mL/min).
In the liquid phase condition of the ultra-high performance liquid phase tandem triple quadrupole mass spectrometry, the sampling amount is preferably 2 muL.
Preferably, the mass spectrum conditions of the ultra-high performance liquid phase tandem triple quadrupole mass spectrometry are as follows: ion source mode: ESI, positive ion MRM mode; temperature of the drying gas: 200 ℃; flow rate of drying gas: 15L/min; temperature of sheath gas: at 330 ℃; flow rate of sheath gas: 12L/min; atomizer pressure: 45 psi; capillary voltage: 3500V; fragmentation voltage: 380V.
When the detection condition of the ultra-high performance liquid phase series triple quadrupole is adopted, the retention time of calycosin is 3.8 min, and the m/z of quantitative ion pairs is 270.0 and 285.0; the retention time of formononetin is 4.5 min, and the quantitative ion pair m/z is 197.0 and 269.1.
In the above identification method, before the detection of the ultra high performance liquid chromatography-tandem triple quadrupole mass spectrometry, the method further comprises a step of pretreating a honey sample, wherein the pretreatment comprises: dissolving the honey sample with water, performing ultrasonic treatment at 50-70kHz power for 5-15 min, performing solid-liquid separation to obtain a honey sample solution, passing the honey sample solution through a solid phase extraction column, eluting the solid phase extraction column with water after the honey sample solution completely flows out, draining, eluting with methanol, collecting the eluate, drying, and re-dissolving with methanol.
The invention has the beneficial effects that: the invention discovers the difference characteristic of the content of calycosin and formononetin in astragalus honey and other honey for the first time, and the characteristic stably exists in astragalus honey and other honey from different years and production places, so that calycosin and formononetin can be used as characteristic markers for identifying the astragalus honey. The method for identifying the astragalus honey provided by the invention has the characteristics of high accuracy, simplicity in operation, high efficiency and the like, and is suitable for identification and quality control of the astragalus honey in practice.
Drawings
FIG. 1 is a UPLC-Q-TOF/MS analysis characteristic ion 285.0752 Extraction Ion Chromatography (EIC) chart of a calycosin standard substance according to example 1 of the present invention.
FIG. 2 is a two-stage scanning mass spectrum of characteristic ion 285.0752 of calycosin standard in example 1 of the present invention.
FIG. 3 is a UPLC-Q-TOF/MS analysis characteristic ion 269.0802 Extraction Ion Chromatography (EIC) diagram of formononetin standard in example 1 of the present invention.
FIG. 4 is a two-stage scanning mass spectrum of characteristic ion 269.0802 of formononetin standard in example 1 of the present invention.
FIG. 5 is a UPLC-Q-TOF/MS analysis characteristic ion 285.0752 Extraction Ion Chromatography (EIC) chart of the astragalus honey extract in example 1 of the present invention.
FIG. 6 is a two-stage scanning mass spectrum of characteristic ion 285.0752 of Astragalus honey extract in example 1 of the present invention.
FIG. 7 is a UPLC-Q-TOF/MS analysis characteristic ion 269.0802 Extraction Ion Chromatography (EIC) chart of the astragalus honey extract in example 1 of the present invention.
FIG. 8 is a two-stage scanning mass spectrum of characteristic ion 269.0802 of Astragalus honey extract in example 1 of the present invention.
FIG. 9 is a UPLC-Q-TOF/MS analysis characteristic ion 285.0752 Extraction Ion Chromatogram (EIC) diagram of Vitex negundo extract in example 1 of the present invention.
FIG. 10 is a UPLC-Q-TOF/MS analysis characteristic ion 269.0802 Extraction Ion Chromatogram (EIC) of Vitex negundo extract in example 1 of the invention.
FIG. 11 is a UPLC-Q-TOF/MS analysis characteristic ion 285.0752 Extraction Ion Chromatography (EIC) chart of acacia honey extract in example 1 of the present invention.
FIG. 12 is a UPLC-Q-TOF/MS analysis characteristic ion 269.0802 Extraction Ion Chromatography (EIC) chart of acacia honey extract in example 1 of the present invention.
FIG. 13 is a UPLC-Q-TOF/MS analysis characteristic ion 285.0752 Extraction Ion Chromatogram (EIC) diagram of the jujube flower honey extract in example 1 of the present invention.
FIG. 14 is a UPLC-Q-TOF/MS analysis characteristic ion 269.0802 Extraction Ion Chromatogram (EIC) diagram of the jujube flower honey extract in example 1 of the present invention.
FIG. 15 is a UPLC-Q-TOF/MS analysis characteristic ion 285.0752 Extraction Ion Chromatography (EIC) diagram of the linden honey extract in example 1 of the present invention.
FIG. 16 is a UPLC-Q-TOF/MS analysis characteristic ion 269.0802 Extraction Ion Chromatography (EIC) diagram of the linden honey extract in example 1 of the present invention.
Detailed Description
The invention provides a method for identifying whether a honey sample is astragalus honey by using calycosin and formononetin as characteristic markers, which adopts UPLC-Q-TOF/MS (ultra high performance liquid phase tandem quadrupole-time of flight mass spectrum) and UPLC-QQQ/MS (ultra high performance liquid phase tandem triple quadrupole mass spectrum) to qualitatively and quantitatively detect the honey sample respectively, the obtained spectrogram contains characteristic peaks corresponding to calycosin and formononetin, and when the content ranges of the calycosin and the formononetin are respectively 20-170 mu g/kg and 20-40 mu g/kg, the honey sample is judged to be astragalus honey; when the honey sample does not contain characteristic peaks corresponding to calycosin and formononetin, the honey sample is judged to be honey other than astragalus honey or adulterated astragalus honey.
The following examples are intended to illustrate the invention but are not intended to limit the scope of the invention.
Example 1 identification of Astragalus honey with Calycosin and formononetin as characteristic markers
Astragalus honey, acacia honey, date honey, vitex honey and linden honey are directly collected from a bee field in different seasons and years across the country, and the source information of part of honey samples is shown in table 1. The honey samples are used for detecting the content of calycosin and formononetin in different honey samples.
TABLE 1 Honey sample Source information
Figure 870160DEST_PATH_IMAGE001
1. Pretreatment of honey samples
Taking 5g of honey sample, adding 10ml of deionized water, shaking for dissolution, performing ultrasonic treatment at 60kHz power for 10min, and performing solid-liquid separation to obtain a honey sample solution. Placing an Oasis HLB solid-phase extraction column on a solid-phase extraction device, sequentially activating the Oasis HLB solid-phase extraction column by using 5 mL of methanol and 5 mL of pure water, placing a honey sample solution on the Oasis HLB solid-phase extraction column, adjusting the flow rate to be 1-1.5 mL/min, rinsing the Oasis HLB solid-phase extraction column by using 10mL of pure water after the sample solution completely flows out, and draining under negative pressure. Eluting with 8 mL of methanol, collecting the eluent, and blowing the eluent with nitrogen at normal temperature. Redissolving with 1ml of methanol, filtering with a 0.22 mu m nylon filter membrane, and carrying out subsequent chromatographic detection.
2. Detection method
2.1 reagents
Acetonitrile: chromatographically pure, available from Fisher Scientific;
formic acid: purchased from Sigma.
2.2 instruments
The Agilent 1290-6545 ultrahigh-performance liquid-phase serial quadrupole-time-of-flight mass spectrometry (UPLC-Q-TOF/MS) combined system is used for qualitative analysis of compounds, the Agilent 1290-6495 ultrahigh-performance liquid-phase serial triple quadrupole mass spectrometry (UPLC-QQQ/MS) combined system is used for quantitative analysis of compounds, and an Agilent Master workstation is adopted for data processing.
2.3 UPLC-Q-TOF/MS Instrument conditions
2.3.1 liquid chromatography conditions
A chromatographic column: ZORBAX Eclipse Plus C18 column (2.1X 100 mm, 1.8 μm);
column temperature: 50 ℃;
sample introduction amount: 2 muL;
mobile phase: A) 0.1% aqueous formic acid; b) 0.1% formic acid acetonitrile solution;
flow rate: 0.3 mL/min;
the gradient elution procedure is shown in table 2.
TABLE 2 gradient elution procedure
Figure 866934DEST_PATH_IMAGE002
2.3.2 Mass Spectrometry conditions
Ion source mode: ESI, positive ion mode;
temperature of the drying gas: 325 ℃;
flow rate of drying gas: 10L/min;
temperature of sheath gas: 370 ℃;
flow rate of sheath gas: 12L/min;
atomizer pressure: 35 psi;
capillary voltage: 3500V;
fragmentation voltage: 135V;
an acquisition mode: full Scan and Auto MS/MS (20 eV collision energy);
2.4 UPLC-QqQ-MS/MS Instrument conditions
2.4.1 conditions of liquid chromatography
And (3) chromatographic column: ZORBAX Eclipse Plus C18 column (2.1X 100 mm, 1.8 μm);
column temperature: at 40 ℃;
sample injection amount: 2 muL;
mobile phase: A) 0.1% aqueous formic acid; b) Acetonitrile;
flow rate: 0.3 mL/min;
the gradient elution procedure is shown in table 3.
TABLE 3 gradient elution procedure
Figure 673348DEST_PATH_IMAGE003
2.4.2 Mass Spectrometry conditions
Ion source mode: ESI, positive ion MRM mode;
temperature of the drying gas: 200 ℃;
flow rate of drying gas: 15L/min;
temperature of sheath gas: at 330 ℃;
flow rate of sheath gas: 12L/min;
atomizer pressure: 45 psi;
capillary voltage: 3500V;
fragmentation voltage: 380V.
The retention time, MS/MS information, linear range, detection limit, and quantitation limit of calycosin and formononetin in UPLC-QqQ-MS/MS detection are shown in Table 4.
TABLE 4 Retention time, MS/MS information, Linear Range, detection Limit and quantitation Limit of target Compounds in UPLC-QqQ-MS/MS detection
Figure 506306DEST_PATH_IMAGE004
Note: in table 4,. denotes a quantitative ion; a: the detection limit is 3 times the signal-to-noise ratio; b: the limit of quantitation is 10 times the signal-to-noise ratio.
3. Results of the experiment
3.1 qualitative analysis results
3.1.1 extraction of chromatographic peak with accurate mass number of 285.0752 in TIC, extraction in TIC chart of radix astragali Mel extract and calycosin standard solution, retention time is 9.87 min.
The chromatographic peak with the accurate mass number of 269.0802 is extracted from TIC, and can be extracted from TIC chart of radix astragali Mel extract and formononetin standard solution, and the retention time is 11.83 min.
3.1.2 parent ion 285.0752 of the compound calycosin produced daughter ions 213.0540 and 270.0512 in a second mass spectrometric assay. The two-stage mass spectrometric detection of the astragalus honey extract and the calycosin standard solution both contain 213.0540 and 270.0512 daughter ions.
The parent ion 269.0802 of the compound formononetin can generate daughter ions 197.0596 and 237.0543 in secondary mass spectrometric detection. The two-stage mass spectrometric detection of the astragalus honey extract and the formononetin standard solution both contained 197.0596 and 237.0543 daughter ions.
The mass error (mass error) was less than 5 ppm for the extracted mass numbers of 3.1.1 and 3.1.2 above.
The results show that astragalus honey contains calycosin and formononetin.
3.2 quantitative analysis results
3.2.1 use of UPLC-QqQ-MS/MS to establish matrix standard curve, linear range, detection limit and quantitative limit of calycosin and formononetin
The calycosin retention time is 3.8 min, the quantitative ion pair is 270.0/285.0, the linear range of the substrate standard curve is 12-200 μ g/kg, R 2=0.9987, detection limit is 4 mug/kg, quantification limit is 12 mug/kg;
the retention time of formononetin is 4.5 min,the quantitative ion pair is 197.0/269.1, the linear range of the substrate standard curve is 12-200 mug/kg, R2=0.9992, detection limit 4 μ g/kg, quantification limit 12 μ g/kg.
3.2.2 method recovery and reproducibility validation
Calycosin was added to the blank honey samples at 20. mu.g/kg, 50. mu.g/kg, 500. mu.g/kg, respectively, for 5 replicates. The recovery rate of calycosin obtained by the quantitative method established based on UPLC-QqQ-MS/MS is 65.6-105.0%, the relative standard deviation is 3.1-9.3% (Table 5), and the method has the recovery rate and reproducibility meeting the quantitative requirements and can be used for real sample detection.
Formononetin of 20. mu.g/kg, 50. mu.g/kg, 500. mu.g/kg was added to the blank honey samples, respectively, setting 5 replicates. The recovery rate of formononetin obtained by the detection of a quantitative method established based on UPLC-QqQ-MS/MS is 78.5-93.0%, the relative standard deviation is 2.8-11.4% (Table 5), the recovery rate and the reproducibility of the method meet the quantitative requirements, and the method can be used for real sample detection.
TABLE 5 test recovery and Relative Standard Deviation (RSD) of target compounds added to a sample of honey blank
Figure 305634DEST_PATH_IMAGE005
3.2.3 Calycosin and formononetin content in Astragalus honey
Detecting the content of calycosin and formononetin in the astragalus honey to obtain 20-170 mu g/kg of calycosin and 20-40 mu g/kg of formononetin in the astragalus honey.
3.2.4 Calycosin and formononetin content in other Mel
Calycosin and formononetin are not detected in acacia honey, date flower honey, wattle honey and linden honey. However, the detected contents of the two in the astragalus honey are 20-170 mu g/kg and 20-40 mu g/kg respectively. Therefore, calycosin and formononetin can be used as characteristic markers of astragalus honey.
A summary of the test results for the above honey samples is shown in Table 6.
TABLE 6 actual honey sample test results
Figure 907648DEST_PATH_IMAGE006
Specific test results of the partial astragalus honey samples shown in table 1 are shown in table 7.
TABLE 7 results of determination of content of calycosin and formononetin in part of radix astragali honey samples
Figure 399809DEST_PATH_IMAGE007
In the above test, the test results of calycosin standard are shown in FIGS. 1 and 2, and the test results of formononetin standard are shown in FIGS. 3 and 4. The detection results of one astragalus honey sample are shown in fig. 5, 6, 7 and 8, the detection results of one vitex honey sample are shown in fig. 9 and 10, the detection results of acacia honey are shown in fig. 11 and 12, the detection results of date honey are shown in fig. 13 and 14, and the detection results of linden honey are shown in fig. 15 and 16.
According to the detection result, if the content of calycosin in the honey sample is not lower than 20 mug/kg, the honey sample is judged to be astragalus honey. If the content of formononetin in the honey sample is not lower than 20 mug/kg, the honey sample is judged to be astragalus honey.
Although the invention has been described in detail hereinabove with respect to a general description and specific embodiments thereof, it will be apparent to those skilled in the art that modifications or improvements may be made thereto based on the invention. Accordingly, such modifications and improvements are intended to be within the scope of the invention as claimed.

Claims (10)

1. Application of calycosin and/or formononetin as radix astragali honey characteristic marker is provided.
2. Application of calycosin and/or formononetin in identification of radix astragali Mel is provided.
3. Application of calycosin and/or formononetin in detecting authenticity of radix astragali Mel is provided.
4. An identification method of astragalus honey is characterized in that calycosin and/or formononetin is used as a characteristic marker to detect a honey sample, and if the content of calycosin in the honey sample is not lower than 20 mug/kg and/or the content of formononetin is not lower than 20 mug/kg, the honey sample is judged to be astragalus honey; if the content of calycosin in the honey sample is less than 20 mug/kg and/or the content of formononetin in the honey sample is less than 20 mug/kg, the honey sample is judged to be honey other than astragalus honey or adulterated astragalus honey.
5. The method for identifying the astragalus honey as claimed in claim 4, wherein the honey sample is detected by ultra high performance liquid tandem triple quadrupole mass spectrometry.
6. The method for identifying the astragalus honey as claimed in claim 5, wherein the liquid phase conditions of the ultra high performance liquid tandem triple quadrupole mass spectrometry are as follows: adopting a C18 chromatographic column, taking 0.1% formic acid aqueous solution as a mobile phase A, and taking acetonitrile as a mobile phase B; separation was performed using a gradient elution procedure: 0min, 95% mobile phase a; 1min, 95% mobile phase a; 2min, 65% mobile phase a; 3min, 50% mobile phase A; 5min, 25% mobile phase A; 5.1min, 0% mobile phase A; 7min, 0% mobile phase A; after 3min run, 95% mobile phase a.
7. The method for identifying astragalus honey as claimed in claim 6, wherein the column temperature is 39-41 ℃ and/or the flow rate of the mobile phase is 0.25-0.4 mL/min.
8. The method for identifying the astragalus honey as claimed in claim 7, wherein the mass spectrum conditions of the ultra high performance liquid tandem triple quadrupole mass spectrometry are as follows: ion source mode: ESI, positive ion MRM mode; temperature of the drying gas: 200 ℃; flow rate of drying gas: 15L/min; temperature of sheath gas: at 330 ℃; flow rate of sheath gas: 12L/min; atomizer pressure: 45 psi; capillary voltage: 3500V; fragmentation voltage: 380V.
9. The method for discriminating milkvetch root honey according to claim 8, wherein the holding time of calycosin is 3.8 min, and the quantitative ion pair m/z is 270.0 and 285.0;
and/or formononetin has a retention time of 4.5 min, and quantitative ion pair m/z of 197.0 and 269.1.
10. The method for identifying the astragalus honey as claimed in any one of claims 5 to 9, further comprising a step of pretreating a honey sample before ultra high performance liquid tandem triple quadrupole mass spectrometry detection, wherein the pretreatment comprises: dissolving the honey sample with water, performing ultrasonic treatment at 50-70kHz power for 5-15 min, performing solid-liquid separation to obtain a honey sample solution, passing the honey sample solution through a solid phase extraction column, eluting the solid phase extraction column with water after the honey sample solution completely flows out, draining, eluting with methanol, collecting the eluate, drying, and re-dissolving with methanol.
CN202210664107.0A 2022-06-14 2022-06-14 Method for identifying astragalus mongholicus honey Pending CN114755350A (en)

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