CN104090059A - Method for simultaneously measuring eight components in edible flavors and fragrances - Google Patents
Method for simultaneously measuring eight components in edible flavors and fragrances Download PDFInfo
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Abstract
The invention discloses a method for simultaneously measuring eight components in edible flavors and fragrances. The method comprises the steps of preparing an extraction solution, preparing a standard solution, preparing a sample solution, carrying out ultra-high performance liquid chromatography-tandem mass spectrometry and calculating the content. A simple, quick and high-flux measurement method is established by optimizing a pretreatment condition, a liquid chromatography condition and a mass spectrum condition of the method for measuring the eight components in the edible flavors and fragrances. The method for simultaneously measuring the eight components in the edible flavors and fragrances has the advantages of simplicity in operation, high sensitivity, high recycling rate, high reproducibility, short detection time, low solvent consumption, high detection flux and high detection efficiency.
Description
technical field:
The invention belongs to physical and chemical inspection technical field, be specifically related to eight kinds of composition thiocarbamide, sesamol, dehydroactic acid, P-ethoxyphellylurea, ortho-aminobenzoic acid cinnamic ester, nordihydroguaiaretic acid, aristolochic acid A and sorbic acid methods for measuring simultaneously in a kind of flavoring essence spices.
background technology:
Thiocarbamide is thio urea, is a kind of important industrial chemicals, is mainly used in organic synthesis, and textile industry etc., because it has certain bleaching function, also can be applicable to the bleaching of textile.Report, contact thiocarbamide may cause the diseases such as thyroid gland enlargement, and thiocarbamide is also irritant to the upper respiratory tract, eye, intestines and stomach, and skin is had to infringement, and national regulation strictly forbids thiocarbamide to add in food.The assay method of thiocarbamide mainly contains liquid phase chromatography, photometry, Liquid Chromatography-Tandem Mass Spectrometry and electrochemical process etc.
Sesamol is aromatic series phenolic compound, natural being present in sesame oil, there is oxidation resistant effect, it is the important stay in grade agent of sesame oil, this material has stronger radical scavenging activity, is widely used in preparing hair conditioner of antimildew and antibacterial agent, anticancer, anti-hypertension medicine, high-grade shampoo etc.Based on this material cause tumour possibility, 42 kinds of food additives names that in forbidding is listed sesamol by Japanese MHLW in 2006 are single.In GB GB2760-2011 " food additives use hygienic standard ", also sesamol is not listed in to the food additives list of permission to use.The detection method of the sesamol of research report mainly concentrates on By Gas Chromatography-mass Spectrometry, Ultra Performance Liquid Chromatography method etc.
Dehydroactic acid all has certain inhibiting effect to mould, saccharomycete, spoilage organisms.In food, be used as antiseptic, mildewproof agent, for the food antiseptic taking carbohydrates as principal ingredient, be mainly used in pickles, fermented bean curd, fruit juice, cake etc.Dehydroactic acid is acid type antiseptic, and its effect is determined with pH value, and the more potent fruit of acidity is better, and nearly unavailable under alkali condition.Dehydroactic acid can be absorbed by human body rapidly, is scattered in blood plasma and many organs after entering human body, has multiple oxidasic effect in the body of inhibition.At present, the European Community clearly bans use of, and Japan also limits its use, and China allows it to use within the specific limits, but maximum permission use amount is 300 μ g/mL.The detection method of dehydroactic acid mainly contains vapor-phase chromatography, high performance liquid chromatography etc.
Dulcin, formal name used at school is P-ethoxyphellylurea, is a kind of artificial synthesis edulcorant, its sugariness is equivalent to 200 times of sucrose.Dulcin has carcinogenicity, thereby is listed in banned substance.The assay method of dulcin mainly contains liquid phase chromatography, Liquid Chromatography/Mass Spectrometry, capillary electrophoresis etc.
Ortho-aminobenzoic acid cinnamic ester can be used as flavoring additives or flavouring agent, thereby is used to the industry such as cosmetics, food, and in finally adding Fragrant food, concentration is about 1.7-730mg/Kg.Because ortho-aminobenzoic acid cinnamic ester is a kind of sintetics, some researchs show that this material has carcinogenicity, human body is had to larger harm, thereby be prohibited from using in multiple countries.1940 start, ortho-aminobenzoic acid cinnamic ester is the food for grape or cherry flavor as a kind of synthetic flavoring additives, and obtain food and drug administration and ratify as food additives, European Union is also using ortho-aminobenzoic acid cinnamic ester as artificial fragrance matter, and suggestion working concentration is 25mg/Kg.1980, the research of National Cancer Institute of Britain found that high concentration ground makes an addition to ortho-aminobenzoic acid cinnamic ester in food and causes liver cell generation canceration in hybridize mice body, and therefore the Something English Committee of Experts is forbidden being applied to food industry.Subsequently, food and drug administration also classifies ortho-aminobenzoic acid cinnamic ester as ban use of in food adjuvant.The detection method of ortho-aminobenzoic acid cinnamic ester mainly contains high performance liquid chromatography etc.
Nordihydroguaiaretic acid (NDGA) is antioxidant, is a kind of natural products extracting from evergreen shrub, has multiple biologically active, comprises lipoxidase, papillomavirus, herpes simplex virus and the hypoglycemic effect etc. of suppressing.In recent years NDGA tumor vessel generates inhibiting discovery, and has the generation of kinds of tumors of inhibition and the effect of development, the especially NDGA therapeutic action to people's Malignant cerebral gliomas, for treatment tumour, cancer provide new approach.High-efficient solid optionally synthetic NDGA, for Development of Novel is treated tumour, cancer new drug provides material base, is carried out the R&D work of PTS.The assay method of nordihydroguaiaretic acid mainly contains high liquid chromatography method, liquid chromatography-mass spectrography etc.
Aristolochic acid is the luxuriant and rich with fragrance compounds of nitro, has the body of raising antibacterial ability, and the effect that improves cellular immune function, is prevalent in Aristolochiaceae birthwort.Such plant is widely used for medicine preparation in states such as China, India.In May, 2000 FDA (Food and Drug Adminstration) (FDA) has forbidden comprising homonym, the relevant herbal medicine of different name jljl kind and import and the sale of Chinese patent drug such as caulis aristologhiae manshuriensis, birthwort, dutchmanspipe root, herba aristolochiae, Aristolochia fangchi, Cocculus trilobus.Subsequently, the country such as Britain, Canada, Australia, New Zealand and Malaysia has also taked same measure.In 6 kinds of aristolochic acids finding at nature, the toxicity of aristolochic acid A is the strongest.The detection method of bibliographical information Aristolochic Acid material is existing a lot, mainly contains thin-layered chromatography, high performance liquid chromatography (HPLC), Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS) etc.
Sorbic acid is that China allows the food preservative using, and its chemical name is 2,4-hexadienoic acid, has good stability.Because sorbic acid has good inhibiting effect to mould, saccharomycete, aerophile bacterium, and its toxicity is less, security is higher, the antiseptic of Chang Zuowei food uses, but excessive, long-term edible meeting harmful to human liver, renal function, clearly specified in standard GB/T 2760-2007 that the maximum of sorbic acid is used limitation.The antiseptic effect of sorbic acid reduces with the rising of pH value, and its pH value is generally 5.0-6.0.The assay method of Determination of sorbic mainly contains spectrophotometric method, chromatography, ultra-violet absorption spectrometry etc.
At present, open source literature has nothing to do temporarily in these eight kinds of methods that composition detects simultaneously both at home and abroad.
summary of the invention:
The object of the present invention is to provide eight kinds of composition thiocarbamide, sesamol, dehydroactic acid, P-ethoxyphellylurea, ortho-aminobenzoic acid cinnamic ester, nordihydroguaiaretic acid, aristolochic acid A and sorbic acid methods for measuring simultaneously in a kind of flavoring essence spices.
The present invention is directed to the deficiencies in the prior art, by long-term research and exploration, eight kinds of compositions (thiocarbamide, sesamol, dehydroactic acid, P-ethoxyphellylurea, ortho-aminobenzoic acid cinnamic ester, nordihydroguaiaretic acid, aristolochic acid A and sorbic acid) method for measuring proposed a kind of flavoring essence spices on the basis of method contrast, optimization in, for detection of thiocarbamide, sesamol, dehydroactic acid, P-ethoxyphellylurea, ortho-aminobenzoic acid cinnamic ester, nordihydroguaiaretic acid, aristolochic acid A and sorbic acid in flavoring essence spices.The present invention has investigated Pretreatment, liquid phase chromatogram condition and the mass spectrum condition of thiocarbamide, sesamol, dehydroactic acid, P-ethoxyphellylurea, ortho-aminobenzoic acid cinnamic ester, nordihydroguaiaretic acid, aristolochic acid A and sorbic acid in flavoring essence spices, for eight kinds of compositions in easy, quick, high-throughout mensuration flavoring essence spices provide technical support.
Technical scheme of the present invention is: described assay method comprises the following steps:
The preparation of A, extraction solution: the methanol-water solution of getting the standby volume ratio 7:3 of one-tenth of Methanol is extraction solution;
The preparation of B, standard solution: with the standard solution of preparing respectively thiocarbamide, sesamol, dehydroactic acid, P-ethoxyphellylurea, ortho-aminobenzoic acid cinnamic ester, nordihydroguaiaretic acid, aristolochic acid A and sorbic acid containing interior target extraction solution;
The preparation of C, sample solution: use containing interior target extraction solution extraction flavoring essence spices sample, excessively obtain sample solution after organic phase filter membrane;
D, the analysis of Ultra Performance Liquid Chromatography-tandem mass spectrometer: utilize Ultra Performance Liquid Chromatography-tandem mass spectrometer respectively standard solution and sample solution to be detected to analysis;
The calculating of eight kinds of composition thiocarbamides, sesamol, dehydroactic acid, P-ethoxyphellylurea, ortho-aminobenzoic acid cinnamic ester, nordihydroguaiaretic acid, aristolochic acid A and Determination of sorbics in E, flavoring essence spices.
The preparation of standard solution described in B step is: take respectively thiocarbamide, sesamol, dehydroactic acid, P-ethoxyphellylurea, ortho-aminobenzoic acid cinnamic ester, nordihydroguaiaretic acid, aristolochic acid A and the each 20mg of sorbic acid, after dissolving with methyl alcohol, be transferred in the brown volumetric flask of 100mL, be settled to scale, obtain respectively single standard storing solution; Take salicylic acid-d4 10mg, after dissolving, be transferred in the brown volumetric flask of 50mL with methyl alcohol, methanol constant volume, to scale, obtains interior mark storing solution; Get respectively in the brown volumetric flask of single standard storing solution 0.1-1mL to 50mL, to scale, obtain one-level mixed standard solution by methanol constant volume; Get in the brown volumetric flask of interior mark storing solution 1mL to 50mL, to scale, obtain mark standard solution in one-level by methanol constant volume; Accurately pipette respectively the one-level mixed standard solution of 0.1mL, 0.5mL, 1mL, 2mL, 5mL to the brown volumetric flask of 50mL, each brown volumetric flask all adds mark standard solution in 2mL one-level, is settled to scale with extraction solution, obtains series standard solution.
Get in 0.8mL mark storing solution to the brown volumetric flask of 1L, be settled to scale with extraction solution, obtain containing interior target extraction solution.Described is salicylic acid-d4 containing the internal standard compound in interior target extraction solution.
Described in C step, the preparation method of sample solution is: take 0.5g sample in 50mL tool plug conical flask, accurately add 25mL to contain interior target extraction solution, with 150r/min oscillation extraction 20min, get supernatant and cross organic phase filter membrane, filtrate is sample solution.Described organic phase filter membrane is 0.22 μ m polyethersulfone filter membrane.The oscillation frequency of described oscillation extraction is 120-180r/min, and extraction time is 10-30min.
Chromatographiccondition described in D step is chromatographic column adopting C18,1.7 μ m, 2.1 × 100mm chromatographic column; Mobile phase is methyl alcohol A and 0.05% aqueous formic acid B; Flow velocity: 0.4mL/min; Sample size: 5 μ L; Column temperature: 40 DEG C; Sample temperature: 10 DEG C; Adopt gradient elution program, initial A be 10%, B while being 90%, 1min A be that 35%, B is 65%, when 4min A be 65%, B while being 35%, 6min initial A be 90%, B while being 10%, 7min A be 90%, when B is 10%, 8min A be 10%, B while being 90%, 10min A be that 10%, B is 90%; Described mass spectrum condition is: electric spray ion source (ESI) positive ion mode, scan mode is multiple-reaction monitoring pattern, electron spray voltage is 5500V, 600 DEG C of ion source temperatures, gas curtain gas: 25Psi, atomization gas: 55Psi, auxiliary heating gas: 50Psi, the quota ion of thiocarbamide, sesamol, dehydroactic acid, P-ethoxyphellylurea, ortho-aminobenzoic acid cinnamic ester, nordihydroguaiaretic acid, aristolochic acid A, sorbic acid and salicylic acid-d4 is to being respectively: m/z 60.0,109.1,127.1,108.0,117.1,193.2,265.1,95.1,97.1.Other mass spectrum parameter is in table 1.
The calculating of content described in E step is: first carry out regretional analysis with the chromatographic peak area of the detected object of standard solution and internal standard compound than corresponding its respective concentration ratio and obtain typical curve linear equation, the chromatographic peak area of the object then sample solution being detected, substitution typical curve linear equation, obtains the content of thiocarbamide in sample, sesamol, dehydroactic acid, P-ethoxyphellylurea, ortho-aminobenzoic acid cinnamic ester, nordihydroguaiaretic acid, aristolochic acid A, sorbic acid.
The present invention is directed to the deficiencies in the prior art, by the optimization of Pretreatment, chromatographic condition and mass spectrum condition to thiocarbamide, sesamol, dehydroactic acid, P-ethoxyphellylurea, ortho-aminobenzoic acid cinnamic ester, nordihydroguaiaretic acid, aristolochic acid A and sorbic acid in flavoring essence spices, set up a kind of easy, quick, high-throughout assay method, for the detection of eight kinds of compositions in flavoring essence spices provides technical support.That the present invention has advantages of is simple to operate, highly sensitive, the recovery is good, favorable reproducibility and detection time is short, solvent consumption is little, detect that flux is high, detection efficiency is high, has very important significance for the quality safety of supervision flavoring essence spices.
Brief description of the drawings
Fig. 1 is the process flow diagram of assay method of the present invention.
Fig. 2 is typical curve linear equation and the linear regression coeffficient coordinate diagram of thiocarbamide.
Fig. 3 is typical curve linear equation and the linear regression coeffficient coordinate diagram of sesamol.
Fig. 4 is typical curve linear equation and the linear regression coeffficient coordinate diagram of dehydroactic acid.
Fig. 5 is typical curve linear equation and the linear regression coeffficient coordinate diagram of P-ethoxyphellylurea.
Fig. 6 is typical curve linear equation and the linear regression coeffficient coordinate diagram of ortho-aminobenzoic acid cinnamic ester.
Fig. 7 is typical curve linear equation and the linear regression coeffficient coordinate diagram of nordihydroguaiaretic acid.
Fig. 8 is typical curve linear equation and the linear regression coeffficient coordinate diagram of aristolochic acid A.
Fig. 9 is typical curve linear equation and the linear regression coeffficient coordinate diagram of sorbic acid.
Figure 10 is the chromatogram of thiocarbamide in standard solution, sesamol, dehydroactic acid, P-ethoxyphellylurea, ortho-aminobenzoic acid cinnamic ester, nordihydroguaiaretic acid, aristolochic acid A and sorbic acid.
Embodiment
Below in conjunction with embodiment and accompanying drawing, the present invention is further illustrated, but never in any form the present invention is limited, and any conversion or the replacement done based on training centre of the present invention, all belong to protection scope of the present invention.
Embodiment 1:
Eight kinds of composition methods for measuring simultaneously in flavoring essence spices of the present invention, comprise the following steps:
(1) preparation of extraction solution: the methanol-water solution of getting the standby volume ratio 7:3 of one-tenth of Methanol is extraction solution;
(2) preparation of standard solution: with the standard solution of preparing respectively thiocarbamide, sesamol, dehydroactic acid, P-ethoxyphellylurea, ortho-aminobenzoic acid cinnamic ester, nordihydroguaiaretic acid, aristolochic acid A and sorbic acid containing interior target extraction solution;
(3) preparation of sample solution: use containing interior target extraction solution extraction flavoring essence spices sample, excessively obtain sample solution after organic phase filter membrane;
(4) Ultra Performance Liquid Chromatography-tandem mass spectrometer analysis: utilize Ultra Performance Liquid Chromatography-tandem mass spectrometer respectively standard solution and sample solution to be detected to analysis;
(5) calculating of eight kinds of composition thiocarbamides, sesamol, dehydroactic acid, P-ethoxyphellylurea, ortho-aminobenzoic acid cinnamic ester, nordihydroguaiaretic acid, aristolochic acid A and Determination of sorbics in flavoring essence spices.
The preparation of standard solution described in B step is: take respectively thiocarbamide, sesamol, dehydroactic acid, P-ethoxyphellylurea, ortho-aminobenzoic acid cinnamic ester, nordihydroguaiaretic acid, aristolochic acid A and the each 20mg of sorbic acid, after dissolving with methyl alcohol, be transferred in the brown volumetric flask of 100mL, be settled to scale, obtain respectively single standard storing solution; Take salicylic acid-d4 10mg, after dissolving, be transferred in the brown volumetric flask of 50mL with methyl alcohol, methanol constant volume, to scale, obtains interior mark storing solution; Get respectively in the brown volumetric flask of single standard storing solution 0.1-1mL to 50mL, to scale, obtain one-level mixed standard solution by methanol constant volume; Get in the brown volumetric flask of interior mark storing solution 1mL to 50mL, to scale, obtain mark standard solution in one-level by methanol constant volume; Accurately pipette respectively the one-level mixed standard solution of 0.1mL, 0.5mL, 1mL, 2mL, 5mL to the brown volumetric flask of 50mL, each brown volumetric flask all adds mark standard solution in 2mL one-level, is settled to scale with extraction solution, obtains series standard solution.The described single standard storing solution 0.1mL-1mL that gets is respectively specially and gets respectively thiocarbamide, sesamol, P-ethoxyphellylurea, nordihydroguaiaretic acid, aristolochic acid A, sorbic acid single standard storing solution 1mL and dehydroactic acid, ortho-aminobenzoic acid cinnamic ester 0.1mL.
Get in 0.8mL mark storing solution to the brown volumetric flask of 1L, be settled to scale with extraction solution, obtain containing interior target extraction solution.Described is salicylic acid-d4 containing the internal standard compound in interior target extraction solution.
Described in C step, the preparation method of sample solution is: take 0.5g sample in 50mL tool plug conical flask, accurately add 25mL to contain interior target extraction solution, with 150r/min oscillation extraction 20min, get supernatant and cross organic phase filter membrane, filtrate is sample solution.Sample solution is analyzed with Ultra Performance Liquid Chromatography-tandem mass spectrometer.
Described organic phase filter membrane is 0.22 μ m polyethersulfone filter membrane.Liquid-phase chromatographic analysis condition described in D step is: chromatographic column: C18,1.7 μ m, 2.1 × 100mm chromatographic column; Mobile phase is methyl alcohol A and 0.05% aqueous formic acid B; Flow velocity: 0.4mL/min; Sample size: 5 μ L; Column temperature: 40 DEG C; Sample temperature: 10 DEG C; Adopt gradient elution program, initial A be 10%, B while being 90%, 1min A be that 35%, B is 65%, when 4min A be 65%, B while being 35%, 6min initial A be 90%, B while being 10%, 7min A be 90%, when B is 10%, 8min A be 10%, B while being 90%, 10min A be that 10%, B is 90%; Described mass spectrum condition is for adopting electric spray ion source (ESI) positive ion mode, scan mode is multiple-reaction monitoring pattern, electron spray voltage is 5500V, 600 DEG C of ion source temperatures, gas curtain gas: 25Psi, atomization gas: 55Psi, auxiliary heating gas: 50Psi, the quota ion of thiocarbamide, sesamol, dehydroactic acid, P-ethoxyphellylurea, ortho-aminobenzoic acid cinnamic ester, nordihydroguaiaretic acid, aristolochic acid A, sorbic acid and salicylic acid-d4 is to being respectively:
m/z?60.0,109.1,127.1,?108.0,117.1,193.2,265.1,95.1，97.1。Other mass spectrum parameter is in table 1.
The mass spectrum parameter list of eight kinds of compositions in table 1 flavoring essence spices
The calculating of content described in E step is: first carry out regretional analysis with the chromatographic peak area of the detected object of standard solution and internal standard compound than corresponding its respective concentration ratio and obtain typical curve linear equation, the chromatographic peak area of the object then sample solution being detected, substitution typical curve linear equation, obtains the content of thiocarbamide in sample, sesamol, dehydroactic acid, P-ethoxyphellylurea, ortho-aminobenzoic acid cinnamic ester, nordihydroguaiaretic acid, aristolochic acid A, sorbic acid.
Regression equation, the related coefficient etc. corresponding with typical curve are as shown in table 2.
Typical curve equation and the detection limit of eight kinds of compositions in table 2 flavoring essence spices
Then the chromatographic peak area that detects object sample being recorded, substitution typical curve linear equation, obtain the concentration of thiocarbamide in sample, sesamol, dehydroactic acid, P-ethoxyphellylurea, ortho-aminobenzoic acid cinnamic ester, nordihydroguaiaretic acid, aristolochic acid A and sorbic acid, calculate thus the content of thiocarbamide in flavoring essence spices, sesamol, dehydroactic acid, P-ethoxyphellylurea, ortho-aminobenzoic acid cinnamic ester, nordihydroguaiaretic acid, aristolochic acid A and sorbic acid.
In flavoring essence spices sample, the cubage formula of eight kinds of compositions is as follows:
In formula:
The content of eight kinds of compositions in Xi-sample, unit is milligrams per kilogram (mg/kg);
The concentration of eight kinds of compositions in the sample of Ci-drawn by standard working curve, unit is milligrams per liter (mg/L);
The concentration of eight kinds of compositions in the blank of Ci0-drawn by standard working curve, unit is milligrams per liter (mg/L);
V-extraction solution volume, unit is milliliter (mL);
M-sample mass, unit is gram (g).
The precision of the inventive method and the detection method of recovery of standard addition are as follows: adopt sample to carry out recovery of standard addition test, in sample, add respectively the standard solution of basic, normal, high three variable concentrations levels to carry out recovery of standard addition test, each sample replicate determination 6 times, analysis condition is with embodiment 1, then the relative standard deviation of calculating measured value after the recovery of standard addition of eight kinds of compositions in flavoring essence spices and mark-on, the results are shown in Table 3.
The precision of eight kinds of compositions and the recovery (n=6) in table 3 flavoring essence spices
As can be seen from Table 3, under three mark-on levels, 91.55%, 97.36%, 97.34%, 94.87%, 97.83%, 95.06%, 95.03%, 95.38 % the average recovery rate that adopts the method to measure eight kinds of compositions in flavoring essence spices is respectively:, 3.62%, 3.34%, 3.05%, 3.20%, 4.07%, 2.66%, 2.53 %, 3.93% relative standard deviation of sample determination result is respectively:, illustrate that the method is stable, the recovery of method is high, favorable reproducibility.
The present invention has examined or check repeatability, repeatability and the universality etc. of method by experiment, Ultra Performance Liquid Chromatography-tandem mass spectrum detection method of having determined thiocarbamide, sesamol, dehydroactic acid, P-ethoxyphellylurea, ortho-aminobenzoic acid cinnamic ester, nordihydroguaiaretic acid, aristolochic acid A and sorbic acid in flavoring essence spices, has reached following effect:
(1) detection time is short: adopt the present invention to measure the only 10min of content cycle of thiocarbamide in flavoring essence spices, sesamol, dehydroactic acid, P-ethoxyphellylurea, ortho-aminobenzoic acid cinnamic ester, nordihydroguaiaretic acid, aristolochic acid A and sorbic acid.
(2) solvent consumption is little.
(3) detection flux is high, and detection efficiency is high.
(4) the present invention has simple to operate, highly sensitive, good and the high repeatability and other advantages of the recovery, eight kinds of compositions all have good linear relationship, the detection limit of the method is between 0.010-0.023mg/kg, 3.62%, 3.34%, 3.05%, 3.20%, 4.07%, 2.66%, 2.53 %, 3.93% 91.55%, 97.36%, 97.34%, 94.87%, 97.83%, 95.06%, 95.03%, 95.38 % average recovery rate is respectively:, the relative standard deviation of sample determination result is respectively:.
(5) foundation of the method has very important significance for the quality safety of supervision flavoring essence spices.
Claims (8)
1. eight kinds of composition methods for measuring simultaneously in flavoring essence spices, is characterized in that comprising the following steps:
The preparation of A, extraction solution: the methanol-water solution of getting the standby volume ratio 7:3 of one-tenth of Methanol is extraction solution;
The preparation of B, standard solution: with the standard solution of preparing respectively thiocarbamide, sesamol, dehydroactic acid, P-ethoxyphellylurea, ortho-aminobenzoic acid cinnamic ester, nordihydroguaiaretic acid, aristolochic acid A and sorbic acid containing interior target extraction solution;
The preparation of C, sample solution: use containing interior target extraction solution extraction flavoring essence spices sample, excessively obtain sample solution after organic phase filter membrane;
D, the analysis of Ultra Performance Liquid Chromatography-tandem mass spectrometer: utilize Ultra Performance Liquid Chromatography-tandem mass spectrometer respectively standard solution and sample solution to be detected to analysis;
The calculating of eight kinds of composition thiocarbamides, sesamol, dehydroactic acid, P-ethoxyphellylurea, ortho-aminobenzoic acid cinnamic ester, nordihydroguaiaretic acid, aristolochic acid A and Determination of sorbics in E, flavoring essence spices.
2. eight kinds of composition methods for measuring simultaneously in flavoring essence spices according to claim 1, the preparation that it is characterized in that standard solution described in B step is: take respectively thiocarbamide, sesamol, dehydroactic acid, P-ethoxyphellylurea, ortho-aminobenzoic acid cinnamic ester, nordihydroguaiaretic acid, aristolochic acid A and the each 20mg of sorbic acid, after dissolving with methyl alcohol, be transferred in the brown volumetric flask of 100mL, be settled to scale, obtain respectively single standard storing solution; Take salicylic acid-d4 10mg, after dissolving, be transferred in the brown volumetric flask of 50mL with methyl alcohol, methanol constant volume, to scale, obtains interior mark storing solution; Get respectively in the brown volumetric flask of single standard storing solution 0.1-1mL to 50mL, to scale, obtain one-level mixed standard solution by methanol constant volume; Get in the brown volumetric flask of interior mark storing solution 1mL to 50mL, to scale, obtain mark standard solution in one-level by methanol constant volume; Accurately pipette respectively the one-level mixed standard solution of 0.1mL, 0.5mL, 1mL, 2mL, 5mL to the brown volumetric flask of 50mL, each brown volumetric flask all adds mark standard solution in 2mL one-level, is settled to scale with extraction solution, obtains series standard solution.
3. eight kinds of composition methods for measuring simultaneously in flavoring essence spices according to claim 1, is characterized in that getting in 0.8mL mark storing solution to the brown volumetric flask of 1L, are settled to scale with extraction solution, obtain containing interior target extraction solution.
4. according to eight kinds of composition methods for measuring simultaneously in flavoring essence spices described in claim 1 or 3, it is characterized in that described is salicylic acid-d4 containing the internal standard compound in interior target extraction solution.
5. eight kinds of composition methods for measuring simultaneously in flavoring essence spices according to claim 1, the preparation method who it is characterized in that sample solution described in C step is: take 0.5g sample in 50mL tool plug conical flask, accurately add 25mL to contain interior target extraction solution, with 150r/min oscillation extraction 20min, get supernatant and cross organic phase filter membrane, filtrate is sample solution.
6. according to eight kinds of composition methods for measuring simultaneously in flavoring essence spices described in claim 1 or 5, it is characterized in that described organic phase filter membrane is 0.22 μ m polyethersulfone filter membrane.
7. eight kinds of composition methods for measuring simultaneously in flavoring essence spices according to claim 1, the liquid-phase chromatographic analysis condition described in D step that it is characterized in that is: chromatographic column adopting C18,1.7 μ m, 2.1 × 100mm chromatographic column; Mobile phase is methyl alcohol A and 0.05% aqueous formic acid B; Flow velocity: 0.4mL/min; Sample size: 5 μ L; Column temperature: 40 DEG C; Sample temperature: 10 DEG C; Adopt gradient elution program, initial A be 10%, B while being 90%, 1min A be that 35%, B is 65%, when 4min A be 65%, B while being 35%, 6min initial A be 90%, B while being 10%, 7min A be 90%, when B is 10%, 8min A be 10%, B while being 90%, 10min A be that 10%, B is 90%; Described mass spectrum condition is for adopting electric spray ion source (ESI) positive ion mode, scan mode is multiple-reaction monitoring pattern, electron spray voltage is 5500V, 600 DEG C of ion source temperatures, gas curtain gas: 25Psi, atomization gas: 55Psi, auxiliary heating gas: 50Psi, the quota ion of thiocarbamide, sesamol, dehydroactic acid, P-ethoxyphellylurea, ortho-aminobenzoic acid cinnamic ester, nordihydroguaiaretic acid, aristolochic acid A, sorbic acid and salicylic acid-d4 is to being respectively: m/z 60.0,109.1,127.1,108.0,117.1,193.2,265.1,95.1,97.1.
8. eight kinds of composition methods for measuring simultaneously in flavoring essence spices according to claim 1, the calculating that it is characterized in that content described in E step is: first carry out regretional analysis with the chromatographic peak area of the detected object of standard solution and internal standard compound than corresponding its respective concentration ratio and obtain typical curve linear equation, the chromatographic peak area of the object then sample solution being detected, substitution typical curve linear equation, obtain thiocarbamide in sample, sesamol, dehydroactic acid, P-ethoxyphellylurea, ortho-aminobenzoic acid cinnamic ester, nordihydroguaiaretic acid, aristolochic acid A, the content of sorbic acid.
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| CN111024851A (en) * | 2019-12-26 | 2020-04-17 | 江苏傲农生物科技有限公司 | Method for simultaneously detecting contents of various additives |
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| CN111024851A (en) * | 2019-12-26 | 2020-04-17 | 江苏傲农生物科技有限公司 | Method for simultaneously detecting contents of various additives |
| CN112098550A (en) * | 2020-09-04 | 2020-12-18 | 深圳天祥质量技术服务有限公司 | Method for determining thiourea in daily consumer products |
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