• 专利附录
  • 专利说明
  • 权利要求
  • 类似技术
  • 同族专利
  • 引用文献
  • 法律状态
  • 优先权
    • 专利摘要:
    Method of determining the pungency index in a sample by extracting the analytes from the samples, obtaining the fluorometric signals of the samples (obtaining the excitation-emission matrices) and applying a chemometric algorithm that allows determining the concentration of capsaicinoids to express it later in the corresponding units of the pungency index (Scoville index). (Machine-translation by Google Translate, not legally binding)
    公开号:ES2703807A1
    申请号:ES201700713
    申请日:2017-09-12
    公开日:2019-03-12
    发明作者:Marana Olga Monago；Diaz Teresa Galeano；De La Pena Castrillo Arsenio Munoz
    申请人:Universidad de Extremadura；
    IPC主号:
    专利说明:

    [0001]
    [0002] Method of determining the pungency index in a sample.
    [0003]
    [0004] The present invention relates to a method for measuring the pungency index in a sample, by means of total fluorescence and the analysis of the obtained data.
    [0005]
    [0006] BACKGROUND OF THE INVENTION
    [0007]
    [0008] The pungency of the genus capsicum is due to a group of compounds called capsaicinoids, which are composed of an acid vanillylamine amine linked to chains of fatty acids of length C9-C11.
    [0009]
    [0010] The conventional method used for this purpose was the one developed in 1912 by W. Scoville, which determined the pungency in what are known as Scoville units. This method is an organoleptic method which has been subject to many criticisms given the subjectivity of the same, since each consumer perceives the itching in a different way. Therefore, in 1977 Todd et al. (PH Todd JR, Journal of Food Science, 1977, 42 (3), 660-665) established the pungency values of the different pure capsaicinoids that allow establishing the relationship between concentration of capsaicinoids ( pg / g) and Scoville units.
    [0011]
    [0012] A multitude of methods have been developed for the determination of capsaicinoids in different samples of spicy foods (peppers, chilies, paprika, sauces ...). However, practically all employ separative techniques. These techniques require generally long analysis times, the use of a expensive instrumentation, a mobile phase composed of organic solvents, and, as a consequence, a high consumption of organic solvents, which, in addition to their cost, suppose a greater contamination to the environment.
    [0013]
    [0014] There are patent applications that describe other ways to determine pungency, such as the following:
    [0015]
    [0016] The US application with publication number US201410014534 describes a method of electrochemical detection of capsaicinoids. The US application with publication number US20110218739 describes an apparatus for measuring the pungency of red pepper powder, which includes: a near infrared measuring unit.
    [0017]
    [0018] None of these patent applications describes the use of total fluorescence for the determination of this parameter.
    [0019]
    [0020] Description of the invention
    [0021]
    [0022] The present invention describes a method of determining the pungency index in a sample by means of total fluorescence and mathematical analysis of the obtained data, which is an improvement as described above, since it is a faster, less expensive and more expensive procedure. environmentally friendly.
    [0023]
    [0024] In the method described in the present invention it is necessary to extract the analytes responsible for the pungent character, capsaicinoids and to take advantage of their fluorescent properties which, in combination with the use of mathematical analysis, allows carrying out a determination of this parameter without the need of having a chromatographic equipment. The analysis is carried out in a shorter time, with more economically affordable instrumentation and a lower consumption of solvents, which also results in savings, being a method more respectful with the environment.
    [0025] The method presents good precision and reproducibility.
    [0026] Therefore a first aspect of the invention refers to a method of determining the pungency index in a sample comprising the steps of:
    [0027] a) extraction of the capsaicinoids from the sample;
    [0028] b) measurement by total fluorescence of the capsaicinoids extracted in step a) and recording of the total fluorescence data of the capsaicinoids;
    [0029] c) analysis of the data obtained in stage b).
    [0030] The term "total fluorescence" refers to molecular fluorescence with excitation-emission matrices.
    [0031] Detailed description of the invention
    [0032] As said, a first aspect of the invention refers to a method of determining the pungency index in a sample comprising the steps of:
    [0033] a) extraction of the capsaicinoids from the sample;
    [0034] b) measurement by total fluorescence of the capsaicinoids extracted in step a) and recording of the total fluorescence data of the capsaicinoids;
    [0035] c) analysis of the data obtained in stage b).
    [0036] In the method described, the sample is preferably a food sample or a pharmaceutical sample. In particular, the food sample is paprika or a sauce.
    [0037] In the described method the capsaicinoids are preferably: capsaicin, dihydrocapsaicin, nordihydrocapsaicin, homocapsaicin and homodihydrocapsacin. In particular, the capsaicinoids are capsaicin and dihydrocapsaicin.
    [0038] In a preferred embodiment, the extraction step is a solid phase extraction step. More preferably the solid phase extraction step presents the sub-steps of: sample preparation, retention of the sample in a column, elution of the undesirable components, elution of the capsaicinoids. Still more preferably, the sample preparation step comprises in turn the steps of: weighing the sample, adding a volume of an extraction solvent to the sample in an ultrasonic bath, centrifuging said volume and evaporating the supernatant.
    [0039] The extraction solvent chosen must have generally poor polar characteristics, since these analytes do not have a high polarity. Therefore, the extraction solvent is preferably selected from: methanol, acetonitrile or a mixture of both. In particular, methanol is selected. The volume of extraction is selected in proportion to the solid. Generally between 1:50 and 1: 250 (p / v). Particularly 1: 100 (p / v). The ultrasound extraction time is selected in order to obtain the highest possible fluorimetric signal. Preferably, between 3 and 15 min will be selected. In particular, 10 min is selected. The sample is centrifuged preferably 3-10 min.
    [0040] Particularly 5 min. In particular, the supernatant is evaporated to dryness in a rotary evaporator.
    [0041]
    [0042] Preferably, after evaporating the supernatant, the evaporated residue is reconstructed with a reconstruction solvent. Preferably said solvent is a mixture of methanol: water. More preferably, it is a mixture 10:90 (v / v) - 40:60 (v / v). Particularly 30:70 (v / v).
    [0043]
    [0044] Solid phase extraction is carried out. Preferably said extraction in solid phase is carried out in a reverse phase cartridge; preferably C8 or C18; in a particular way C18.
    [0045]
    [0046] Preferably, the step of eluting the undesirable components is carried out with a mixture of methanol: water 40:60 (v / v) - methanol: water 60:40 (v / v). Preferably methanol: water 60:40 (v / v). The volume to be passed will preferably be between 2 and 6 mL. Particularly 4 mL.
    [0047]
    [0048] As for the elution of the capsaicinoids, it is carried out with a methanol: water mixture. Preferably a methanol: water mixture 70:30 (v / v) - methanol: water 90:10 (v / v). In particular, a methanol: water mixture 80:20 (v / v).
    [0049]
    [0050] Preferably the extract of capsaicinoids is diluted with methanol or acetonitrile. Particularly with acetonitrile.
    [0051]
    [0052] Preferably, step b) is carried out at a range of excitation wavelengths comprised in a range between 200 nm and 350 nm and a range of emission lengths in a range between 250 and 600 nm. In particular, the excitation will be recorded between 210-300 nm and the emission between 280-400 nm.
    [0053]
    [0054] Preferably, the analysis of data obtained in step b) is carried out by means of a mathematical treatment of the signals with second-order chemometric algorithms and the quantification of the total content of capsaicinoids is carried out; in particular, parallel analysis of split partial-minimum factors or factors with residual bilinearization. In particular, parallel analysis of factors.
    [0055]
    [0056] Examples
    [0057]
    [0058] Example 1
    [0059]
    [0060] 0.2 g of paprika are weighed and 20 mL of methanol are added. It is extracted for 10 minutes in an ultrasonic bath and centrifuged for 5 min. The supernatant is evaporated to dryness and reconstituted in 15 mL of a methanol: water mixture 30:70 (v / v). Subsequently, a solid phase extraction is carried out using a C18 cartridge. For this, 5 mL of sample is passed, followed by 4 mL of a methanol: water mixture 40:60 (v / v) and, finally, 4 mL of a methanol: water mixture 80:20 (v / v). Take an aliquot of 0.2 mL of this extract and dilute to 3 mL with acetonitrile. The total fluorescence (molecular fluorescence with excitation - emission matrices) is recorded by setting the excitation wavelength range between 210 and 300 nm and the emission between 260 and 400 nm. The treatment of the signals obtained by means of the algorithm of second order parallel analysis of factors is carried out and the concentration of capsaicinoids is obtained, expressed in Scoville units, that is, the pungency index of the paprika. This result is compared with that obtained by means of a chromatographic analysis and it is verified that the combination of this extraction of the sample, with the obtaining of its spectrofluorimetric signal and the treatment of it by means of an algorithm mathematical is an alternative method for determining the pungency index of paprika.
    [0061] The results were the following:
    [0062] Scoville units according to the method used in Example 1: 12349;
    [0063] Scoville units according to chromatographic analysis: 12188.
    [0064] Example 2
    [0065] Weigh 1.0 g of sauce and add methanol: water 30:70 (v / v) to a fixed volume of 10 mL. Centrifuge for 5 min. Subsequently, a solid phase extraction is carried out using a C18 cartridge. For this, 5 mL of sample is passed, followed by 4 mL of a methanol: water mixture 40:60 (v / v) and, finally, 4 mL of a methanol: water mixture 80:20 (v / v). Take an aliquot of 0.5 mL of this extract and dilute to 3 mL with acetonitrile. The total fluorescence (molecular fluorescence with excitation - emission matrices) is recorded by setting the excitation wavelength range between 210 and 300 nm and the emission between 260 and 400 nm. The treatment of the signals obtained by means of the algorithm of second order parallel analysis of factors is carried out and the concentration of capsaicinoids is obtained, expressed in Scoville units, that is, the pungency index of the paprika. This result is compared with that obtained by means of a chromatographic analysis and it is verified that there is a good correlation between both.
    [0066] The results were the following:
    [0067] Scoville units according to the method used in Example 2: 1336;
    [0068] Scoville units according to chromatographic analysis: 1385.
    权利要求:
    Claims (12)
    [1]
    1. Method of determining the pungency index in a sample characterized in that it comprises the steps of:
    a) extraction of the capsaicinoids from the sample;
    b) measurement by total fluorescence of the capsaicinoids extracted in step a) and recording of the total fluorescence data of the capsaicinoids;
    c) analysis of the data obtained in stage b).
    [2]
    2. Method according to claim 1, characterized in that the sample is a food sample.
    [3]
    3. Method according to claim 1 characterized in that the sample is a pharmaceutical sample.
    [4]
    4. Method according to claim 2 wherein the food sample is paprika.
    [5]
    5. Method according to claim 2 wherein the food sample is a sauce.
    [6]
    Method according to any of claims 1 to 5, characterized in that the capsaicinoids are capsaicin and dihydrocapsaicin.
    [7]
    Method according to any of claims 1 to 6, characterized in that the extraction step is a solid phase extraction step.
    [8]
    8. Method according to claim 7, characterized in that the step of extraction in solid phase presents the sub-steps of: sample preparation, retention of the sample in a column, elution of the undesirable components, elution of the capsaicinoids.
    [9]
    9. Method of determination according to claim 8, characterized in that the sub-step of preparing the sample comprises in turn the steps of: weighing the sample, adding a volume of an extraction solvent to the sample in an ultrasonic bath, centrifuging said volume and evaporate the supernatant.
    [10]
    10. Method of determination according to claim 9, characterized in that after evaporating the supernatant, the evaporated residue is reconstructed with a reconstruction solvent.
    [11]
    11. Method of determination according to claims 1-10, characterized in that step b) is carried out at a range of excitation wavelengths comprised in a range between 200 nm and 350 nm and a range of emission lengths in a range between 250 and 600 nm.
    [12]
    12. Determination method according to claims 1-11, characterized in that step c) is carried out by means of parallel analysis of factors.
    类似技术:
    公开号 | 公开日 | 专利标题
    Yardım et al.2013|Voltammetric determination of mixtures of caffeine and chlorogenic acid in beverage samples using a boron-doped diamond electrode
    Higashi et al.2011|A specific LC/ESI‐MS/MS method for determination of 25‐hydroxyvitamin D3 in neonatal dried blood spots containing a potential interfering metabolite, 3‐epi‐25‐hydroxyvitamin D3
    Li et al.2015|Regioisomeric and enantiomeric analyses of 24 designer cathinones and phenethylamines using ultra high performance liquid chromatography and capillary electrophoresis with added cyclodextrins
    Shamsipur et al.2015|Extraction and determination of polycyclic aromatic hydrocarbons in water samples using stir bar sorptive extraction | combined with dispersive liquid–liquid microextraction based on the solidification of floating organic drop | followed by HPLC-UV
    Tsai et al.2010|Determination of sulfonamides in swine muscle after salting-out assisted liquid extraction with acetonitrile coupled with back-extraction by a water/acetonitrile/dichloromethane ternary component system prior to high-performance liquid chromatography
    Kudo et al.2014|Rapid and simultaneous extraction of acidic and basic drugs from human whole blood for reliable semi-quantitative NAGINATA drug screening by GC–MS
    Biziuk et al.2015|Multiresidue methods for determination of currently used pesticides in fruits and vegetables using QuEChERS technique
    El Deeb et al.2008|Strategies in method development to quantify enantiomeric impurities using CE
    Welch et al.2014|Evaluation of capsaicin in chili peppers and hot sauces by MISER HPLC-ESIMS
    De Girolamo et al.2012|Analytical performances of a DNA-ligand system using time-resolved fluorescence for the determination of ochratoxin A in wheat
    van Duynhoven et al.2013|Quantification of complex mixtures by NMR
    Liu et al.2019|A highly efficient vortex-assisted liquid–liquid microextraction based on natural deep eutectic solvent for the determination of Sudan I in food samples
    Gilbert-López et al.2014|Determination of polyphenols in commercial extra virgin olive oils from different origins | by liquid chromatography–electrospray time-of-flight mass spectrometry
    Ali et al.2013|Chromatographic determination of aliphatic aldehydes in human serum after pre-column derivatization using 2, 2′-furil, a novel fluorogenic reagent
    Xiong et al.2014|Separation and quantitation of three acidic herbicide residues in tobacco and soil by dispersive solid-phase extraction and UPLC–MS/MS
    Boxler et al.2019|Analytical considerations for |‐targeted metabolomic studies with special focus on forensic applications
    Jia et al.2016|Rapid selective accelerated solvent extraction and simultaneous determination of herbicide atrazine and its metabolites in fruit by ultra high performance liquid chromatography
    Bane et al.2016|LC-MS/MS method for the determination of tetrodotoxin | on a triple quadruple mass spectrometer
    Amraei et al.2019|Cloud point extraction and simultaneous spectrophotometric determination of Allura red and carmoisine using wavelet orthogonal signal correction–partial least squares method
    Filho et al.2011|Multi‐residue analysis of pesticide residues in mangoes using solid‐phase microextraction coupled to liquid chromatography and UV–Vis detection
    ES2703807B2|2019-09-30|Method of determining the pungency index in a sample
    Başkan et al.2010|Non‐ionic micellar electrokinetic chromatography with laser‐induced fluorescence: A new method tested with biogenic amines in brined and dry‐salted fish
    Er et al.2021|Determination of seventeen free amino acids in human urine and plasma samples using quadruple isotope dilution mass spectrometry combined with hydrophilic interaction liquid chromatography–Tandem mass spectrometry
    Tymm et al.2021|A single laboratory validation for the analysis of underivatized β-N-methylamino-L-alanine |
    Aral et al.2014|HPLC separation of different groups of small polar compounds on a novel amide-embedded stationary phase
    同族专利:
    公开号 | 公开日
    ES2703807B2|2019-09-30|
    引用文献:
    公开号 | 申请日 | 公开日 | 申请人 | 专利标题
    KR100871681B1|2008-08-22|2008-12-03|대한민국|Analytical method using infrared rays for examining the content of red pepper seed in red pepper powder|
    US20110218739A1|2010-03-03|2011-09-08|Rural Development Administration|Apparatus and method for measuring pungency of red pepper powder|
    法律状态:
    2019-03-12| BA2A| Patent application published|Ref document number: 2703807 Country of ref document: ES Kind code of ref document: A1 Effective date: 20190312 |
    2019-09-30| FG2A| Definitive protection|Ref document number: 2703807 Country of ref document: ES Kind code of ref document: B2 Effective date: 20190930 |
    优先权:
    申请号 | 申请日 | 专利标题
    ES201700713A|ES2703807B2|2017-09-12|2017-09-12|Method of determining the pungency index in a sample|ES201700713A| ES2703807B2|2017-09-12|2017-09-12|Method of determining the pungency index in a sample|
    [返回顶部]