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  • 专利说明
  • 权利要求
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    • 专利摘要:
    Device and method for measuring ascorbic acid and anthocyanins in juices. A device and method for measuring ascorbic acid and anthocyanins in juices comprising an amperometric sensor (1) comprising a first working electrode (3) and a second working electrode (4); electronic means for capturing and processing data (6, 7, 8, 9); a graphic management and/or configuration interface (10); and where the first working electrode (3) is not treated with ascorbate oxidase and a second working electrode (4) that is treated with ascorbate oxidase; and where the electronic means (6, 7, 8, 9) are configured to stimulate the sensor (1) according to a predefined range of voltages depending on the type of juice selected by a user in the graphic management interface and/or configuration (10), and simultaneously obtain the current reading due to the oxidation of the juice studied. (Machine-translation by Google Translate, not legally binding)
    公开号:ES2728448A1
    申请号:ES201930555
    申请日:2019-06-18
    公开日:2019-10-24
    发明作者:Gimenez Encarnacion Pilar Aguayo；Poveda Juan Aznar；Sanchez Jose Francisco Beltran；Haro Juan Garcia；Sanchez Antonio Javier Garcia；Pastor Jose Antonio Lopez；Sanchez Ascension Martinez
    申请人:Universidad Politecnica de Cartagena；
    IPC主号:
    专利说明:

    [0001]
    [0002] Device and method for measuring ascorbic acid and anthocyanins in juices
    [0003]
    [0004] Field of the Invention
    [0005]
    [0006] The present invention relates to a programmable electronic device and a method by which the concentration of ascorbic acid (AA) and anthocyanins present in juices can be measured amperometrically. The present invention has application in the agri-food industry and specifically in the industry of obtaining natural juices.
    [0007]
    [0008] Prior art
    [0009]
    [0010] The food industry, and in particular, the manufacture of juices requires the addition of antioxidant during processing to avoid rust-reduction reactions, ensure the quality of the final product and prolong its shelf life. Ascorbic acid (AA), is a reducing agent and therefore a natural antioxidant present in fruits and vegetables, but also has a wide use as an additive in juices, jams, dairy products and other foods, in addition to the nutraceutical industry and pharmaceutical.
    [0011]
    [0012] Other bioactive compounds of interest in the food industry are anthocyanins, compounds responsible for the orange, red and purple colors of plant-based foods, which have a high antioxidant capacity with highly beneficial health properties. Anthocyanins possess pharmacological and functional properties related to reducing the incidence of cardiovascular diseases and cancer, improving memory or treating urinary tract disorders. These properties make anthocyanins a very attractive ingredient for the food industry and increasingly valued and demanded by consumers.
    [0013]
    [0014] However, both AA and anthocyanins are substances sensitive to high temperatures, the presence of oxygen or the pH of the matrix, so that their concentration can be reduced after processing. In addition, in the case of juices, the criteria of quality and authenticity must be met, the analysis confirming their composition being necessary. The food industry needs to control the AA content of the product in each of the processing stages, as well as in the final products (eg juices) continuously in order to ensure at all times, the quality of said product. On the other hand, in the particular case of juices containing anthocyanins, it is necessary to control their content / concentration in each of the stages to maintain an appropriate composition and characteristics of the final product.
    [0015]
    [0016] A variety of methods can be found in the scientific literature to measure and quantify ascorbic acid (AA). Among them, the most prominent and widespread are spectrometry, titrimetry, colorimetric method with test strips and high efficiency liquid chromatography (HPLC). The latter technology is the most widespread due to its high precision and its ability to measure different compounds in the same analysis. In the case of vitamin C, by means of HPLC, both AA and its oxidized form, dehydroascorbic acid, can be rigorously quantified.
    [0017]
    [0018] With respect to anthocyanins, the most common methods of determination that can be found are spectroscopy, HPCL and nuclear magnetic resonance imaging of one and two dimensions. Although AA and anthocyanins can be identified and quantified by HPLC, the extraction and characteristics of HPLC analysis are different. As a consequence, both substances - anthocyanins and AA - cannot be determined in the same analysis.
    [0019]
    [0020] All these methods have some important disadvantages in common, the most prominent being the following: (i) high cost per determination; (ii) require highly qualified, specialized and experienced laboratory personnel; (iii) implement complex protocols that include laborious sample preparations; (iv) they need considerable analysis time and a large amount of reagents; (v) there must be a laboratory equipped with instrumentation - often expensive - that demands a controlled environment to carry out the analytics; e (vi) implies a different analysis for each substance under study.
    [0021]
    [0022] Due to the inconveniences (i) - (vi) previously stated, the agrifood industry lacks a simultaneous measurement system of AA and anthocyanins that allows carrying out sample measurements with a periodicity and economic amount suitable for industrial requirements. In this sense, the agri-food industry requires (i) low cost per measurement and determination device; (ii) portability; (iii) ease of use and reduced determination time; (iv) avoid sample processing; and (v) measure several substances of interest using a single analytical. These requirements are in line with the new paradigm of quality and safety in food control under European Union regulation ( European Commission Priorities: Agriculture and Food Security (2018). Obtained at https://ec.europa.eu/jrc/en/science-area/ agriculture-and-food-security).
    [0023]
    [0024] In order to solve the demands of the food industry and deal with the indicated inconveniences, different methods have been developed in order to determine the AA and other antioxidant substances. These are based on the reduction power of the analyte to be measured, in this case the AA and the anthocyanins, both natural donors of electrons and, therefore, electroactive. This feature allows quantification by electrochemical techniques. However, because both substances have similar oxidizing potential, the joint presence of both substances interferes with the analysis and makes their measurement difficult.
    [0025]
    [0026] The use of electrochemical techniques includes the use of printed carbon electrodes (SPE in English terminology). The use and modification of the SPE allow obtaining the concentration of AA in different matrices such as juices, fruits and / or food supplements. Although the SPE are clear exponents of a disposable and low-priced technology, the modifications in its structure require the use of nanomaterials, antibodies and oxidation reaction enhancers such as hydroquinone or hydrogen peroxide, which implies the loss of simplicity and an increase in cost and maintenance because they are solutions that, in general, require a very specific work environment - temperature, sample pH, laboratory conditions or others - and special storage conditions for proper stabilization and durability over time. However, when non-SPE carbon electrodes are used without modifying the detection of AA in different juices, their use in complex matrices, such as juices, which have known interferents, such as anthocyanins, is not validated.
    [0027]
    [0028] In the state of the art, the described works that try to determine the AA by means of SPE without modifying in complex matrices are not able to reach the required precision at the time of obtaining the concentration of AA motivated by interferences produced by polyphenols and anthocyanins, given which oxidize to a potential similar to that of AA. On the other hand, the works that try to determine the concentration of AA through modified sensors are expensive to manufacture, do not have or have few reuses and must be stored and used in special laboratory conditions. With respect to the anthocyanins, the works that integrate SPE in their work method have not been validated in complex matrices.
    [0029] In addition, it can be concluded that in the state of the art it does not describe (i) simultaneously perform the determination of AA and anthocyanins by means of a dual SPE formed by two working electrodes in the same analysis, (ii) provide a complete system , and (iii) a method capable of satisfying the needs of the food industry with respect to the quantification of the content of AA and anthocyanins such as that which is intended to be protected with the present invention.
    [0030]
    [0031] Explanation of the invention.
    [0032]
    [0033] The object of the present invention is a device and a method for the measurement of ascorbic acid and anthocyanins in juices, which comprises an electronic device that implements a detection method that simultaneously measures the concentration of AA and anthocyanins. This object is achieved by the device and the method of the independent claims that are included herein. Other embodiments and aspects of the present invention are protected in the dependent claims.
    [0034]
    [0035] It is another object of the present invention a device and a method that allows its transport in a simple way, allowing the realization of analytics in the field or where the user requires it, such as, for example, industrial environments or quality control departments of food businesses In addition, it is an additional object that the device is easy to handle, that is, that does not require specializing in AA and anthocyan analysis techniques or in the handling of specific laboratory instrumentation. Additionally, the device has a low cost per measure of interest, which translates into reduced cost of both the electronic equipment that performs the analyzes and the sensors used in each determination.
    [0036]
    [0037] It is another object of the invention that the analysis occurs in a limited time interval, approximately in 30 seconds. In addition, the determination of the measure, from the original matrix of the sample itself, without the need for extraction stages of the compounds to be analyzed. In addition, not requiring calibration through the use of chemical standards. The simultaneous measurement capacity of AA and anthocyanins using a single analytical sensor must also have a range of detection and precision comparable to other conventional laboratory techniques with greater cost and complexity.
    [0038]
    [0039] More specifically, the device for measuring ascorbic acid and anthocyanins in juices of the invention comprises an amperometric sensor comprising a first electrode of work and a second work electrode, an auxiliary electrode and a reference electrode that are printed on a ceramic surface configured to receive at least one sample of the juice to determine its ascorbic acid and anthocyanin content; electronic means of data capture and processing; a graphic management and / or configuration interface; and that it is characterized in that a first working electrode is not treated with ascorbate oxidase and a second working electrode that is treated with ascorbate oxidase; and where the electronic means are configured to: stimulate the sensor according to a predefined range of voltages depending on the type of juice selected by a user in the graphic management interface and / or configuration, and obtain the current reading due to oxidation of the studied juice, so that the current readings of the first working electrode, the current generated by the anthocyanins and ascorbic acid is obtained, while in the second working electrode treated with ascorbate oxidase only the current generated exclusively by the ascorbic acid.
    [0040]
    [0041] In a particular embodiment, the second working electrode is treated with immobilized ascorbate oxidase by 5% polyethylene glycol or 0.05% glutaraldehyde.
    [0042]
    [0043] The method for the measurement of ascorbic acid and anthocyanins in juices, which is implemented in the previous device and comprising the steps of: depositing a sample of juice on the surface of an amperometric sensor; select the calibration line of the type of juice of the sample deposited in the sensor; execute an amperometry process with the start voltage, end voltage levels and a number of previously established incremental voltage jumps along with the calibration line selected for the type of juice in the sample; simultaneously measure the ascorbic acid and anthocyanins in the juice sample deposited on a first working electrode and a second working electrode of the sensor being said second working electrode previously treated with ascorbate oxidase; and obtain the current reading due to the oxidation of the studied juice, so that the current readings of the first working electrode obtain the current generated by the anthocyanins and ascorbic acid, while in the second working electrode treated with Ascorbate oxidase only determines the current generated exclusively by ascorbic acid.
    [0044]
    [0045] In a particular embodiment, the method of the invention can execute a cyclic voltammetry process or by means of a fixed potential. In another embodiment, the method further comprises the steps of displaying the measurement results in a graphical interface and / or saving the measurement results in an internal memory of the device of the invention; and / or send the results to a remote database.
    [0046] Throughout the description and the claims, the word "comprises" and its variants are not intended to exclude other technical characteristics, additives, components or steps. For those skilled in the art, other objects, advantages and features of the invention will be derived partly from the invention and partly from the practice of the invention. The following examples and drawings are provided by way of illustration and are not intended to restrict the present invention. In addition, the invention covers all possible combinations of particular and preferred embodiments indicated herein.
    [0047]
    [0048] Brief description of the drawings
    [0049]
    [0050] Next, a drawing that helps to better understand the invention and that relates expressly to an embodiment of said invention, which is illustrated as a non-limiting example thereof, is described very briefly.
    [0051]
    [0052] FIG. 1 shows a block diagram of the device for measuring ascorbic acid and anthocyanins in juices, object of the present invention.
    [0053]
    [0054] References:
    [0055]
    [0056] 1. Sensor
    [0057] 2. Auxiliary electrode
    [0058] 3. First working electrode
    [0059] 4. Second working electrode
    [0060] 5. Reference electrode
    [0061] 6. Potentiostat
    [0062] 7. Analog-digital converter
    [0063] 8. Control unit
    [0064] 9. Processor
    [0065] 10. Graphical management and / or configuration interface
    [0066]
    [0067] Explanation of a detailed embodiment of the invention and examples
    [0068]
    [0069] Figure 1 shows a block diagram of the measurement device of AA and anthocyanin in juices that allows simultaneous measurement, according to the object of the present invention. Thus, in a practical example of application of the invention, the juice sample which it is desired to analyze is deposited on a dual printed carbon electrode sensor 1 - hereinafter sensor 1 - which has been partially modified for the specific use according to the present invention. For the deposit of the sample, the device housing - not shown in the attached figure will logically have a slot for this purpose.
    [0070]
    [0071] In particular, the method of analysis consists in depositing a drop of 50 microliters on the surface of the SPE covering all electrodes 2, 3, 4 and 5. For the deposition of the drop a configurable micropipette with said quantity or any other can be used system that allows obtaining said exact amount of liquid.
    [0072]
    [0073] The second working electrode 4 has been previously treated with immobilized ascorbate oxidase by 5% polyethylene glycol or 0.05% glutaraldehyde. The object is to determine in this second working electrode 4 the exact amount of ascorbic acid, while in the first working electrode 3, which has not been treated and / or modified, the concentration of ascorbic acid is provided together with the anthocyanins, thus obtaining the quantity of both substances simultaneously.
    [0074]
    [0075] In order for the device to function correctly and accurately, the calibration lines for ascorbic acid and anthocyanins have been previously obtained for each juice to be analyzed. That is to say, in the memory of processor 9, both the calibration line of ascorbic acid and anthocyanins are preloaded for (i) all kinds of juices of a single flavor that can be found in the market (pineapple, grape, apple, peach, carrot, beet, tomato, guava, mango, kiwi, coconut, orange with pulp, orange without pulp, tangerine, pomegranate, grapefruit, pear and banana, among others) and, (ii) a plurality of juices formed by the combination of other juices and additional supplements added. Thanks to the fact that the calibration lines are stored or pre-loaded in the device, both the concentrations of AA and the anthocyanins of most of the juices on the market can be determined and can be expanded and easily configured in the event that new juices to measure arise. That is, the device proposed by the present invention is scalable being able to store new calibration lines of new juices or juice matrices. Simply, when the drop has been correctly deposited in the sensor 1, the type of juice under study is selected through the graphic management and / or configuration interface 10.
    [0076]
    [0077] When the drop has been deposited in the sensor 1, the control unit 8 begins to execute a cyclic voltammetry (CV) process with the starting voltage, voltage levels of final and the number of previously established incremental tension jumps, together with the selected calibration line of the juice to be analyzed for each substance - ascorbic acid and anthocyanins are understood as substance.
    [0078]
    [0079] When carrying out the voltage sweep, the control unit 8 sends the necessary instructions to an analog-to-digital converter 7 and a potentiostat 6 that integrates the two measurement channels that correspond to the working electrodes 3 and 4 As the potential sweep progresses, potentiostat 6 obtains the current readings due to the oxidation of the juice under study. In the current readings of the first working electrode 3 - the unmodified electrode - the current generated by the anthocyanins and ascorbic acid is obtained, while in the second working electrode 4 - the modified electrode only the current generated is determined only by ascorbic acid
    [0080]
    [0081] The readings of both working electrodes 3 and 4 are sent through the analog-digital converter 7 to the control unit 8 where they are stored in a memory for this purpose. Once the control unit 8 receives all the current data, these are sent to the processor 9 which calculates the concentration of ascorbic acid and anthocyanins present in the sample volume added to the sensor based on the current and straight results. of particularized calibration for each of the two substances and the juice under study. The results are shown to the user through a graphic management interface of the device 10.
    [0082]
    [0083] The software component of the device is configurable according to the needs of the user and / or the analysis to be performed. Thus, once the device is turned on, the management software of the device is loaded and started. In the graphical interface 10 it is possible to choose between several configuration parameters, the most relevant being the type of juice to be analyzed, but it is also possible to select the amperometric measurement method or the sensitivity of the measurement or other more specific operating modes intended to load new calibration lines for other -new-juices or juice matrices, so that the system is scalable.
    [0084]
    [0085] The mode of selection of the amperometric method allows both cyclic voltammetry and fixed potential to be performed, depending on the technique that best suits the determination of the concentration of ascorbic acid and anthocyanins for a given juice. On the other hand, the sensitivity selection mode is designed to vary the internal resistance of the system and increase its sensitivity, so that the accuracy is improved, although, on the contrary, a smaller measuring range is available. In the normal mode of operation of the determination of ascorbic acid and anthocyanin from a sample, the type of juice to be analyzed is selected, together with additional data related to the batch, the date of manufacture or other data introduced by the sample itself. Username. This information, together with the result of the analysis, is sent to a remote database through a means of communication, which may be whatever the person skilled in the art deems most suitable for the transmission of said data (cable communication, serial, wireless or any other communication).
    [0086]
    [0087] Once the desired amperometric technique has been decided for the selected juice and after loading the calibration line, the measurement is started and the concentration of ascorbic acid and anthocyanins in moles / 100 ml and milligrams / 100 is shown in graphical interface 10 ml. Once the data is displayed, the user can carry out a series of options: (i) end the measurement to perform another, (ii) turn off the device, (iii) save the result in the device's internal memory and (iv) send the result to the remote database, or (v) save only one graph with the data obtained.
    [0088]
    [0089] Notwithstanding the practical embodiment described above, it is evident to one skilled in the art that the software incorporated in the device may be exposed to changes, so that it can be extended to determine other substances of interest other than ascorbic acid and Anthocyans simply by selecting a new device configuration, loading new calibration lines for new substances and using a specific sensor modified or not to carry out such an effect.
    权利要求:
    Claims (4)
    [1]
    1 A device for measuring ascorbic acid and anthocyanins in juices comprising:
    an amperometric sensor (1) comprising a first working electrode (3) and a second working electrode (4), an auxiliary electrode (2) and a reference electrode (5) that are printed on a ceramic surface configured to receive at least one sample of the juice to determine its content of ascorbic acid and anthocyanins;
    electronic means for capturing and processing data (6, 7, 8, 9);
    a graphic management and / or configuration interface (10);
    and characterized in that a first working electrode (3) is not treated with ascorbate oxidase and a second working electrode (4) that is treated with ascorbate oxidase;
    and where the electronic media (6, 7, 8, 9) are configured to:
    stimulate the sensor (1) according to a predefined range of voltages depending on the type of juice selected by a user in the graphic management and / or configuration interface (10), and
    simultaneously obtain the current reading due to the oxidation of the studied juice, so that the current readings of the first working electrode (3) obtains the current generated by the anthocyanins and ascorbic acid, while in the second electrode of Work (4) treated with ascorbate oxidase only determines the current generated exclusively by ascorbic acid.
    [2]
    2. - The device according to claim 1 wherein the second working electrode (4) is treated with immobilized ascorbate oxidase by 5% polyethylene glycol or 0.05% glutaraldehyde.
    [3]
    3. - A method for measuring ascorbic acid and anthocyanins in juices,
    which is implemented in the device of claims 1 or 2
    and that includes the stages of:
    deposit a sample of juice on the surface of an amperometric sensor (1), select the calibration line of the type of juice of the sample deposited in the sensor (1),
    run an amperometry process with the start voltage, end voltage levels and a number of previously established incremental voltage jumps along with the selected calibration line of the sample juice type,
    simultaneously measure ascorbic acid and anthocyanins in the juice sample deposited on a first working electrode (3) and on a second working electrode (4) of the sensor (1), said second working electrode (4) being previously treated with ascorbate oxidase, and
    obtain the current reading due to the oxidation of the studied juice, in such a way that the current generated by the anthocyanins and the ascorbic acid is obtained from the current readings of the first working electrode, while in the second electrode of Work (4) treated with ascorbate oxidase only determines the current generated exclusively by ascorbic acid.
    [4]
    4. - The method according to claim 3 wherein the amperometry is cyclic or fixed potential.
    - The method according to one of claims 3 or 4, further comprising the steps of displaying the measurement results in a graphical interface (10) and / or saving the measurement results in an internal memory of the device claims 1 to 2; and / or send the results to a remote database.
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    优先权:
    申请号 | 申请日 | 专利标题
    ES201930555A|ES2728448B2|2019-06-18|2019-06-18|Device and method for the measurement of ascorbic acid and anthocyanins in juices|ES201930555A| ES2728448B2|2019-06-18|2019-06-18|Device and method for the measurement of ascorbic acid and anthocyanins in juices|
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