Analysis method and evaluation method

Abstract

A cultured meat analysis method according to the present disclosure includes: a step (S104) for acquiring a first weight of cultured meat; and a step (S112) for crushing the freeze-dried cultured meat into a powder. In addition, the cultured meat analysis method includes: a step for preparing an analysis sample using the powder; and a step for acquiring the amount of components contained in the cultured meat by using the result of measuring the analysis sample with an analysis device. The cultured meat analysis method further includes a step for using the first weight and the amount of the components to calculate the amount of components per unit wet weight of the cultured meat.

Description

Analysis method and evaluation method

[0001] The present invention relates to an analysis method and an evaluation method, and more specifically, to a method for evaluating the quality of cultured meat based on the amount of components contained in the cultured meat.

[0002] Due to the increase in the global population and the impact of climate change, it is expected that the world's food supply and demand will be strained in the future. In particular, the consumption of meat is expected to increase with the increase in income in emerging countries, etc. However, it is difficult to increase the supply of meat due to the limited production of grains for livestock feed and the limitations of breeding areas, etc. Therefore, cultured meat has attracted attention as one of the sustainable alternative means to meet the increasing demand for meat.

[0003] Cultured meat is artificial meat made from animal cells. Feed grains and vast land are not required for the production of cultured meat., Therefore, it is expected that the production of cultured meat will meet the increasing demand for meat.

[0004] In order to ensure the safety of cultured meat as a food and maintain and improve its quality, it is important to evaluate the component amount and physical properties of the produced cultured meat. Regarding the conventional evaluation method for meat, the Ministry of Education, Culture, Sports, Science and Technology, the Science and Technology Council, the Subcommittee on Resource Survey, the Food Composition Committee, "Analysis Manual of the Japanese Food Standard Composition Table 2020 Edition (Eighth Edition)", February 2022 (Non-Patent Document 1) discloses a method for evaluating meat.

[0005] Ministry of Education, Culture, Sports, Science and Technology, Science and Technology Council, Subcommittee on Resource Survey, Food Composition Committee, "Analysis Manual of the Japanese Food Standard Composition Table 2020 Edition (Eighth Edition)", February 2022

[0006] By analyzing the amount of components in cultured meat according to conventional meat evaluation methods, such as those disclosed in Non-Patent Document 1, it becomes possible to compare the components of cultured meat with those of meat and other cultured meats, thereby evaluating the quality of the produced cultured meat. However, for example, the meat evaluation method disclosed in Non-Patent Document 1 requires the preparation of a sample of about 300 to 500 g, while preparing several hundred g of cultured meat can be a time and cost burden on the user. Thus, it can be difficult to use evaluation methods that have been used for meat to evaluate cultured meat. Therefore, there is a need for the development of evaluation methods for assessing the quality of cultured meat.

[0007] This disclosure is made in light of these circumstances, and its purpose is to provide a technology for evaluating the quality of cultured meat based on data on the amount of components obtained from a smaller sample volume compared to conventional meat evaluation methods.

[0008] An analytical method according to an aspect of the present disclosure includes the steps of (a) obtaining a first weight of cultured meat; (b) crushing the freeze-dried cultured meat into powder; (c) preparing an analytical sample using the powder; (d) obtaining the amount of components contained in the cultured meat using the results of measuring the analytical sample with an analytical instrument; and (e) calculating the amount of components per unit wet weight of the cultured meat using the first weight and the amount of components.

[0009] An evaluation method according to an aspect of this disclosure includes the steps of: (A) comparing the moisture content of cultured meat calculated by a method for analyzing cultured meat with a first reference value; and (B) evaluating the quality of the cultured meat using the results of the comparison step. The method for analyzing cultured meat includes the steps of: (A-1) obtaining a first weight of cultured meat; (A-2) crushing freeze-dried cultured meat into powder; (A-3) preparing an analytical sample using the powder; (A-4) obtaining the amount of components contained in the cultured meat using the results of measuring the analytical sample with an analytical instrument; (A-5) calculating the amount of components per unit wet weight of cultured meat using the first weight and the amount of components; (A-6) obtaining a second weight of freeze-dried cultured meat; and (A-7) calculating the moisture content of the cultured meat using the first weight and the second weight, and comparing the moisture content of the cultured meat calculated by a method for analyzing cultured meat with a first reference value; and (A-8) evaluating the quality of the cultured meat using the results of the comparison step.

[0010] According to this disclosure, compared to conventional meat evaluation methods, the quality of cultured meat can be evaluated based on data on the amount of components obtained from a smaller sample volume.

[0011] This figure shows an example of the configuration of an analysis system. This figure shows the configuration of a mass spectrometer. This figure is for explaining the method of creating cultured meat. This is a flowchart of the process for evaluating the components of cultured meat. This is a flowchart of the subroutine for step S100 shown in Figure 4. This is a flowchart of the subroutine for step S200 shown in Figure 4. This is a flowchart of the subroutine for step S300 shown in Figure 4. This is a flowchart of the subroutine for step S400 shown in Figure 4. This is a flowchart of the subroutine for step S402 shown in Figure 8. This is a flowchart of the process for evaluating the texture of cultured meat. This figure shows the amount of amino acids per unit wet weight of cultured meat in an experimental example. This figure shows the amount of amino acids per unit wet weight of meat in an experimental example. This figure shows the amount of fatty acids per unit wet weight of cultured meat in an experimental example. This figure shows the amount of fatty acids per unit wet weight of meat in an experimental example. This figure shows the amount of lactones per unit wet weight of cultured meat in an experimental example. This figure shows the amount of lactones per unit wet weight of meat in an experimental example.

[0012] The embodiments of this disclosure will be described in detail below with reference to the drawings. In the drawings, the same or corresponding parts are denoted by the same reference numerals, and their descriptions will not be repeated.

[0013] [Configuration of the Analysis System] Figure 1 shows an example of the configuration of the analysis system. As shown in Figure 1, the analysis system 100 includes a liquid chromatograph (LC) 10, a mass spectrometer (MS) 20, a control device 30, an input device 40, and a display device 50. In this embodiment, the analysis system 100 will be described using a configuration that includes a liquid chromatograph tandem mass spectrometer (LC-MS / MS) as an example, but the analytical instruments included in the analysis system 100 are not limited to LC-MS / MS, and may include, for example, a liquid chromatograph mass spectrometer (LC-MS), a gas chromatograph tandem mass spectrometer (GC-MS / MS), and a gas chromatograph mass spectrometer (GC-MS). When the analysis system 100 is configured to include LC-MS / MS and LC-MS, the components to be analyzed are soluble components, such as amino acids, fatty acids, and nucleic acid-related substances. Nucleic acid-related substances include, for example, nucleosides, ribonucleotides, and deoxyribonucleotides. When the analytical system 100 is configured to include GC-MS / MS and GC-MS, the analyte is a volatile component, such as fatty acids and fragrance components. Fragrance components include, for example, lactones, ketones, and aldehydes.

[0014] LC10 separates the components contained in a sample by introducing the sample to be analyzed together with the eluent, which is the mobile phase, into a column. MS20 performs mass spectrometry on the components introduced from LC10. MS20 includes an ionization chamber 21, a mass separation unit 22 (a first intermediate vacuum chamber 221, a second intermediate vacuum chamber 222, and an analysis chamber 223 in Figure 2), and an ion detector 23. The structure of MS20 will be described in detail with reference to Figure 2.

[0015] LC10 includes a mobile phase container 11, a liquid delivery pump 12, an injector 13, a flow path switching valve 14, and a column 15. The column 15 can be any column for reversed-phase chromatography, but for example, an ODS (Octa Decyl Silyl) column can be suitably used. The flow path switching valve 14 is configured to switch between a state in which the multiple ports provided on the valve 14 are connected as shown by the dotted lines in Figure 1 and a state in which they are connected as shown by the solid lines in Figure 1.

[0016] In LC10, the liquid delivery pump 12 draws the mobile phase from the mobile phase container 11 and delivers it to the column 15 at a constant flow rate. The injector 13 injects a fixed amount of sample (analytical sample) into the mobile phase at predetermined timings. The analytical sample is prepared from cultured meat and meat using a method described later. The injected sample is introduced into the column 15 along with the flow of the mobile phase. As it passes through the column 15, multiple compounds (components) contained in the sample are separated and eluted from the outlet of the column 15 at different times. The eluted components are introduced into MS20.

[0017] Figure 2 shows the configuration of the MS20. Referring to Figure 2, the MS20 has a multi-stage differential pumping system configuration that includes first and second intermediate vacuum chambers 221 and 222, in which the vacuum level is gradually increased, between an ionization chamber 21 which is at approximately atmospheric pressure and a high-vacuum analysis chamber 223 which is evacuated by a vacuum pump (not shown).

[0018] An electrospray ionization (ESI) probe 211 is installed in the ionization chamber 21, which sprays the sample solution while applying an electric charge. The ionization chamber 21 and the next stage first intermediate vacuum chamber 221 are connected through a small-diameter heated capillary 212. The first intermediate vacuum chamber 221 and the second intermediate vacuum chamber 222 are separated by a skimmer 2211 with a small hole at the top, and ion guides 2212 and 2221 are installed in the first intermediate vacuum chamber 221 and the second intermediate vacuum chamber 222, respectively, for transporting ions to the next stage while focusing them.

[0019] In the analysis chamber 223, a collision cell 2232, which has a multipole ion guide 2233 installed inside, is flanked by a pre-stage quadrupole mass filter 2231 capable of separating ions according to their mass-charge ratio, and a post-stage quadrupole mass filter 2234, which also separates ions according to their mass-charge ratio. Furthermore, an ion detector 23 is installed following the post-stage quadrupole mass filter 2234. The collision cell 2232 is connected to a collision-induced dissociation (CID) gas supply mechanism (not shown), which introduces CID gas into the collision cell 2232. The CID gas promotes ion dissociation. The ESI probe 211, ion guides 2212, 2221, 2233, quadrupole mass filters 2231, 2234, etc., are each connected to a power supply (not shown) to which a predetermined voltage is applied. Furthermore, the quadrupole mass filters 2231 and 2234 each have a pre-rod electrode in front of the main rod electrode to correct for electric field disturbances at the input end, and a different voltage from that applied to the main rod electrode can be applied to the pre-rod electrode.

[0020] In the MS20, when the eluate from LC10 reaches the electrospray ionization probe 211, the eluate is sprayed while an electric charge is applied to the tip of the probe 211. The charged droplets formed by the spray are divided and miniaturized by the electrostatic force caused by the applied charge, and in the process the solvent vaporizes and ions derived from the compound are released. The ions thus generated are sent to the first intermediate vacuum chamber 221 through the heated capillary 212, focused by the ion guide 2212, and sent to the second intermediate vacuum chamber 222 through the small pore at the top of the skimmer 2211. The ions derived from the compound are then focused by the ion guide 2221 and sent to the analysis chamber 223, where they are introduced into the space along the long axis of the pre-stage quadrupole mass filter 2231. Note that the ionization method is not limited to electrospray ionization; atmospheric pressure chemical ionization or atmospheric pressure photoionization may also be used.

[0021] The MS20 has a configuration that enables MS / MS analysis. In MS / MS analysis, a predetermined voltage (a voltage obtained by superimposing a high-frequency voltage and a DC voltage) is applied from the power supply to each rod electrode of the pre-stage quadrupole mass filter 2231. Among the various ions sent to the pre-stage quadrupole mass filter 2231, only ions having a specific mass-to-charge ratio corresponding to the voltage applied to each rod electrode of the pre-stage quadrupole mass filter 2231 pass through the filter 2231. These ions are called precursor ions.

[0022] Precursor ions are introduced into the subsequent collision cell 2232. A predetermined voltage is applied to the electrodes placed in the collision cell 2232. The precursor ions introduced into the collision cell 2232 are accelerated within the collision cell 2232 in accordance with the voltage applied to the electrodes. CID gas is also supplied into the collision cell 2232 at a predetermined pressure. As a result, the accelerated precursor ions collide with the CID gas at a predetermined collision energy and dissociate, generating multiple ions. The ions generated from the precursor ions are called product ions. In MS / MS analysis, the ion detector 23 detects the signal of the product ions.

[0023] Furthermore, the MS20 can also use only the pre-stage quadrupole mass filter 2231 as the mass separation unit. In this case, CID gas is not supplied into the collision cell 2232, and a voltage is applied to the multipole ion guide 2233 and the subsequent quadrupole mass filter 2234 such that the ions mass-separated in the pre-stage quadrupole mass filter 2231 pass through directly. When mass spectrometry is performed under the above conditions, the ion detector 23 detects the precursor ion signal.

[0024] When analyzing components using MS20, it is possible to analyze using either product ions or precursor ions as indicators. Using product ions as indicators improves measurement accuracy but decreases measurement sensitivity compared to using precursor ions as indicators. Therefore, the user can select whether the analysis system 100 analyzes using precursor ions or product ions as indicators, depending on the amount of component in the sample. For example, consider a case where the amount of precursor ions of component A generated exceeds the amount of ions detectable by the ion detector 23. In such a case, the user switches to MS / MS analysis and analyzes component A using product ions derived from component A as indicators. This allows the amount of ions derived from component A to be kept within the range of ions detectable by the ion detector 23, and component A can be analyzed.

[0025] Returning to Figure 1, the control device 30 includes a processor 31, a memory 32, and an input / output interface (I / F) 33. These components are connected to each other via a bus so as to be able to communicate with one another. The control device 30 is configured, for example, by a general-purpose computer and is connected to the LC 10, MS 20, input device 40, and display device 50. The control device 30 controls the operation of the LC 10 and MS 20.

[0026] The processor 31 is an example of an electrical circuit and controls the operation of the control device 30 by executing a given program. The program executed by the processor 31 may be stored in the memory 32 or in a storage device (not shown) located outside the control device 30. The processor 31 is, for example, a CPU (Central Processing Unit).

[0027] Memory 32 can store programs executed by the processor 31, chromatogram data, and component measurements. Chromatogram data is created by the control device 30 based on the output of the ion detector 23. The chromatogram data includes a total ion chromatogram, where the sum of all ions generated by the ion source is represented on the vertical axis and time on the horizontal axis, and a chromatogram, where the signals of ions with a specific mass-to-charge ratio are represented on the vertical axis and time on the horizontal axis. Component measurements are calculated, for example, from the peak area values ​​and peak heights in the chromatogram data. Memory 32 stores the moisture content of the cultured meat, the amount of components per unit wet weight of the cultured meat, the amount of components per unit dry weight of the cultured meat, and an evaluation of the quality of the cultured meat, all linked together. Memory 32 includes volatile memory (e.g., RAM (Random Access Memory)) and non-volatile memory (e.g., ROM (Read Only Memory), hard disk drives, and solid-state drives). The above program may be stored in an external storage device accessible by the processor 31.

[0028] The input / output interface 33 is an interface for exchanging various types of data between the processor 31 and the devices connected to the input / output interface 33. LC10, MS20, input device 40, and display device 50 are connected to the input / output interface 33. The input / output interface 33 is implemented, for example, by terminal blocks, connectors, and network adapters. Data transfer via the input / output interface 33 may be performed wirelessly, such as via Bluetooth® or wireless LAN, or via a wired connection using USB (Universal Serial Bus).

[0029] The input device 40 receives information from the user to the control device 30. This information includes, for example, the analysis conditions for LC10, the analysis conditions for MS20, the total number of samples, the type of sample, the components to be analyzed, the moisture content of the cultured meat, the wet weight of the cultured meat, the dry weight of the cultured meat, the moisture content of the meat, the wet weight of the meat, and the dry weight of the meat. The input device 40 is composed of, for example, a touch panel, a mouse, and a keyboard.

[0030] The display device 50 displays information according to the instructions of the control device 30. This information includes, for example, chromatogram data, the amount of components, the moisture content of cultured meat, the amount of components per unit wet weight of cultured meat, the amount of components per unit dry weight of cultured meat, the moisture content of meat, the amount of components per unit wet weight of meat, and the amount of components per unit dry weight of meat. The display device 50 is composed of, for example, a liquid crystal display capable of displaying images.

[0031] [Method for Creating Cultured Meat] Next, we will explain how to create cultured meat. Figure 3 is a schematic diagram illustrating the method for creating cultured meat. Figure 3 illustrates the method for creating cultured meat derived from cattle as an example.

[0032] Referring to Figure 3, the user collects muscle tissue (Lean) and adipose tissue (Fat) from livestock.

[0033] Next, the user collects bovine satellite cells (bSCs) from the collected Lean and bovine adipose-derived stem cells (bADSCs) from the Fat. The user then cultures the collected cells to increase their number.

[0034] The user uses a 3D printer to place a culture medium containing cultured cells in a long, narrow shape within a gel-like support. This allows for the creation of muscle fibers and fat fibers.

[0035] The user arranges the created muscle and fat fibers three-dimensionally, bonding the fibers together to form a structure. This completes a block-shaped cultured meat (CM). In one example, the weight of the cultured meat produced by the above method is approximately 10-20 mg.

[0036] [Comparative Example] Cultured meat is attracting attention as one of the sustainable alternatives to meet the increasing demand for meat. Cultured meat, which is artificial meat made from animal cells, is expected to meet the expected increase in demand for meat in the future.

[0037] To ensure the safety of cultured meat as a food product and to maintain and improve its quality, it is important to evaluate the component amounts and physical properties of the cultured meat produced.

[0038] Regarding the evaluation of the quality of cultured meat, it is conceivable to apply conventional meat evaluation methods. However, according to the conventional meat evaluation method disclosed in Non-Patent Document 1, for example, it is necessary to prepare a sample of about 300 to 500 g for evaluation. The reason why such a large sample is required for evaluation is thought to be to prevent the bias in components in a part of the meat from affecting the analytical results used in the evaluation. For example, if a part with a relatively high lipid content is used for analysis, the analytical result for the entire meat may result in a high lipid content. Such an analytical result cannot be said to accurately reflect the amount of components (lipids) in the entire meat and is undesirable as an analytical result for the meat. Therefore, it is expected that by uniformly mixing several hundred g of the sample in a mixer or the like and using a portion of it for component quantity analysis, it will be possible to prevent the bias in components in a part of the meat from affecting the analytical results.

[0039] Preparing several hundred grams of cultured meat for quality evaluation can be time-consuming and costly for users, and the amount of cultured meat that users can prepare for evaluation is typically only a few milligrams to tens of milligrams. Even if this amount of cultured meat is mixed using a mixer or similar device according to conventional meat evaluation methods, it is difficult to achieve uniform mixing because the blades of the mixer cannot finely grind the cultured meat. Thus, it is difficult to use evaluation methods that have been used for meat to evaluate cultured meat. Therefore, there is a need to develop evaluation methods for assessing the quality of cultured meat.

[0040] [Method for Evaluating the Components of Cultured Meat] The method for evaluating cultured meat in this embodiment includes a step of freeze-drying and crushing the cultured meat into a powder when analyzing it. This step allows for the preparation of a uniform sample for analysis even when using a small amount of cultured meat. Furthermore, since the water content in the cultured meat is removed by freeze-drying, a uniform sample for analysis can be prepared regardless of the water content in the cultured meat. By providing the uniformly mixed sample to an analytical device, the user can analyze the components contained in the cultured meat and evaluate the cultured meat based on the analysis results.

[0041] The cultured meat analysis method in this embodiment allows for the preparation of a uniformly mixed analytical sample with a smaller sample volume compared to conventional meat evaluation methods, and enables the acquisition of analytical results for the components of that sample. The cultured meat evaluation method in this embodiment utilizes the analytical results calculated by the above analysis method. Therefore, the cultured meat evaluation method in this embodiment allows for the evaluation of the quality of cultured meat based on component quantity data obtained with a smaller sample volume compared to conventional meat evaluation methods.

[0042] The method for evaluating cultured meat in this embodiment includes the steps described below. <Measurement of the wet weight of cultured meat> The method for evaluating cultured meat in this embodiment includes the step of measuring the wet weight of cultured meat. The wet weight of cultured meat is the weight before freeze-drying the cultured meat and contains moisture. For example, the user measures the wet weight of the cultured meat after washing the surface of the formed cultured meat with a phosphate buffer solution and wiping off the surface moisture with a paper wiper for 1 minute. For example, the user measures the wet weight using an electronic balance. In this embodiment, the wet weight corresponds to the first weight.

[0043] <Freeze-drying> The method for evaluating cultured meat in this embodiment includes the step of freeze-drying the cultured meat. Freeze-drying is a technique for removing moisture in a specimen by sublimation of ice by reducing the pressure at a vacuum level that does not melt the frozen specimen. The user freezes the cultured meat after measuring the wet weight and subjects it to a freeze-dryer. Since freeze-drying can remove the moisture in the cultured meat while maintaining the frozen state of the cultured meat, there is less risk of denaturation of the components in the cultured meat compared to drying by heating or dehydration using centrifugal force.

[0044] <Measurement of the dry weight of cultured meat after freeze-drying> The method for evaluating cultured meat in this embodiment includes the step of measuring the dry weight of the cultured meat. The dry weight of the cultured meat is the weight of the cultured meat after freeze-drying. For example, the user measures the dry weight using an electronic balance. In this embodiment, the dry weight corresponds to the second weight.

[0045] <Crushing of cultured meat after freeze-drying> The method for evaluating cultured meat in this embodiment includes the step of crushing the cultured meat after freeze-drying into a powder. By freeze-drying, the cultured meat becomes a porous dried product with cavities in the part where the evaporated moisture was. When the cultured meat in this state is physically crushed, it can be made into a powder. The crushing of the cultured meat is performed, for example, by putting 3 stainless steel balls (diameter 5 mm) into a container containing the cultured meat after freeze-drying and shaking it at a shaking frequency of 30 Hz for 3 minutes. By making it into a powder, it can be uniformly mixed.

[0046] <Measurement of Component Amounts>The method for evaluating cultured meat in the present embodiment includes a step of measuring components in an analysis sample by the analysis system 100. The user provides an analysis sample prepared from powdered cultured meat to the analysis system 100. Since the concentrations of components in the analysis sample are different, the user prepares and analyzes the prepared analysis sample and a sample obtained by diluting it. Assuming the prepared sample as the stock solution, the user prepares, for example, a sample diluted 10-fold and a sample diluted 100-fold from the stock solution.

[0047] In this step, the amount of the component is analyzed using chromatogram data generated based on the output of the ion detector 23. Specifically, the amount of the component is calculated based on the area value and peak height in the chromatogram of the ions derived from the component. When a standard sample for calibration curve creation can be prepared, the amount of substance and weight of the component are calculated based on the area value, peak height, and calibration curve.

[0048] When it is difficult to create a calibration curve for a predetermined component, the cultured meat is evaluated for the predetermined component by comparing the area value and peak height of the predetermined component in the reference sample. For example, assume that the reference sample is a sample derived from meat. At this time, when the area value of component B per unit wet weight of the cultured meat is larger than the area value of component B per unit wet weight of the meat, it can be evaluated that component B is contained more in the cultured meat than in the meat.

[0049] As described above, when comparing multiple analyzed samples using area values ​​and peak heights as indicators of component quantities, it is preferable that the samples being compared be analyzed by continuous analysis. Continuous analysis means analyzing samples using the same instrument without any time intervals between them. For example, if one sample is measured immediately before the next, the two samples can be said to have been analyzed continuously. Continuous analysis may also be performed in the same batch. Specifically, if the difference in the start times of the analysis of two samples is within 72 hours, the two samples can be said to have been analyzed continuously. In the analysis system 100, differences in conditions such as temperature and humidity during analysis may affect the measurement results. Even if the area in the chromatogram is the same, the amount of component may differ from analysis to analysis. By performing continuous analysis, the differences in the above conditions can be reduced. Therefore, when using area values ​​and peak heights as indicators of component quantities, it is preferable that the samples to be compared be analyzed continuously.

[0050] Furthermore, if there is a component C whose ion detection amount is low in cultured meat and low in meat, the precursor ion may be used as the indicator in the analysis of cultured meat, and the product ion may be used as the indicator in the analysis of meat. By doing so, sensitivity can be improved in cultured meat where the ion detection amount is low, preventing the amount of component C ions from falling below the detection limit, and sensitivity can be reduced in meat where the ion detection amount is high, preventing the amount of component C ions from exceeding the detectable amount. Note that when comparing multiple analyzed samples using area value and peak height as the amount of the component, the area value in the chromatogram of the product ion and the area value in the chromatogram of the precursor ion cannot be directly compared, so it is necessary to correct the area value of the precursor ion to the area value of the product ion. Components whose ion detection amount is low in cultured meat and high in meat include, for example, acetylcarnitine, alanine, carnitine, carnosine, creatine, and inosine. When analyzing these components in cultured meat, precursor ions are used as indicators, while when analyzing these components in meat, product ions are used as indicators.

[0051] <Calculation of moisture content and evaluation based on moisture content> The method for evaluating cultured meat in this embodiment includes the steps of calculating the moisture content of the cultured meat and evaluating the cultured meat based on the moisture content. The moisture content of the cultured meat can be calculated using the wet weight and dry weight of the cultured meat. For example, the moisture content of cultured meat is calculated as (wet weight - dry weight) / wet weight.

[0052] Cultured meat tends to have a higher water content than edible meat. Generally, as the differentiation of the cells that make up cultured meat progresses and the degree of differentiation increases, the water content in the cultured meat decreases. A higher degree of differentiation means that the cells are closer to those that make up edible meat. Therefore, the water content of cultured meat serves as an indicator when evaluating its quality. This quality, for example, indicates the degree of differentiation and represents the degree of similarity between cultured meat cells and edible meat cells.

[0053] Specifically, the moisture content of the cultured meat is compared with a first reference value, and the quality of the cultured meat is evaluated based on the results of this comparison. For example, if the moisture content of the cultured meat is below the first reference value, it is evaluated as having a high degree of differentiation. The first reference value is, for example, the moisture content of meat obtained by cutting meat to the size of the cultured meat and processing it in the same way. The first reference value may be determined by the user.

[0054] <Calculation of component amounts per unit wet weight and evaluation thereunder> The method for evaluating cultured meat in this embodiment includes the steps of calculating the component amounts per unit wet weight of the cultured meat and evaluating the cultured meat based on the component amounts. The component amounts per unit wet weight of the cultured meat can be calculated using the wet weight of the cultured meat obtained in <Measurement of wet weight of cultured meat> and the component amounts of the cultured meat obtained in <Measurement of component amounts>. The unit wet weight is, for example, 100 g and 1 g.

[0055] Furthermore, the amount of a component per unit wet weight serves as an indicator for evaluating cultured meat. Specifically, the amount of a component per unit wet weight of cultured meat is compared with a second standard value, and the quality of the cultured meat is evaluated based on the results of this comparison. This quality refers to the nutritional value, sweetness, umami, bitterness, and sourness of the cultured meat. For example, if the amount of a component per unit wet weight of cultured meat is equal to or greater than the second standard value, the cultured meat is evaluated as having high nutritional value. Also, regarding the component that exhibits sweetness, if the amount of the component in the cultured meat is equal to or greater than the second standard value, the cultured meat can be evaluated as sweet. The second standard value is, for example, the amount of the component per unit wet weight of meat obtained by cutting meat to the size of cultured meat and processing it in the same way. If the second standard value is analytical data for meat, for example, if the amount of a component per unit wet weight of cultured meat is within a predetermined range relative to the second standard value, the cultured meat may be evaluated as containing the same amount of the component as meat. The predetermined range is, for example, ±80%. The second reference value may be determined by the user. The second reference value may also be determined for each component. Furthermore, the quality of the cultured meat may be evaluated by comparing the amount of each component per unit wet weight of the cultured meat with the corresponding second reference value. For example, if there are 20 components, the cultured meat may be evaluated as having high nutritional value if the amount of 15 of the components per unit wet weight of the cultured meat is equal to or greater than the second reference value.

[0056] <Calculation of the amount of components per unit dry weight and evaluation thereunder> The method for evaluating cultured meat in this embodiment includes the steps of calculating the amount of components per unit dry weight of the cultured meat and evaluating the cultured meat based on the amount of said components. The amount of components per unit dry weight of the cultured meat can be calculated using the dry weight of the cultured meat obtained in <Measurement of the dry weight of the cultured meat> and the amount of components of the cultured meat obtained in <Measurement of the amount of components>. The unit dry weight is, for example, 100 g and 1 g.

[0057] Furthermore, the amount of components per unit dry weight serves as an indicator for evaluating cultured meat. Specifically, the amount of components per unit dry weight of cultured meat is compared with a third standard value, and the quality of the cultured meat is evaluated based on the results of this comparison. This quality refers to the nutritional value, sweetness, umami, bitterness, and sourness of the cultured meat. For example, if the amount of components per unit dry weight of cultured meat is equal to or greater than the third standard value, the cultured meat is evaluated as having high nutritional value. Also, regarding components that exhibit sweetness, if the amount of components in cultured meat is equal to or greater than the third standard value, the cultured meat can be evaluated as sweet. The third standard value is, for example, the amount of the relevant component per unit dry weight of meat obtained by cutting meat to the size of cultured meat and processing it in the same way. If the third standard value is analytical data for meat, for example, if the amount of components per unit dry weight of cultured meat is within a predetermined range relative to the third standard value, the cultured meat may be evaluated as containing the same amount of the relevant component as meat. The predetermined range is, for example, ±80%. The third criterion value may be determined by the user. The third criterion value may also be determined for each component. Alternatively, the quality of cultured meat may be evaluated by comparing the amount of each component per unit dry weight of cultured meat with the corresponding third criterion value. For example, if there are 20 components, cultured meat may be evaluated as having high nutritional value if the amount of 15 of the components per unit dry weight of cultured meat is equal to or greater than the third criterion value.

[0058] The moisture content, component amounts per unit wet weight, and evaluation of the cultured meat obtained are linked and stored in memory 32. The stored data, including the moisture content of the cultured meat, are used as training data to predict the component amounts per unit wet weight and the evaluation of the cultured meat.

[0059] [Flowchart for Evaluating Cultured Meat] The method for evaluating cultured meat in this embodiment will now be explained. Figure 4 is a flowchart illustrating the method for evaluating cultured meat. The user prepares an analytical sample from the cultured meat. The user then uses the analysis system 100 to analyze the components contained in the analytical sample. The user can evaluate the cultured meat based on the measured values ​​of the obtained components.

[0060] First, in step S100, the user performs pre-treatment on the cultured meat to be evaluated. Figure 5 is a flowchart of the pre-treatment subroutine in step S100. The pre-treatment process will be explained with reference to Figure 5.

[0061] Referring to Figure 5, in step S102, the user prepares the cultured meat to be evaluated. An example of the weight of the prepared cultured meat is approximately 20 mg.

[0062] In step S104, the user measures the wet weight of the cultured meat prepared in step S102.

[0063] In step S106, the user freezes the cultured meat at -80°C. In step S108, the user freeze-dries the cultured meat frozen in step S106.

[0064] In step S110, the user measures the dry weight of the freeze-dried cultured meat. In step S112, the user places three stainless steel balls (5 mm in diameter) into container (A) containing the freeze-dried cultured meat and shakes it at a frequency of 30 Hz for 3 minutes. This causes the freeze-dried cultured meat to become powdery. After obtaining the powdery cultured meat, the user terminates the pre-processing subroutine and returns the process to Figure 4.

[0065] Returning to Figure 4, in step S200, the user prepares the sample for analysis to be used in the analysis system 100. Figure 6 is a flowchart of the sample preparation subroutine. Referring to Figure 6, the process of preparing the sample for analysis will be explained.

[0066] Referring to Figure 6, in step S202, the user adds 10 μL (equivalent to 5 μg) of 0.5 mg / mL monofluoroethanesulfonic acid (MES) internal standard solution to container (A) containing the sample to be analyzed. In this embodiment, the sample to be analyzed is the powdered cultured meat obtained in step S112.

[0067] In step S204, the user puts 900 μL of methanol:water:chloroform (2.5:1:1) mixed solution into container (A) and mixes it.

[0068] In step S206, the user heats container (A) to 37°C and shakes it at a speed of 1200 rpm (revolutions per minute) for 30 minutes.

[0069] In step S208, the user cools container (A) to 4°C and centrifuges it with a relative centrifugal force of 15,000 g for 5 minutes. As a result, the protein components contained in container (A) settle at the bottom of container (A).

[0070] In step S210, the user transfers 630 μL of the supernatant from container (A) after centrifugation in step S208 into container (B).

[0071] In step S212, the user puts 280 μL of ultrapure water into container (B) and mixes for 2 minutes.

[0072] In step S214, the user cools container (B) to 4°C and centrifuges it with a relative centrifugal force of 15,000 g for 3 minutes. This separates the liquid in container (B) into an upper layer and a lower layer.

[0073] In step S216, the user adds 500 μL of the upper layer after centrifugation in step S214 to a protein-removing centrifugal filter. The protein-removing centrifugal filter is a filter that does not allow protein components to pass through.

[0074] In step S218, the user cools the protein removal centrifugal filter to 4°C and rotates it for 60 minutes with a relative centrifugal force of 15,000 g. A container (C) is provided at the bottom of the filter, and the liquid that has passed through the filter due to the centrifugal force from the rotation is stored in container (C).

[0075] In step S220, the user covers the opening of container (C) in which the liquid that has passed through the protein-removing centrifugal filter in step S218 is stored with a PTFE (polytetrafluoroethylene) membrane filter.

[0076] In step S222, the user heats the container (C) covered with the membrane filter to 40°C using a centrifugal evaporator and processes it twice for 90 minutes at a rotation speed of 2500 rpm. This process allows the methanol in the container (C) to be evaporated and removed.

[0077] In step S224, the user freezes the liquid in container (C) at -80°C and freeze-dries container (C).

[0078] In step S226, the user adds 100 μL of ultrapure water to the freeze-dried container (C) and mixes it. The resulting solution is referred to as the stock solution.

[0079] In step S228, the user prepares a 10-fold dilution and a 100-fold dilution of the stock solution obtained in step S226 by diluting the stock solution with ultrapure water. After obtaining the stock solution and dilutions, the user terminates the sample preparation subroutine and returns the process to Figure 4.

[0080] Returning to Figure 4, in step S300, the analysis system 100 analyzes the amount of components contained in the sample for analysis. Figure 7 is a flowchart of the analysis subroutine. Referring to Figure 7, the process of analyzing the amount of components contained in the sample for analysis will be explained. Note that in Figure 7, step S302 is performed by the user, and steps S304 to S324 are performed by the analysis system 100.

[0081] Referring to Figure 7, in step S302, the user introduces the analytical sample (undiluted solution, a 10-fold dilution, and a 100-fold dilution) into the LC10 from the injector 13.

[0082] In step S304, the analysis system 100 receives analysis conditions from the user. In step S306, the analysis system 100 separates the introduced sample into its components based on the analysis conditions received in step S304.

[0083] In step S308, the analysis system 100 introduces the components separated in LC10 into MS20.

[0084] In step S310, the analysis system 100 performs mass spectrometry of the components introduced from LC10 using MS20.

[0085] In step S312, the analysis system 100 creates a chromatogram based on the mass-to-charge ratio of the ions detected by the MS20.

[0086] In step S314, the analysis system 100 calculates the amount of each component in the sample based on the chromatogram created in step S312. The amount of each component may be expressed in terms of amount of substance (mol), mass, and weight using a calibration curve, or it may be expressed as the area value and peak height in the chromatogram.

[0087] In step S316, the analysis system 100 receives the wet weight of the cultured meat obtained in step S104 and the dry weight of the cultured meat obtained in step S110 from the user.

[0088] In step S318, the analysis system 100 calculates the moisture content of the cultured meat using the wet weight and dry weight of the cultured meat obtained in step S316. The moisture content of the cultured meat is calculated, for example, as: Moisture content = (wet weight - dry weight) / wet weight.

[0089] In step S320, the analysis system 100 calculates the amount of the component per unit wet weight of the cultured meat using the amount of the component calculated in step S314 and the wet weight of the cultured meat obtained in step S316. The unit wet weight is, for example, 100 g, and the amount of the component per unit wet weight of the cultured meat is the amount of that component contained in 100 g of the wet weight of the cultured meat.

[0090] In step S322, the analysis system 100 calculates the amount of the component per unit dry weight of the cultured meat using the amount of the component calculated in step S314 and the dry weight of the cultured meat obtained in step S316. The unit dry weight is, for example, 100 g, and the amount of the component per unit dry weight of the cultured meat is the amount of that component contained in 100 g of dry weight of the cultured meat.

[0091] In step S324, the analysis system 100 stores the moisture content of the cultured meat calculated in step S318, the amount of components per unit wet weight of the cultured meat calculated in step S320, and the amount of components per unit dry weight of the cultured meat calculated in step S322, linked together, in the memory of the control unit 540. After that, the analysis system 100 terminates the analysis subroutine and returns the process to Figure 4.

[0092] Returning to Figure 4, in step S400, the user evaluates the cultured meat based on the moisture content of the cultured meat, the amount of components per unit wet weight of the cultured meat, and the amount of components per unit dry weight of the cultured meat. Figure 8 is a flowchart of the evaluation subroutine. Referring to Figure 8, the process by which the user evaluates the cultured meat will be explained.

[0093] Referring to Figure 8, in step S402, the user sets a first reference value used when evaluating the moisture content of the cultured meat, a second reference value used when evaluating the amount of components per unit wet weight of the cultured meat, and a third reference value used when evaluating the amount of components per unit dry weight of the cultured meat.

[0094] Figure 9 is a flowchart of the reference value setting subroutine (S402). The process of setting the reference value will be explained with reference to Figure 9. In Figure 9, steps that are the same as those in Figure 4 are denoted by the same reference numerals and their explanation is omitted.

[0095] Referring to Figure 9, in step S100A, the user performs pre-processing on the meat to be measured. Step S100A is the same as step S102, except that the user prepares cultured meat, while in step S100A the user cuts the meat to approximately the same size as the cultured meat. In other words, the same processing performed on the cultured meat is performed on the meat.

[0096] In step S452, the user sets the moisture content of the meat as the first reference value. In step S454, the user sets the amount of components per unit of wet weight of the meat as the second reference value.

[0097] In step S456, the user sets the amount of the component per unit dry weight of the meat as the third reference value.

[0098] Returning to Figure 8, in step S404, the user compares the moisture content of the cultured meat with the first reference value.

[0099] In step S406, the user evaluates the cultured meat using the comparison results from step S404.

[0100] In step S408, the user compares the amount of the component per unit wet weight of the cultured meat with the second reference value.

[0101] In step S410, the user evaluates the cultured meat using the comparison results from step S408.

[0102] In step S412, the user compares the amount of the component per unit dry weight of the cultured meat with the third reference value.

[0103] In step S414, the user evaluates the cultured meat using the comparison results from step S412. Afterward, the process shown in Figure 8 is terminated.

[0104] Steps S404, S410, and S414 describe the user evaluating the cultured meat, but are not limited to this, and the control device 30 may perform processing for evaluating the cultured meat.

[0105] The cultured meat analysis method described in this disclosure allows for the preparation of a uniformly mixed analytical sample with a smaller sample volume compared to conventional meat evaluation methods, and enables the acquisition of analytical results for the components of that analytical sample. The cultured meat evaluation method described in this disclosure utilizes the analytical results calculated by the above analysis method. Therefore, the cultured meat evaluation method described in this disclosure allows for the evaluation of the quality of cultured meat based on component quantity data obtained with a smaller sample volume compared to conventional meat evaluation methods.

[0106] Furthermore, if the analysis system 100 is equipped with GC-MS or GC-MS / MS, the analyte is a volatile component. In this case, the aroma of cultured meat can be evaluated using the analysis system 100.

[0107] When analyzing the aroma of cultured meat, the user packs individual cultured meat fibers into glass vials and heats them at 200°C for 30 minutes using a fume hood heating device. After that, the vials are left to stand at room temperature for 30 minutes, allowing volatile components to diffuse into the glass vials. The user then concentrates the volatile components in the glass vials using solid-phase micro-extraction (SPME) and introduces them into the analysis system 100.

[0108] When the analysis results of meat are to be used as the second and third reference values, the meat will be crushed with dry ice and freeze-dried before being analyzed.

[0109] [Method for Evaluating the Texture of Cultured Meat] Evaluating the texture based on physical properties is also important for evaluating cultured meat. In this embodiment, the texture of cultured meat is evaluated by preparing meat cut to match the shape of the cultured meat and comparing the elastic modulus of the meat with the elastic modulus of the cultured meat. With this method of evaluating the texture of cultured meat, the meat is prepared according to the amount of cultured meat to be evaluated, so the texture of cultured meat can be evaluated regardless of the amount of cultured meat.

[0110] The method for evaluating the texture of cultured meat in this embodiment includes a step of evaluating the texture of cultured meat based on the elastic modulus of a sample of livestock meat cut into a fiber shape similar to that of cultured meat, as a reference.

[0111] This embodiment describes a method for evaluating the texture of cultured meat. Figure 10 is a flowchart illustrating the method for evaluating the texture of cultured meat. The user prepares samples for analysis from cultured meat and raw meat. The user then measures the elastic modulus of each sample. The user compares the elastic modulus of the raw meat with that of the cultured meat. Based on the obtained comparison results, the user can evaluate the texture of the cultured meat.

[0112] Referring to Figure 10, first, in step S500, the user prepares the cultured meat to be evaluated and the meat to be compared.

[0113] In step S502, the user cuts the meat into fibrous shapes to match the fibrous shape of the cultured meat.

[0114] In step S504, the user washes the cultured meat and meat with phosphate buffer and leaves them on a paper wiper for 30 seconds. This removes moisture from the surface of the cultured meat and meat.

[0115] In step S506, the user lays several cultured meat fibers in a container, according to the size of the container being used. For example, three cultured meat fibers are laid in the container.

[0116] In step S508, the user compresses the cultured meat at 1 mm / second using a plunger with a spherical tip to measure its elastic modulus.

[0117] In step S510, the user lays the cut meat in a container to fit its size.

[0118] In step S512, the user compresses the meat at 1 mm / second using a plunger with a spherical tip to measure its elastic modulus.

[0119] In step S514, the user compares the elastic modulus of cultured meat with that of meat. In step S516, the user evaluates the cultured meat based on the comparison results from step S514. For example, if the elastic modulus of cultured meat is greater than or equal to that of meat, the user evaluates that the cultured meat has a harder texture than meat; if the elastic modulus of cultured meat is less than or equal to that of meat, the user evaluates that the cultured meat has a softer texture than meat; and if the elastic modulus of cultured meat is equal to that of meat, the user evaluates that the cultured meat has a texture equivalent to that of meat.

[0120] In steps S508 and S512, the height of the cultured meat and meat may not match the height of the sample stage. In such cases, the plunger was pushed in excessively, touching the sample stage on which the cultured meat and meat were placed. The displacement at the point where the measured force increased linearly was taken as the height of the cultured meat and meat, and the modulus of elasticity was calculated from the displacement rate.

[0121] According to the method for evaluating the texture of cultured meat in this embodiment, the texture of cultured meat can be evaluated regardless of the amount of cultured meat.

[0122] [Experimental Examples] The present invention will be described in more detail below with reference to examples, but the present invention is not limited to these. In the experimental examples, tissue was collected from three types of cattle (H: Holstein, C: crossbreed, W: Wagyu) and cultured meat was prepared. The amino acids contained in the cultured meat were analyzed by LC-MS / MS, and fatty acids and aroma components were analyzed by GC-MS / MS. Meat collected from each of the three types of cattle was also analyzed.

[0123] For amino acid analysis, fibers created from 20 mg of bSC were analyzed as cultured meat, and 20 mg of Lean was analyzed as meat. For fatty acid analysis, fibers created from 20 mg of bADSC were analyzed as cultured meat, and 100 mg of Fat was analyzed as meat. For aroma component analysis, fibers created from 20 mg of bADSC were analyzed as cultured meat, and 20 mg of adipose tissue was analyzed as meat.

[0124] Amino acids were measured using LC-MS / MS (LCMS-8060NX, Shimadzu Corporation). Fatty acids were measured using GC-MS / MS (GCMS-QP2020NX, Shimadzu Corporation), and aroma components were measured using GC-MS / MS (GCMS-TQ8050NX, Shimadzu Corporation). For the analysis of aroma components, AOC6000 and SPMWALLOW, which are suitable for measuring aroma components, were used. 498 types of aroma components were analyzed. In this experimental example, the analysis results for lactone, a type of sweet aroma component, are shown. The amino acid and fatty acid amounts are shown as analysis results converted to molar or weight based on a calibration curve, but for lactone, relative values ​​to the reference sample (cultured meat derived from Holstein, or Holstein fat) are shown.

[0125] The measurement conditions for LC-MS / MS were as follows: <LC conditions> The analytical column used was Discovery HS F5-3 (2.1 mm I.D. x 150 mmL, 3 μm) (Sigma-Aldrich) P / N 567503-U. Mobile phase A was 0.1% formic acid aqueous solution, and mobile phase B was 0.1% formic acid acetonitrile solution. The gradient program was B Conc. 0% (0 min) → 0% (2 min) → 25% (5 min) → 35% (11 min) → 50% (12.5 min) → 50% (16 min) → 95% (16.01 min) → 95% (19 min) → 0% (19.01 min) → 0% (30 min). The flow rate was 0.25 mL / min. The injection volume was 1 μL. The column temperature was 40°C.

[0126] The nebulizer gas flow rate was set to 3.0 L / min, the drying gas flow rate to 10.0 L / min, and the heating gas flow rate to 10.0 L / min. The interface temperature was set to 270°C. The DL temperature was set to 250°C. The heat block temperature was set to 400°C. ESI was used as the ionization mode.

[0127] Furthermore, for acetylcarnitine, alanine, carnitine, carnosine, creatine, and inosine, the detection levels of these ions are low in cultured meat and high in meat. Therefore, when analyzing these components in cultured meat, precursor ions were used as the indicator, and when analyzing these components in meat, product ions were used as the indicator. Specifically, the precursor ion of acetylcarnitine is m / z 204.1000, and the product ion of acetylcarnitine is m / z 85.0500. The precursor ion of alanine is m / z 90.1000, and the product ion of alanine is m / z 44.1000. The precursor ion of carnitine has a m / z of 162.1000, and the product ion of carnitine has a m / z of 103.0500. The precursor ion of carnosine has a m / z of 227.1000, and the product ion of carnosine has a m / z of 110.0500. The precursor ion of creatine has a m / z of 132.1000, and the product ion of creatine has a m / z of 44.0500. The precursor ion of inosine has a m / z of 269.1000, and the product ion of inosine has a m / z of 137.0500.

[0128] Figure 11 is a graph showing the amount of amino acids (nmol) per unit wet weight (1 mg) of cultured meat. The horizontal axis of Figure 11 shows cultured meat prepared using bSCs extracted from Holstein cattle, cultured meat prepared using bSCs extracted from crossbred cattle, and cultured meat prepared using bSCs extracted from Wagyu cattle, while the vertical axis shows the amount of amino acids per 1 mg of cultured meat.

[0129] Figure 11 shows that when comparing cultured meat derived from Holstein cattle with cultured meat derived from Wagyu cattle, the cultured meat derived from Wagyu cattle had lower levels of, for example, Glycine and Alane (which contribute to sweetness) and Arginine (which contributes to bitterness). From these comparison results, it can be concluded that cultured meat derived from Wagyu cattle has less sweetness and bitterness than cultured meat derived from Holstein cattle.

[0130] Thus, by using the method for evaluating the components of cultured meat according to this disclosure, the analytical data of the cultured meat to be evaluated (for example, cultured meat prepared using bSCs collected from Wagyu beef) can be compared with the analytical data of other cultured meat corresponding to the second reference value (for example, cultured meat prepared using bSCs collected from Holstein beef), and the cultured meat can be evaluated based on the comparison results.

[0131] Figure 12 is a graph showing the amount of amino acids (nmol) per unit wet weight (1 mg) of meat. The horizontal axis of Figure 11 shows Lean from Holstein, Lean from crossbreeds, and Lean from Wagyu, and the vertical axis shows the amount of amino acids per 1 mg of meat.

[0132] Referring to Figures 11 and 12, for example, cultured meat derived from Wagyu beef was shown to have lower levels of each amino acid compared to Lean Wagyu beef. From these comparison results, it can be concluded that cultured meat derived from Wagyu beef has lower nutritional value than Wagyu beef.

[0133] Thus, by using the method for evaluating the components of cultured meat according to this disclosure, the analytical data of the cultured meat to be evaluated (for example, cultured meat prepared using bSCs collected from Wagyu beef) can be compared with the analytical data of meat corresponding to the second reference value (for example, lean Wagyu beef), and the cultured meat can be evaluated based on the comparison results.

[0134] Figure 13 is a graph showing the amount of fatty acids (μg) per unit wet weight (1g) of cultured meat. The horizontal axis of Figure 13 shows cultured meat prepared using bADSCs extracted from Holstein cattle, cultured meat prepared using bADSCs extracted from crossbred cattle, and cultured meat prepared using bADSCs extracted from Wagyu cattle, while the vertical axis shows the weight of fatty acids per 1mg of cultured meat.

[0135] In Figure 13, no statistically significant differences were observed when comparing cultured meat derived from Holstein cattle with cultured meat derived from Wagyu cattle. Based on these results, it can be concluded that cultured meat derived from Wagyu cattle is no different from cultured meat derived from Holstein cattle in terms of fatty acids.

[0136] Thus, by using the method for evaluating the components of cultured meat according to this disclosure, the analytical data of the cultured meat to be evaluated (for example, cultured meat prepared using bACSC taken from Wagyu beef) can be compared with the analytical data of other cultured meat corresponding to the second reference value (for example, cultured meat prepared using bADSC taken from Holstein beef), and the cultured meat can be evaluated based on the comparison results.

[0137] Figure 14 is a graph showing the amount of fatty acids (μg) per unit wet weight (1 mg) of meat. The horizontal axis of Figure 14 shows the fat content from Holstein, crossbred cattle, and Wagyu cattle, while the vertical axis shows the weight of fatty acids per 1 mg of meat.

[0138] Referring to Figures 13 and 14, for example, cultured meat derived from Wagyu beef was shown to have lower amounts of each fatty acid compared to Wagyu beef fat. From these comparison results, it can be concluded that cultured meat derived from Wagyu beef has lower nutritional value than Wagyu beef.

[0139] Thus, by using the method for evaluating the components of cultured meat according to this disclosure, the analytical data of the cultured meat to be evaluated (for example, cultured meat prepared using bADSC taken from Wagyu beef) can be compared with the analytical data of meat corresponding to the second reference value (for example, Fat of Wagyu beef), and the cultured meat can be evaluated based on the comparison results.

[0140] Figure 15 is a graph showing the amount of lactone per unit wet weight of cultured meat. In Figure 15, the amount of lactone per unit wet weight of each cultured meat is shown as a relative value to the analysis data of cultured meat prepared using bADSC collected from Holstein cows.

[0141] In Figure 15, no statistically significant differences were observed when comparing cultured meat derived from Holstein cattle with cultured meat derived from Wagyu cattle. From these comparison results, it can be concluded that cultured meat derived from Wagyu cattle is no different from cultured meat derived from Holstein cattle in terms of lactones, which are responsible for the sweet aroma.

[0142] Thus, by using the method for evaluating the components of cultured meat according to this disclosure, the analytical data of the cultured meat to be evaluated (for example, cultured meat prepared using bACSC taken from Wagyu beef) can be compared with the analytical data of other cultured meat corresponding to the second reference value (for example, cultured meat prepared using bADSC taken from Holstein beef), and the cultured meat can be evaluated based on the comparison results.

[0143] Figure 16 is a graph showing the amount of lactones per unit wet weight of meat. In Figure 16, the amount of lactones per unit wet weight of each meat is shown as a relative value to the analysis data for Holstein fat.

[0144] Figure 16 shows that when comparing Holstein fat and Wagyu fat, Wagyu fat contains more of all 13 types of lactones than Holstein fat. From this comparison, it can be concluded that Wagyu beef contains more lactones, which are responsible for the sweet aroma, than Holstein beef.

[0145] Furthermore, in the experimental examples, some components were found to be present in higher concentrations in cultured meat than in natural meat. One such component is adenosine.

[0146] Thus, by using the method for evaluating the components of cultured meat described herein, it is possible to compare the analytical data of meat with the analytical data of other meats and evaluate the meat based on the comparison results.

[0147] [Aspects] The above-described exemplary embodiments will be understood by those skilled in the art to be specific examples of the following aspects.

[0148] (Section 1) An analytical method in one embodiment may include the steps of obtaining a first weight of cultured meat, crushing the freeze-dried cultured meat into powder, preparing an analytical sample using the powder, obtaining the amount of components contained in the cultured meat using the results of measuring the sample with an analytical device, and calculating the amount of the components per unit wet weight of the cultured meat using the first weight and the amount of the components.

[0149] According to the analytical method described in paragraph 1, the amount of components contained in cultured meat can be obtained with a smaller sample volume compared to the evaluation method for meat. Users can evaluate the cultured meat using the amount of components contained in it.

[0150] (Clause 2) The analytical method described in paragraph 1 may further include the steps of obtaining a second weight of the freeze-dried cultured meat and calculating the amount of the component per unit dry weight of the cultured meat using the second weight and the amount of the component.

[0151] According to the analysis method described in Section 2, the user can obtain the amount of components per unit dry weight of cultured meat. The user can then evaluate the cultured meat using the amount of components per unit dry weight. Note that the amount of components per unit wet weight decreases as the moisture content of the cultured meat increases. The amount of components per unit dry weight is not affected by the moisture content of the cultured meat. Therefore, by evaluating the cultured meat using the amount of components per unit dry weight, the user can evaluate the cultured meat while excluding the influence of moisture content.

[0152] (Clause 3) The analytical method described in paragraph 1 may further include the steps of obtaining a second weight of the freeze-dried cultured meat and calculating the moisture content of the cultured meat using the first and second weights.

[0153] According to the analysis method described in paragraph 3, the user can obtain the moisture content of cultured meat. The user can then evaluate the cultured meat using its moisture content.

[0154] (Clause 4) An evaluation method in one embodiment may include the steps of comparing the moisture content of cultured meat calculated by the analytical method described in paragraph 3 with a first reference value, and evaluating the quality of the cultured meat using the results of the comparison step.

[0155] According to the evaluation method described in Section 4, users can compare the moisture content of cultured meat with a first reference value and evaluate the quality of the cultured meat based on the results. The quality of cultured meat is, for example, the degree of differentiation, which is one indicator of how similar the cells that make up the cultured meat are to the cells that make up meat.

[0156] (Clause 5) The evaluation method described in paragraph 4 may further include the steps of obtaining a third weight of meat, obtaining a fourth weight of freeze-dried meat, calculating the moisture content of the meat using the third and fourth weights, and determining the first reference value based on the moisture content of the meat.

[0157] According to the evaluation method described in paragraph 5, cultured meat can be evaluated based on the moisture content of meat obtained by analyzing it using the same method as cultured meat.

[0158] (Clause 6) An evaluation method in one embodiment may include the steps of comparing the amount of a component per unit wet weight of cultured meat calculated by the analytical method described in paragraph 1 with a second reference value, and evaluating the quality of the cultured meat using the results of the comparison step.

[0159] According to the evaluation method described in Section 6, users can compare the amount of a component per unit wet weight of cultured meat with the second reference value and evaluate the quality of the cultured meat based on the result. The quality of cultured meat includes, for example, nutritional value, sweetness, umami, bitterness, and sourness. For example, if the amount of a component exhibiting sweetness in the cultured meat is equal to or greater than the second reference value, the cultured meat can be evaluated as sweet.

[0160] (Clause 7) The evaluation method described in paragraph 6 may further include the steps of: preparing meat of approximately the same size as the cultured meat; obtaining a fifth weight of the meat; crushing the freeze-dried meat into powder; preparing a reference sample using the powdered meat; obtaining the amount of the component contained in the meat using the results of measuring the reference sample with an analytical device; calculating the amount of the component per unit wet weight of the meat using the fifth weight and the amount of the component; and determining the second reference value based on the amount of the component per unit wet weight of the meat.

[0161] According to the evaluation method described in paragraph 7, cultured meat can be evaluated based on the amount of components per unit wet weight of meat obtained by analysis using the same method as for cultured meat.

[0162] (Clause 8) In the evaluation method described in paragraph 7, the amount of the component contained in the cultured meat and the amount of the component contained in the meat may be obtained by continuous analysis.

[0163] According to the evaluation method described in Section 8, the analytical data of cultured meat can be compared with comparable data without correcting for interanalytical errors.

[0164] (Clause 9) In the evaluation method described in either paragraph 7 or 8, when measured by a liquid chromatograph tandem mass spectrometer (LC-MS / MS) or a gas chromatograph tandem mass spectrometer (GC-MS / MS), the step of obtaining the amount of a component contained in the cultured meat may include the step of quantifying the component using a signal derived from the precursor ion of the component as an indicator, and the step of obtaining the amount of a component contained in the meat may include the step of quantifying the component using a signal derived from the product ion of the component as an indicator.

[0165] According to the analytical method described in paragraph 9, when the amount of a given component in meat is greater than the amount in cultured meat, the detection sensitivity can be reduced by analyzing the component in the meat using product ions as an indicator. This makes it possible to keep the amount of ions derived from the component below the detection limit of the ion detector, and to analyze the component in the meat.

[0166] (Clause 10) An evaluation method in one embodiment may include the steps of: obtaining a first elastic modulus obtained by measuring the elastic modulus of cultured meat; preparing meat having substantially the same shape as the cultured meat; obtaining a second elastic modulus obtained by measuring the elastic modulus of the meat; comparing the first elastic modulus and the second elastic modulus; and evaluating the quality of the cultured meat using the results of the comparison step.

[0167] According to the evaluation method described in Section 10, the user can obtain the elastic modulus of meat cut to match the shape of the cultured meat being evaluated, as well as the elastic modulus of the cultured meat itself. This allows the user to evaluate the texture of the cultured meat.

[0168] The embodiments disclosed herein should be considered in all respects to be illustrative and not restrictive. The scope of this disclosure is indicated by the claims rather than by the description of the embodiments above, and all modifications within the meaning and scope of the claims are intended to be included. Furthermore, each technique in the embodiments is intended to be practiced individually or, as far as possible, in combination with other techniques in the embodiments.

[0169] 11 Mobile phase container, 12 Liquid delivery pump, 13 Injector, 14 Flow path switching valve, 15 Column, 21 Ionization chamber, 22 Mass separation unit, 23 Ion detector, 30 Control device, 31 Processor, 32 Memory, 33 Input / Output I / F, 40 Input device, 50 Display device, 100 Analysis system, 211 ESI probe, 212 Heated capillary, 221 First intermediate vacuum chamber, 222 Second intermediate vacuum chamber, 223 Analysis chamber, 2211 Skimmer, 2212, 2221, 2233 Ion guide, 2231 Pre-stage quadrupole mass filter, 2232 Collision cell, 2234 Post-stage quadrupole mass filter.

Claims

1. An analytical method comprising the steps of: obtaining a first weight of cultured meat; crushing the freeze-dried cultured meat into powder; preparing an analytical sample using the powder; obtaining the amount of components contained in the cultured meat using the results of measuring the analytical sample with an analytical instrument; and calculating the amount of the components per unit wet weight of the cultured meat using the first weight and the amount of the components.

2. The analytical method according to claim 1, further comprising the steps of obtaining a second weight of the freeze-dried cultured meat, and calculating the amount of the component per unit dry weight of the cultured meat using the second weight and the amount of the component.

3. The analytical method according to claim 1, further comprising the steps of obtaining a second weight of the freeze-dried cultured meat and calculating the moisture content of the cultured meat using the first and second weights.

4. An evaluation method comprising the steps of: comparing the moisture content of cultured meat calculated by the analytical method described in claim 3 with a first reference value; and evaluating the quality of the cultured meat using the results of the comparison step.

5. The evaluation method according to claim 4, further comprising the steps of: obtaining a third weight of meat; obtaining a fourth weight of freeze-dried meat; calculating the moisture content of the meat using the third and fourth weights; and determining the first reference value based on the moisture content of the meat.

6. An evaluation method comprising the steps of: comparing the amount of a component per unit wet weight of cultured meat calculated by the analytical method described in claim 1 with a second reference value; and evaluating the quality of the cultured meat using the results of the comparison step.

7. The evaluation method according to claim 6, further comprising the steps of: preparing meat of approximately the same size as the cultured meat; obtaining a fifth weight of the meat; crushing the freeze-dried meat into powder; preparing a reference sample using the meat powder; obtaining the amount of the component contained in the meat using the results of measuring the reference sample with an analyzer; calculating the amount of the component per unit wet weight of the meat using the fifth weight and the amount of the component; and determining the second reference value based on the amount of the component per unit wet weight of the meat.

8. The evaluation method according to claim 7, wherein the amount of the component contained in the cultured meat and the amount of the component contained in the meat are obtained by continuous analysis.

9. The evaluation method according to claim 7, wherein, when measured by liquid chromatography-tandem mass spectrometer (LC-MS / MS) or gas chromatography-tandem mass spectrometer (GC-MS / MS), the step of obtaining the amount of a component contained in the cultured meat includes the step of quantifying the component using a signal derived from the precursor ion of the component as an indicator, and the step of obtaining the amount of a component contained in the meat includes the step of quantifying the component using a signal derived from the product ion of the component as an indicator.

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