Application and method of curcumin in reducing chicken fat rancidity odor

Abstract

This invention belongs to the field of livestock and poultry feed nutrition regulation, and specifically relates to the application and method of curcumin in reducing the rancid odor of chicken fat. This application found significant differences in the volatile flavor compounds and total lipid molecular profiles between the control group and the curcumin group. Curcumin can reduce the formation of MDA and rancid odor compounds in fats, and reduce the formation of oxidative odor compounds such as diphenyl sulfone, trans-2-nonenal, trans-2-octenal, acetaldehyde, and heptanal. Its mechanism for reducing MDA is to regulate the activity of PEMT enzyme in chicken skeletal muscle, promoting the conversion of PE to MePC and dMePE. Correlation analysis showed that 2-methylbutanal, 2-ethyl-1-hexanol, 2-undecane, acetaldehyde, and various lipid molecules were significantly correlated, and the abundance of flavor compounds was regulated by lipid composition. This application provides a theoretical basis for regulating the volatile matter spectrum of chicken flavor and also provides theoretical support for developing high-quality poultry products.

Description

Technical Field

[0001] This invention belongs to the field of livestock and poultry feed nutrition regulation technology, and specifically relates to a method for reducing the rancid odor of chicken fat. Background Technology

[0002] Chicken is one of the world's most consumed sources of animal protein. Its popularity stems from its balanced nutrition, excellent taste, and ease of processing and storage. Globally, the poultry industry provides high-quality animal protein, playing a crucial role in ensuring food security and improving dietary nutrition. The core of sustainable development in the poultry industry lies in continuously improving meat quality, which directly determines consumer acceptance and product market competitiveness. Flavor, as a core indicator of meat's sensory quality, is key to influencing consumers' repurchase intentions. Meat flavor originates from volatile substances generated through multiple biochemical pathways, primarily including lipid oxidation, Maillard reactions, and protein and nucleotide degradation. Fresh chicken itself has a mild flavor, naturally possessing a bloody and metallic odor induced by heme proteins; its rich meaty aroma largely depends on the subsequent heating process for generation and accumulation. Fresh chicken tissue is rich in flavor precursors, forming the basis for cooked flavors, but also providing substrate conditions for undesirable post-slaughter oxidative reactions. Volatile metabolites in chicken muscle and fat tissue are subject to dual regulation by in vivo metabolism and post-slaughter storage and processing biochemical reactions. They are highly susceptible to deterioration and the generation of off-flavors throughout the entire storage, transportation and processing process.

[0003] Compared to other livestock and poultry meats, chicken muscle tissue is rich in myoglobin, free iron ions, and highly unsaturated fatty acids, making it extremely sensitive to oxidation. It readily undergoes lipid-protein co-oxidation chain reactions, significantly accelerating the deterioration process. Lipid oxidation in chicken is a typical free radical chain reaction: unsaturated fatty acids combine with oxygen to generate primary oxidation products such as hydroperoxides. These products are extremely unstable and further decompose to generate secondary oxidation metabolites such as aldehydes, alcohols, and ketones. These secondary products have extremely low odor thresholds and are the core source of rancid, stale, and rotten odors during chicken storage. Once accumulated in large quantities, they significantly degrade the sensory quality of chicken, drastically reducing consumer acceptance and severely restricting the shelf life and market promotion of fresh and processed chicken products. Therefore, improving chicken flavor and inhibiting the formation of oxidative odors are key technical challenges that the chicken processing industry urgently needs to overcome. The invention patent with publication number CN121621446A discloses a broiler feed premix containing curcumin. Curcumin is used as a flavor-enhancing component and combined with various additives to improve the meat quality and flavor of chicken and weaken lipid oxidation through oral feeding. The core focus is on improving the tenderness of chicken, intramuscular fat content, and deposition of umami substances. Curcumin is only used as an auxiliary flavor component in the compound. It does not build a specific inhibition mechanism for the putrid odor of chicken fat. Moreover, the superposition of multiple components is prone to functional antagonism and cannot focus on the core functions of curcumin itself, such as anti-oxidation of fat and removal of odor.

[0004] In summary, there is an urgent need to develop a green preservation technology that precisely targets the oxidation system of chicken fat and can efficiently block the generation of spoilage odors. This technology would significantly improve the flavor and quality of chicken, meet the high-quality development needs of the modern chicken deep processing industry, and provide theoretical support for natural feed additives to improve meat flavor and develop high-quality poultry products. Summary of the Invention

[0005] To address the aforementioned problems, this invention proposes an application and method for curcumin in reducing the rancid odor of chicken fat.

[0006] The technical solution of this invention is implemented as follows:

[0007] This application clarifies the molecular mechanism by which curcumin regulates the flavor of chicken, providing theoretical support for its use as a natural feed additive to improve meat flavor and develop high-quality poultry products.

[0008] Based on this, the present invention provides an application of curcumin in reducing the rancid odor of chicken fat.

[0009] Preferably, the reduction of the rancid odor of chicken fat is achieved by reducing the content of MDA and rancid odor substances.

[0010] Preferably, the rancid odor substances in the above-mentioned oils are diphenyl sulfones and aldehyde rancid substances.

[0011] Preferably, the aforementioned aldehyde rancid substances include any one or more of trans-2-nonenal, trans-2-octenal, acetaldehyde, and heptanal.

[0012] The curcumin group exhibited higher relative intensities of sensory characteristics such as grassy aroma, fruity aroma, waxy aroma, lipid aroma, citrus aroma, fresh aroma, nutty aroma, and floral aroma. The curcumin group also showed significant upregulation of the following substances: ethyl 2-methylbutyrate (fruity aroma), 2-undecone (orange aroma, fresh aroma, grassy aroma), ethyl 3-methylbutyrate (apple aroma), and 2,3-dimethylpyrazine (aroma of toasted bread, roasted corn, and roasted peanuts). The content of diphenyl sulfone was significantly decreased.

[0013] The ROAV of key aldehydes all decreased: trans-2-nonenal (lipid aroma, cucumber flavor), trans-2-octenal (nut, grass, lipid aroma), acetaldehyde (pungent fruit aroma), and heptanal (citrus, oil, rancid aroma); only the content of 2-methylbutanal increased (cocoa, almond aroma).

[0014] Preferably, the reduction of MDA is achieved by blocking the downregulation of phospholipids. Blocking the downregulation of phospholipids involves regulating the activity of PEMT enzyme in chicken skeletal muscle, promoting the conversion of phosphatidylethanolamine (PE) to methylphosphatidylcholine (MePC) and dimethylphosphatidylethanolamine (dMePE). This results in the upregulation of methylphosphatidylcholine (MePC) and dimethylphosphatidylethanolamine (dMePE), and the downregulation of phosphatidylethanolamine (PE), phosphatidylcholine (PC), and dimethylphosphatidic acid (BisMePA).

[0015] The vast majority of phosphatidylcholine and phosphatidylethanolamine molecules were downregulated in the curcumin group (25 upregulated, 59 downregulated), indicating that curcumin primarily reduces muscle phospholipid content. Simultaneously, the curcumin group exhibited a greater variety of volatile compounds, confirming that dietary curcumin reshapes the meat lipid profile. Unsaturated fatty acids in meat are readily oxidized, generating volatile substances such as aldehydes, ketones, alcohols, hydrocarbons, esters, and heterocyclic compounds, directly determining meat flavor. The upregulation of triglycerides and some phospholipids in the curcumin group may be related to curcumin's enhancement of the broiler's antioxidant capacity. Dietary curcumin can significantly reshape the overall lipid profile of chicken breast; methylated phospholipids (MePC, dMePE, PE) are precursors to lipid oxidation flavor, and changes in lipid content are the core factor influencing differences in the volatile flavor of chicken.

[0016] 2-Methylbutyraldehyde was significantly positively correlated with CL (74:7), PE (18:0-18:3), Hex1Cer (d35:3+O), MePC (38:9), and dMePE (16:0-20:5), and significantly negatively correlated with PE (20:0-20:4) and LPE (18:2); 2-Ethyl-1-hexanol was strongly correlated with Cer (d32:0) and MGDG (42:7e); Acetaldehyde was negatively correlated with DG (16:0-20:4) and PE (22:1-20:4); 2-Undecanone was positively correlated with MePC (38:9) and dMePE (16:0-20:5), and negatively correlated with various lipids such as LPE (18:2), LPC (20:4), LPE (22:5), PE (20:0-20:4), LPE (22:4), and CL (79:5). Key flavor compounds such as aldehydes and ketones are significantly correlated with differentially expressed lipids. Meat flavor generation is regulated by multiple complex metabolic pathways.

[0017] Preferably, the chicken meat mentioned above is chicken breast.

[0018] Secondly, this invention also applies for protection of a method for reducing the rancid odor of chicken breast fat, which involves adding curcumin to the basal diet before feeding chickens (or adding it directly to the chicken breast) to reduce the rancid odor of chicken breast fat.

[0019] The above-mentioned basic diet can be any feed that is available in this field; when added directly to fresh meat, it can reduce the production of rancid and odorous substances in chicken breast by inhibiting lipid oxidation.

[0020] Preferably, the curcumin addition ratio is 400-500 mg / kg, and the feeding cycle is 42 days; the rancid odor substances are diphenyl sulfone and aldehyde rancid substances.

[0021] Preferably, the aforementioned aldehyde rancid substances include any one or more of trans-2-nonenal, trans-2-octenal, acetaldehyde, and heptanal.

[0022] The present invention has the following beneficial effects:

[0023] This invention provides an application of curcumin in reducing the rancid odor of chicken fat. Using multi-omics techniques, the malondialdehyde (MDA) and intramuscular fat content of chicken breast in the control and curcumin groups were detected. The differences in flavor precursor lipids and volatile flavor compounds (VOCs) were systematically analyzed. Results showed no significant difference in intramuscular fat content between the two groups, but significant differences in the molecular profiles of volatile flavor compounds and total lipids. Curcumin reduces the rancid odor of chicken fat by decreasing the formation of MDA and rancid odor substances. The overall flavor of the chicken includes multiple sensory characteristics such as grassy, ​​fruity, waxy, fatty, citrus, fresh, nutty, and floral aromas. Volatile compound analysis confirmed that dietary curcumin supplementation optimizes chicken breast flavor by inhibiting the formation of oxidative odor substances such as diphenyl sulfone, trans-2-nonenal, trans-2-octenal, acetaldehyde, and heptanal. The mechanism of MDA reduction is through regulating the activity of PEMT enzyme in chicken skeletal muscle, promoting the conversion of PE to MePC and dMePE. Correlation analysis showed that 2-methylbutanal, 2-ethyl-1-hexanol, 2-undecane, acetaldehyde, and various lipid molecules were significantly correlated, and the abundance of flavor compounds was regulated by lipid composition. This application comprehensively analyzes the changes in volatile flavor and lipid characteristics of chicken breast treated with curcumin, clarifies its inhibitory effect on undesirable flavor compounds such as diphenyl sulfone and heptanal, provides data support for the development of flavor-improved poultry products, and also provides theoretical support for the improvement of meat flavor and the development of high-quality poultry products using natural feed additives. Attached Figure Description

[0024] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0025] Figure 1 Mass spectra of different chicken breast muscle samples analyzed by GC×GC-TOF MS; Figure A shows the volatile flavor compounds (VOCs) identified in the samples using GC×GC-TOF MS, B shows the relative values ​​of different metabolite categories, and C shows the radar charts of predicted flavor attributes of chicken meat in the CON and CUR groups. The radial axis represents the relative intensity of each flavor attribute, which is derived from the volatile compound spectrum and matched with odor descriptors using the FlavorDB database. No human sensory evaluation was performed.

[0026] Figure 2 GC×GC-TOF MS analysis of different chicken breast muscle samples, where Figure A shows the OPLS-DA analysis of chicken breast muscle samples from the CON group and CUR group. OPLS-DA model parameters: (R 2 Y=1.00), (Q 2=0.34), B is the statistics of differentially expressed metabolites in chicken breast muscle, C is the volcano plot of differential abundance VOCs in chicken breast muscle, D is the heatmap of normalized differential VOC expression in chicken breast muscle of CON group and CUR group (P < 0.05) (relative content is represented by color intensity: red indicates higher content, blue indicates lower content; columns represent samples, rows represent substances; the clustering tree on the left shows the hierarchical clustering of substances), E is the interaction diagram between volatile organic compounds (VOCs) and various flavors, showing the degree of contribution of VOCs to flavor (green circles represent sensory features, red circles represent flavor compounds. For green circles: the larger the circle, the more flavor compounds it connects, and the more important the sensory feature. For red circles: the larger the circle, the more sensory features it is associated with, and the more important the flavor compound).

[0027] Figure 3 Lipid composition analysis of chicken breast muscle, where Figure A shows the proportion of identified lipid classes, and Figure B shows the OPLS-DA score plot distinguishing different chicken breeds. OPLS-DA model parameters: (R 2 Y=1.00), (Q 2 =0.34); C is a heatmap of differentially abundant lipid categories: red indicates relatively high content, blue indicates relatively low content, and D is a volcano plot of differentially abundant lipids.

[0028] Figure 4 This is a heatmap of differentially abundant lipids. Figure A shows the hierarchical clustering of the 20 most abundant differentially abundant lipids (highest VIP values) in chicken breast muscle from the CON and CUR groups. Figure B shows the Pearson correlation heatmap between differentially abundant lipids and key volatile flavor compounds. All correlation analyses were subjected to FDR multiple correction before visualization. Black dots mark statistically significant correlation pairs after FDR correction; cells without black dots correspond to insignificant correlations after correction. * and ** represent... P < 0.05 and P < 0.01. Detailed Implementation

[0029] The technical solution of the present invention will be clearly and completely described below with reference to the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative effort are within the scope of protection of the present invention.

[0030] Unless otherwise specified, the experimental methods used in the following experimental examples are conventional methods; the materials and reagents used are commercially available unless otherwise specified.

[0031] Curcumin (CUR) is a natural polyphenol extracted from turmeric. It is inexpensive, safe, and widely available, possessing antioxidant, anti-inflammatory, and anti-tumor functions, while also protecting mitochondrial function and regulating cell signaling pathways. Curcumin can maintain cell membrane integrity and inhibit lipid and protein oxidation. Existing research mostly focuses on the effects of curcumin on poultry growth performance, antioxidant capacity, and conventional meat quality indicators; systematic studies on curcumin's regulation of the intact volatile flavor profile and lipid molecular profile of chicken breast, as well as the interaction mechanisms among lipid metabolism, lipid oxidation, and flavor formation, remain relatively scarce.

[0032] Based on existing research background and gaps, this invention hypothesizes that dietary addition of curcumin can reshape the molecular profile of muscle lipids and inhibit lipid oxidation, thereby improving the sensory flavor of chicken and reducing the content of unpleasant odor substances. This study combines flavor and lipidomics technologies to detect malondialdehyde and intramuscular fat content in chicken breast, analyze the differences between volatile flavor compounds and total lipid molecules, and elucidate the intrinsic relationship between flavor changes and lipid metabolism. This application clarifies the molecular mechanism by which curcumin regulates chicken flavor, providing theoretical support for its use as a natural feed additive to improve meat flavor and develop high-quality poultry products.

[0033] 1. Ethical Approval

[0034] This animal experiment protocol was approved by the Laboratory Animal Ethics Committee of Henan Agricultural University (Approval No.: 11-0099-2023). All broiler chicken experiments were conducted in strict accordance with the National Institutes of Health's "Guidelines for the Care and Use of Laboratory Animals" and with reference to the "Guidelines for Humane Euthanasia of Laboratory Animals" (T / CALAS 31-2017) issued by the Chinese Association for Laboratory Animal Science.

[0035] 2. Test materials

[0036] Curcumin (CAS No.: 458-37-7, purity 98.85%) was purchased from Nanjing Jingzhu Biotechnology Co., Ltd. (Nanjing, China), batch number JZ23061008; the purity of the product was verified by high performance liquid chromatography (HPLC) in the factory test report.

[0037] 3. Experimental animals and treatment

[0038] A 42-day rearing trial was conducted using 120 healthy one-day-old 817 male broilers purchased from Tongxu Breeding Farm in Henan Province. A completely randomized design was used, with the broilers divided into two groups, each with 6 replicate cages and 10 chickens per cage.

[0039] ① Control group (CON): fed with basal diet;

[0040] ② Curcumin group (CUR): 500 mg / kg curcumin was added to the basal diet and fed continuously for 42 days. The curcumin was mixed evenly according to the manufacturer's specifications using a feed mixer.

[0041] The basic diet formula is shown in Table 1. The experimental chickens had free access to feed and water, and all other feeding and management followed the standardized feeding process for commercial broilers.

[0042] Table 1. Composition and nutritional level of the experimental diet (%, air-dried basis)

[0043]

[0044] Note: The premix provides the following per kilogram of feed: Vitamin A 12000 IU; Vitamin D3 3000 IU; Vitamin E 20 IU; Vitamin K3 1.0 mg; Vitamin B1 2.0 mg; Vitamin B2 6.0 mg; Vitamin B6 3.5 mg; Vitamin B12 0.01 mg; D-Biotin 0.15 mg; Folic acid 1.25 mg; Niacin 35 mg; D-Calcium Pantothenate 10 mg; Copper 8.0 mg; Iron 100 mg; Manganese 80 mg; Zinc 60 mg; Iodine 0.45 mg; Selenium 0.35 mg.

[0045] 4. Sample Collection

[0046] On day 42 of rearing, after the broilers were fasted for 12 hours overnight, one healthy broiler of uniform weight was selected from each replicate cage (6 broilers per group). Selection criteria: broilers in the same cage had consistent growth performance, similar weight, good health, and stable physiological state. Before dissection, the broilers were stunned using a water bath electro-stun device, followed by severing the carotid artery and jugular vein to allow for thorough bloodletting, completing humane euthanasia. Breast tissue was then immediately collected.

[0047] Chicken breast samples were placed in polyethylene sealed bags to eliminate interference from volatile substance detection between groups; the samples were then aged at 4°C for 24 hours to simulate the post-slaughter maturation process, preparing for subsequent detection of malondialdehyde, intramuscular fat, and volatile flavor substances. This maturation method is a standard procedure for meat analysis.

[0048] 5. Data statistical analysis

[0049] Results for malondialdehyde and intramuscular fat are expressed as mean ± standard deviation and independent samples t-tests were performed using SPSS 28.0 software.

[0050] The raw chromatographic data of volatile substances were normalized by total peak area to facilitate comparison of differences between groups; ChromaTOF software was used to match the NIST 2020 mass spectrometry library to complete substance annotation; PubChem database and ClassyFire chemical information software were used to classify and quantify the detected volatile substances, and to statistically analyze the distribution and relative content of each substance.

[0051] After the raw lipid data were standardized, they were imported into LipidSearch 4.0 software for preprocessing and peak alignment. Lipid annotation was completed by matching the mass-to-charge ratio of primary precursor ions and secondary fragment ions with the database, and the total peak area was normalized to eliminate batch effects.

[0052] Orthogonal partial least squares discriminant analysis (OPLS-DA) was used to construct a multivariate statistical model to distinguish samples between groups; the significance of differences between groups was assessed using t-tests and one-way ANOVA. All omics data were log2 transformed to conform to a normal distribution, and missing values ​​were filled with half of the lowest detected concentration; false detection rate (FDR) was used to correct for false positives in high-dimensional omics. Differential lipid screening criteria: variable importance projection VIP > 1, FDR < 0.05.

[0053] Example 1: Determination of malondialdehyde and intramuscular fat content

[0054] Chicken breast samples were collected and homogenized with pre-cooled 0.9% physiological saline to prepare a 10% (w / v) tissue homogenate. The homogenate was centrifuged at 3000×g for 10 min at 4℃, and the supernatant was collected. Malondialdehyde (MDA) content was determined using a commercially available thiobarbituric acid (TBA) kit (catalog number A003-1-2) from Nanjing Jiancheng Bioengineering Institute, according to the kit instructions. A standard curve was established using 0–10 nmol / mL standards prepared with 1,1,3,3-tetraethoxypropane (TEP). Method precision validation: intra-assay coefficient of variation 3.2%, inter-assay coefficient of variation 5.1%.

[0055] Intramuscular fat (IMF) content was determined according to the national standard GB 5009.6-2016, "National Food Safety Standard - Determination of Fat in Food": 1.0 g of freeze-dried chicken breast sample was taken and extracted with petroleum ether at 60-90℃ for 6 h using the Soxhlet extraction method; the intramuscular fat content was expressed as the percentage of fat mass to the mass of the freeze-dried sample.

[0056] Malondialdehyde and intramuscular fat content in chicken breast:

[0057] Malondialdehyde (MDA) is a characteristic product of lipid peroxidation and can directly reflect the degree of oxidative damage in meat; the higher the content, the more severe the deterioration of meat quality. Intramuscular fat determines the fatty acid composition and affects the generation of flavor compounds and the rate of aroma release during cooking. Therefore, complete lipid profile analysis is fundamental to elucidating the mechanism of meat flavor formation.

[0058] Table 2. Effects of different dietary CUR supplements on MDA and IMF content in chicken breast muscle.

[0059]

[0060] Table 2 shows that adding curcumin to the diet can significantly reduce the malondialdehyde content in chicken breast. P The concentration of intramuscular fat in the muscle was <0.05%, but there was no significant difference between the two groups. This study confirms that curcumin can inhibit the accumulation of malondialdehyde in meat. Intramuscular fat is rich in easily oxidized lipids, which directly determine the composition of flavor compounds. Therefore, this study will further conduct a full lipidomics analysis to elucidate the flavor regulation mechanism.

[0061] Example 2: Gas chromatography-time-of-flight mass spectrometry for detecting differences in volatile flavor compounds in chicken breast

[0062] 1. Extraction method

[0063] Solid-phase microextraction (SPME) is a mainstream technique for separating volatile flavor compounds in meat products and can be coupled with static headspace analysis. A 50 / 30 μm × 1 cm DVB / CAR / PDMS coated SPME fiber (catalog number 57330-U, Supelco) was selected. This fiber has a stronger adsorption capacity for volatile substances with a wide polarity range and is suitable for the full flavor profile analysis of chicken breast.

[0064] Weigh 2.00 g of meat homogenate into a 20 mL brown screw-top headspace vial (22.5 × 75.5 mm, Thermo Fisher Scientific, USA), and add 10 mL of 6% sodium chloride solution. Using a microsyringe, precisely add 10.0 μL of internal standard solution (deuterated n-hexanol, 1 mg / L, prepared with 50% ethanol). Method validation showed that deuterated n-hexanol had recoveries of 86.4%–92.7% in the meat matrix, with a relative standard deviation of less than 4.5%, demonstrating strong stability and suitability for semi-quantitative analysis of volatile substances in meat products.

[0065] The sealed headspace vial was placed in an 80℃ constant-temperature shaking water bath for equilibration for 10 min. The SPME fiber was aged at 250℃ for 10 min before use to remove residual impurities. After sample equilibration, the fiber was inserted into the headspace vial and adsorbed at 80℃ for 25 min. Extraction parameters were referenced from literature on meat flavor verification using the same fiber type. After adsorption, the fiber was inserted into the gas chromatograph inlet and thermally desorbed at 250℃ for 5 min.

[0066] The relative odor activity (ROAV) was calculated based on the semi-quantitative results of volatile substances to evaluate the contribution of each substance to the overall flavor; the odor thresholds of all substances were determined by a third-party authoritative testing institution. In this experiment, ROAV was only used for inter-group comparison under the same fresh meat matrix and does not represent the absolute odor activity of cooked meat.

[0067] 2. Detection by full two-dimensional gas chromatography-time-of-flight mass spectrometry (GC×GC-TOF MS)

[0068] Thermal desorption conditions for volatile flavor compounds: injection port 250℃, desorption for 5 min; instrument: LECO Pegasus ® A 4D full two-dimensional chromatography-time-of-flight mass spectrometry system, equipped with an Agilent 8890A gas chromatograph, a two-stage cryogenic modulator, and a TOF mass spectrometer detector.

[0069] One-dimensional chromatographic column: DB-Heavy Wax (30 m × 250 μm inner diameter, 0.5 μm film thickness, Agilent Technologies, USA);

[0070] Two-dimensional chromatographic column: Rxi-5Sil MS (2.0 m × 150 μm inner diameter, 0.15 μm film thickness, Restek, USA);

[0071] Carrier gas: High-purity helium (purity > 99.999%), constant flow rate 1.0 mL / min.

[0072] Gas phase heating program: Initially 50℃ and hold for 2 min; heat up to 230℃ at 5℃ / min and hold for 5 min; the 2D furnace temperature is always 5℃ higher than the 1D furnace temperature; the modulator temperature is 15℃ higher than the 2D furnace temperature, and the modulation cycle is 6 s; the injection port is kept constant at 250℃.

[0073] Mass spectrometry parameters: transfer line and ion source temperatures were both 250℃; data acquisition frequency was 200 frames / second; electron ionization mode (EI) was used, with an electron energy of 70 eV, a mass-to-charge ratio scan range of 35~550, and a detector voltage of 1960 V; dynamic exclusion was used to simplify the secondary mass spectrum.

[0074] Analysis of the types of volatile substances and overall sensory characteristics:

[0075] Two-dimensional gas chromatography-time-of-flight mass spectrometry (GC×GC-TOF MS) was used to detect volatile substances in chicken breast meat from the control group and curcumin group. After removing impurity and contamination peaks, a total of 1449 volatile flavor compounds were identified and quantified. Figure 1 A), divided into 8 categories: alcohols, aldehydes, carboxylic acids, esters, heterocyclic compounds, hydrocarbons, ketones, and other compounds ( Figure 1 B. Table 3). The curcumin group had 34 more volatile substances than the control group, and its flavor diversity was higher.

[0076] Table 3 Classification of Differential Volatile Flavor Compounds

[0077]

[0078] Significant differences were found in the relative contents of various volatile substances between the groups (Table 3): the curcumin group had higher relative contents of carboxylic acids (3.06 vs 2.15), esters (11.14 vs 8.83), heterocyclic compounds (3.39 vs 2.86), hydrocarbons (34.48 vs 24.00), and ketones (2.72 vs 2.10) than the control group; while the relative contents of alcohols (19.64 vs 25.63), aldehydes (8.83 vs 10.64), and other substances (16.75 vs 23.79) were lower than the control group. The abundant volatile substances in chicken breast contribute to its rich and complex flavor.

[0079] Predicting overall sensory flavor based on volatile matter spectra using the FlavorDB database ( Figure 1 C). The overall flavor profiles of the two chicken groups were similar, but the curcumin group exhibited a higher relative intensity of sensory characteristics such as grassy, ​​fruity, waxy, fatty, citrus, fresh, nutty, and floral aromas. Differences in volatile matter composition are the fundamental reason for the differences in sensory flavor, and dietary polyphenol additives are a key factor in regulating the volatile matter spectrum. Polyphenols possess strong antioxidant capabilities, inhibiting lipid oxidation in fresh meat and enabling targeted regulation of flavor profiles and sensory characteristics.

[0080] Differential volatile flavor compound analysis:

[0081] The OPLS-DA model can completely distinguish between the control group and the curcumin group samples. Figure 2 A) With VIP>1, correction P <0.05% yielded 62 differentially volatile substances ( ) Figure 2 B). Compared with the control group, the curcumin group showed a significant upregulation of 39 volatile substances and a significant downregulation of 14 substances (Table 4). These differentially expressed substances shaped the unique sensory flavor of the chicken meat in both groups. The differentially expressed volatile substances were divided into 13 categories: 2 alcohols, 1 aldehyde, 5 benzene ring compounds, 16 esters, 7 hydrocarbons, 6 ketones, 3 lipids, 1 1,3-dipolar organic compound, 1 organic acid derivative, 2 oxygen-containing compounds, 1 halogenated organic compound, 7 heterocyclic compounds, and 1 category of other substances.

[0082] Table 4. Changes in differentially volatile flavor compounds in chicken breast muscle (CUR vs CON)

[0083]

[0084]

[0085] Table 5. ROAV of differentially volatile flavor compounds in chicken breast muscle

[0086]

[0087]

[0088]

[0089] The differential volatile matter clustering heatmap clearly distinguishes the two groups of samples. Figure 2 C); Volcano plots show the top 5 flavor compounds that are most significantly upregulated and downregulated (C); Figure 2 D. Table 5). The curcumin group significantly upregulated the following substances: ethyl 2-methylbutyrate (fruity aroma), 2-undecone (orange, fresh, and grassy aroma), ethyl 3-methylbutyrate (apple aroma), and 2,3-dimethylpyrazine (toasted bread, roasted corn, and roasted peanut aroma); the content of diphenyl sulfone significantly decreased (P < 0.05). Dietary curcumin can optimize the flavor profile of chicken, reduce the formation of off-flavor substances such as diphenyl sulfone, and polyphenols inhibit lipid oxidation in meat through their antioxidant activity, thereby reducing oxidation-derived off-flavor substances.

[0090] 3. Analysis of key flavor active substances

[0091] Different volatile compounds directly cause the sensory flavor differences between the two groups of chicken. Relative odor activity (ROAV) is a classic method for evaluating food flavor and is suitable for analyzing changes in volatile compound content in fresh meat; ROAV ≥ 1 is considered a key aroma active compound, making a significant contribution to the overall flavor. This study calculated the volatile compounds with ROAV ≥ 1 in both groups and quantified the flavor contribution of each compound (Table 5); a network diagram of the association between flavor compounds and sensory characteristics was constructed based on the FlavorDB database. Figure 2 E), the top ten flavor characteristics are: sweet, grassy, ​​fruity, apple, pineapple, fatty, waxy, soapy, fresh, and nutty. This indicates that the flavor of chicken is the result of the synergistic effect of multiple volatile substances.

[0092] 2-Pentylfuran is the heterocyclic volatile substance with the highest content in chicken breast, and has the aroma of green beans and fresh vegetables. The ROAV in the control group is 28.94 and that in the curcumin group is 25.33. It is generated by the oxidation of linoleic acid and is the core flavor substance shared by both groups of chicken breast.

[0093] Aldehydes are the main volatile substances in chicken and other poultry meat, primarily derived from lipid peroxidation and thermal degradation of fatty acids. Aldehydes are the core of poultry flavor, but the ROAV of key aldehydes decreased in the curcumin group: trans-2-nonenal (control group 100.00 / curcumin group 96.99, fatty aroma, cucumber flavor), trans-2-octenal (12.88 / 12.43, nutty, grassy, ​​fatty aroma), acetaldehyde (3.02 / 2.55, pungent fruity aroma), and heptanal (50.85 / 32.16, citrus, oily, rancid flavor); only 2-methylbutanal content increased (50.85 / 32.16, cocoa, almond aroma).

[0094] Heptanal is a typical marker of lipid oxidation; 2-methylbutanal is a branched aldehyde with a low odor threshold and strong sensory effects. This study confirmed that dietary curcumin promotes its accumulation and optimizes the flavor of chicken breast. Trans-2-nonenal and trans-2-octenal are both lipid oxidation products and are negatively correlated with the body's antioxidant level; the stronger the animal's antioxidant capacity, the lower the aldehyde content in the meat in the early post-mortem period. In this study, the curcumin group showed a significant decrease in MDA, which corroborates the trend of aldehyde reduction.

[0095] Ketones originate from amino acid breakdown and microbial β-oxidation of fatty acids. In the curcumin group, ROAV increased for 1-octen-3-one (mushroom aroma) and 2-undecone (orange, fresh, and grassy aroma), while ROAV decreased for 2,3-butanedione (creamy sweet aroma). Most differentially expressed ketones were upregulated in the curcumin group, showing a trend completely opposite to that of MDA and aldehydes. MDA is a product of lipid peroxidation and is directly associated with rancid odor; curcumin blocks free radical-mediated primary lipid oxidation, reducing the formation of MDA and aldehyde rancid substances; simultaneously, it regulates lipid secondary metabolism, generating creamy and fruity flavor compounds such as methyl ketones and diketones (2-nonanone, 2,3-octenedanedione), enhancing the overall flavor of chicken.

[0096] Alcohols are formed through lipid oxidation, sugar metabolism, amino acid decarboxylation and dehydrogenation, and aldehyde reduction, and often possess a sweet, fruity, or floral aroma. Unsaturated alcohols have a low odor threshold and contribute more to the aroma of meat; saturated alcohols have a high odor threshold and a weaker sensory effect. In this study, 2-ethyl-1-hexanol (with rose and grassy aromas) is a key flavor compound in chicken breast. Excessive levels of diphenyl sulfone and heptanal can produce rancid odors in the fat; curcumin can significantly reduce these two substances and optimize the flavor.

[0097] Example 3: Non-targeted lipidomics analysis

[0098] Lipidomics analysis was outsourced to Suzhou Panomics Biomedical Technology Co., Ltd., and the process consisted of two parts: lipid extraction and liquid chromatography-mass spectrometry (LC-MS) detection. Blank matrices and mixed quality control (QC) samples were set up synchronously throughout the experiment to ensure data reliability; all samples were processed from the same batch to eliminate batch-to-batch differences and eliminate the need for subsequent calibration. A universal lipid internal standard was used to achieve relative quantification of various lipids; method validation included the determination of limits of detection (LOD) and limits of quantitation (LOQ).

[0099] 1. Lipid extraction

[0100] Weigh 100 mg of chicken breast sample into a centrifuge tube and add two glass beads; add 750 μL of pre-cooled (-20℃) chloroform-methanol mixture (volume ratio 2:1), oscillate at 50 Hz for 60 s using a high-throughput tissue homogenizer, repeat twice; let stand on ice for 40 min, add 190 μL of ultrapure water, vortex for 30 s, let stand at -20℃ for 10 min; centrifuge at 13000 g for 5 min at room temperature, and aspirate 300 μL of the lower organic phase into a new centrifuge tube; add 500 μL of chloroform-methanol (2:1) again, vortex for 30 s, centrifuge at 13000 g for 5 min, aspirate 400 μL of the lower organic phase, and combine with the previous organic phase.

[0101] The combined organic extracts were vacuum dried, and the residue was reconstituted with 200 μL of isopropanol, filtered through a 0.22 μm filter membrane, and analyzed by LC-MS. Blank samples and mixed quality control samples were prepared simultaneously throughout the extraction process: the mixed quality control was prepared by mixing equal volumes of all samples, with one quality control sample inserted for every 10 samples to monitor instrument stability; all samples were processed from the same batch to eliminate batch error. The limit of detection was defined as a signal-to-noise ratio (S / N) ≥ 3, and the limit of quantitation (S / N) ≥ 10.

[0102] 2. Liquid Chromatography-Mass Spectrometry (LC-MS) Detection

[0103] Column: Waters ACQUITY UPLC ® BEH C18 (2.1×100 mm, 1.7 μm), column temperature 50℃; autosampler temperature 8℃; binary mobile phase system: A2 phase: acetonitrile-water (60:40, v / v), containing 0.1% formic acid and 10 mM ammonium formate; B2 phase: isopropanol-acetonitrile (90:10, v / v), containing 0.1% formic acid and 10 mM ammonium formate; flow rate 0.25 mL / min, injection volume 2 μL, total elution time 28 min.

[0104] Gradient elution program: 0~5 min: 70%~57% A2; 5~5.1 min: 57%~50% A2; 5.1~14 min: 50%~30% A2; 14~14.1 min: 30% A2 at constant temperature; 14.1~21 min: 30%~1% A2; 21~24 min: 1% A2 at constant temperature; 24.0~24.1 min: 1% rapidly increased to 70% A2; 24.1~28 min: 70% A2 equilibrate the column.

[0105] Electrospray mass spectrometry (ESI-MSn) parameters: positive mode spray voltage 3.5 kV, negative mode 2.5 kV; sheath gas 30 arb, auxiliary gas 10 arb; capillary temperature 325℃. Full scan mass-to-charge ratio range 150–2000, orbital trap resolution 35000; data-dependent secondary mass spectrometry uses high-energy collisional dissociation (HCD) with a collision energy of 30 eV; dynamic exclusion is enabled to simplify the secondary spectrum.

[0106] Comparison of lipid composition in chicken breast:

[0107] This study used 6 biological replicates per group and employed LC-MS / MS non-targeted lipidomics to analyze the differences in lipids in chicken breast meat between the control group and the curcumin group. A total of 1977 lipid molecules were detected and classified into 51 major categories (Table 6). Figure 3 A). Two dominant lipid classes: triglycerides (TG), phosphatidylcholine (PC), phosphatidylethanolamine (PE), diglycerides (DG), sphingomyelin (SM), methylated phosphatidylcholine (MePC), phosphatidylserine (PS), cardiolipin (CL), ceramide (Cer), and monogalactosyldiglyceride (MGDG).

[0108] Table 6. Lipid differences between the control group and the curcumin group

[0109]

[0110]

[0111] There was no significant difference in total triglyceride deposition between the two groups (curcumin group 34.77%, control group 34.75%). P >0.05). Triglycerides are neutral storage lipids responsible for energy storage and fatty acid transport in the body; phosphatidylcholine and phosphatidylethanolamine are the main phospholipids in muscle cell membranes, maintaining cell morphology, membrane fluidity, and signal transduction. Phosphatidylethanolamine participates in the development of energy-producing organelles, while phosphatidylcholine regulates brain development, lipoprotein secretion, and determines tissue lipid distribution.

[0112] In this study, the vast majority of phosphatidylcholine and phosphatidylethanolamine molecules were downregulated in the curcumin group (25 upregulated and 59 downregulated), indicating that curcumin primarily reduces muscle phospholipid content. Simultaneously, the curcumin group exhibited a greater variety of volatile compounds, confirming that dietary curcumin reshapes the lipid profile of meat. Unsaturated fatty acids in meat are readily oxidized, generating volatile substances such as aldehydes, ketones, alcohols, hydrocarbons, esters, and heterocyclic compounds, which directly determine the flavor of meat products.

[0113] Differential lipid analysis:

[0114] The OPLS-DA model showed that the lipidomic profiles of the two groups of chicken breast meat were completely separated. Figure 3B); The smaller the p-value and the higher the VIP value, the more significant the difference between lipid groups. Screening criteria: VIP > 1. P <0.05, a total of 153 differential lipids were obtained, with the highest proportion of categories being phosphatidylcholine (31 types), triglycerides (21 types), and phosphatidylethanolamine (19 types).

[0115] Compared with the control group, the curcumin group showed an increase in the levels of 64 lipids and a decrease in 89 lipids. The upregulated lipids included 3 acetylcarnitines, 2 dimethylphosphatidic acids, 1 cardiolipin, 3 cholesterol esters, 4 ceramides, 2 diglycerides, 1 hexosylceramide, 1 lysophosphatidic acid, 2 monogalactosyldiglycerides, 3 methylphosphatidylcholine, 12 phosphatidylcholine, 9 phosphatidylethanolamine, 2 phosphatidylglycerols, 1 phosphatidylserine, 3 sphingomyelins, 1 sphingosine, 12 triglycerides, 1 enyl ether phospholipid, and 1... The lipids downregulated include 3 types of dimethylphosphatidylethanolamine, 6 types of cardiolipin, 1 type of ceramide glycolipid, 3 types of cortisol, 3 types of diglycerides, 17 types of lysophosphatidylcholine, 6 types of lysophosphatidylethanolamine, 4 types of dimethyllysophosphatidylethanolamine, 1 type of monogalactosyldiglyceride, 3 types of methylphosphatidylcholine, 19 types of phosphatidylcholine, 10 types of phosphatidylethanolamine, 2 types of phosphatidylserine, 2 types of sphingomyelin, and 9 types of triglycerides. Chicken breast lipids are mainly triglycerides, phosphatidylcholine, and phosphatidylethanolamine.

[0116] Triglycerides hydrolyze to release free fatty acids, which are core precursors to chicken flavor; phospholipid oxidation directly generates characteristic volatile substances. The curcumin group showed an overall increase in triglyceride content, which is beneficial to human health; phosphatidylcholine is the main glycerophospholipid in skeletal muscle cell membranes, regulating lipoprotein secretion and tissue lipid distribution; phosphatidylethanolamine ensures the development of energy-producing organelles. The upregulation of triglycerides and some phospholipids in the curcumin group may be related to curcumin's ability to enhance the antioxidant capacity of broilers.

[0117] Triglycerides and phospholipids are rich in unsaturated fatty acids, making them highly susceptible to oxidation during processing and storage. Phospholipids, with their higher unsaturated fatty acid content, are more easily exposed to aqueous oxidizing agents, resulting in a higher oxidation risk than triglycerides. Phospholipid oxidation targets unsaturated double bonds, generating phospholipid hydroperoxides, which further decompose into long-chain aldehydes and short-chain carbonyl volatiles, damaging flavor and reducing nutritional value. Phospholipids are fundamental components of cell membranes; oxidation disrupts membrane integrity, inducing intracellular oxidation and amplifying oxidative chain reactions. Dietary curcumin inhibits MDA accumulation in chicken breast, possibly by blocking phospholipid downregulation, thereby improving meat flavor.

[0118] Differential lipid clustering heatmaps showed tight intra-group repetitive aggregation and complete inter-group separation, indicating good experimental repeatability. Figure 3C); Volcano plot screening revealed the five lipids with the most significant differences: in the curcumin group, methylphosphatidylcholine (MePC) (38:9) and dimethylphosphatidylethanolamine (dMePE) (16:0-20:5) were significantly upregulated, while phosphatidylethanolamine (PE) (18:1-22:0), phosphatidylcholine (PC) (17:0-22:4), and dimethylphosphatidylcholine (BisMePA) (42:5) were significantly downregulated. Figure 3 D). In summary, dietary curcumin can significantly reshape the total lipid profile of chicken breast; methylated phospholipids (MePC, dMePE, PE) are precursors to lipid oxidation flavor, and changes in lipid content are the core factor influencing the differences in volatile flavor in chicken. Subsequent lipid-flavor correlation analysis will further verify this inference.

[0119] Curcumin treatment significantly increased the content of methylated phospholipids (MePC, dMePE) in chicken breast. These lipids are generated by the stepwise methylation of phosphatidylethanolamine via PEMT enzyme. PEMT is mainly expressed in the liver, but it is also present in skeletal muscle and participates in the regulation of phospholipid homeostasis. Curcumin has strong antioxidant capacity, and it is speculated that it regulates the activity of PEMT enzyme in skeletal muscle, promotes the conversion of phosphatidylethanolamine to MePC and dMePE, alters the content of lipid oxidation precursors, and ultimately regulates the generation of meat flavor.

[0120] 4. Correlation analysis between volatile flavor compounds and differential lipids

[0121] The chicken flavors of the control group and the curcumin group differed significantly; key flavor compounds common to both groups (ROAV > 1) were screened. P <0.05): trans-2-nonenal, heptanal, 2-pentylfuran, 2-methylbutanal, 2,3-butanedione, 2-ethyl-1-hexanol, trans-2-octenal, acetaldehyde, 2-undecenone, 1-octen-3-one. Except for 2-ethyl-1-hexanol, 2-methylbutanal, 1-octen-3-one, and 2-undecenone, the contents of the other flavor compounds were reduced in the curcumin group.

[0122] Correlation analysis was conducted on the top 20 differentially expressed lipids in the VIP ranking. Figure 4 A is a heatmap of the top 20 differential lipid clusters, showing a clear distinction in lipid abundance between groups. Figure 4 B is a heatmap showing the correlation between key flavor compounds and differential lipids after FDR correction, only displaying significantly correlated combinations.

[0123] The results showed that 2-methylbutyraldehyde was significantly positively correlated with CL (74:7), PE (18:0-18:3), Hex1Cer (d35:3+O), MePC (38:9), and dMePE (16:0-20:5), and significantly negatively correlated with PE (20:0-20:4) and LPE (18:2); 2-ethyl-1-hexanol was strongly correlated with Cer (d32:0) and MGDG (42:7e); acetaldehyde was negatively correlated with DG (16:0-20:4) and PE (22:1-20:4); 2-undecanone was positively correlated with MePC (38:9) and dMePE (16:0-20:5), and negatively correlated with various lipids such as LPE (18:2), LPC (20:4), LPE (22:5), PE (20:0-20:4), LPE (22:4), and CL (79:5). Key flavor compounds such as aldehydes and ketones are significantly correlated with differentially expressed lipids. Meat flavor generation is regulated by multiple complex metabolic pathways.

[0124] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. Application of curcumin in reducing the rancid odor of chicken fat.

2. Use according to claim 1, characterized in that: The reduction of the rancid odor of chicken fat is achieved by reducing the content of MDA and rancid odor-causing substances in the fat.

3. Use according to claim 2, characterized in that: The rancid odor substances in the oils are diphenyl sulfone and aldehyde rancid substances.

4. Use according to claim 3, characterized in that: The aldehyde rancid substances include any one or more of trans-2-nonenal, trans-2-octenal, acetaldehyde, and heptanal.

5. Use according to claim 4, characterized in that: The reduction of MDA is achieved by blocking the downregulation of phospholipids.

6. Use according to claim 5, characterized in that: The blocking of phospholipid downregulation regulates the activity of PEMT enzyme in chicken skeletal muscle, promoting the conversion of phosphatidylethanolamine to methylphosphatidylcholine and dimethylphosphatidylethanolamine.

7. Use according to claim 6, characterized in that: The chicken meat in question is chicken breast.

8. A method for reducing the rancid odor of chicken breast fat, characterized in that: Adding curcumin to the basal diet of chickens can reduce the rancid odor of chicken breast fat.

9. The method according to claim 8, characterized in that: The curcumin addition ratio is 400-500 mg / kg, and the feeding cycle is 42 days; the rancid odor substances in the oil are diphenyl sulfone and aldehyde rancid substances.

10. The method according to claim 9, characterized in that: The aldehyde rancid substances include any one or more of trans-2-nonenal, trans-2-octenal, acetaldehyde, and heptanal.

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