1. Introduction
Whole-plant corn silage (WPCS) is typically harvested and chopped from the end of milk ripening to the early wax ripening stage, packed into the cellar layer by layer, suppressed, and sealed. WPCS has become a key component in dairy cow diets worldwide [1] because of its high fiber digestibility [2] and nutritional value [3]. The high adoption of WPCS can improve the economic utilization of crops and effectively solve the problem of balanced feed supply in spring and winter in northern regions [4]. At the same time, it optimizes the feed structure for large-scale livestock farms, reduces costs, and increases efficiency in healthy livestock farming.
A major challenge for dairy and corn farmers today is the need to produce larger quantities of WPCS to meet the nutritional demands of high-yielding cows [5]. However, variations in fermentation processes and preservation conditions can lead to significant differences in silage flavor. The formation of unique odors in silage is believed to result from the dynamic relationship between different microorganisms, which influence the metabolic pathways that lead to the accumulation of small molecules [6]. For example, corn stalk silage with Lactobacillus brucei added has a reduced butyric acid taste, a fresh fruit aroma, and a high-quality green fruit color [7]. Li Hongshen et al. found that sweet sorghum silage, rich in starch, protein, fat, and tannins, enhances the flavor of fragrant wine, producing an ethyl acetate aroma and increasing the ethanol content through anaerobic fermentation by brewing yeast [8]. Meanwhile, different microbial populations and contamination by exogenous pathogenic bacteria can lead to variations in silage quality, including differences in flavor compounds [9]. P.H. Robinson et al. discovered that the outer surface of the bale that comes into contact with air has a higher probability of emitting pungent moldy odors in haylage [10]. Silage fermented predominantly by anaerobic spoilage bacteria, such as Butyricicoccus species, can decompose proteins to produce ammonia or amines, resulting in unpleasant odors [6]. This demonstrates that VOCs are formed through interactions between microbial populations or non-biochemical processes. We found that, among various animal management factors, diet is considered to be the most important and sensitive factor influencing the flavor of milk and dairy products [11]. There are three primary ways in which raw milk flavor compounds are synthesized. Firstly, flavor compounds in the diet can be directly absorbed through the digestive tract, enter the bloodstream, and reach peripheral tissues (such as the mammary glands) [12]. Secondly, nutrients such as carbohydrates, fats, and proteins in the diet can influence the formation of flavor substances in milk through rumen metabolism [13]. Some studies have highlighted the potential for direct transfer of compounds from the diet to raw milk. An important finding was that the flavor intensity of cheeses made from milk of indoor green-fed cows was more similar to that of cheeses made from milk of cows fed conserved forage rather than that of cheeses made from milk of cows that graze on pasture [14]. Often, artisanal hard and semi-hard cheese producers often avoid milk from silage-fed farms to prevent late swelling and off-flavors during ripening, which can occur if butyric acid bacteria spores from silage are present [15]. VOCs may be a key factor in reducing DMI to some extent. A study on the effect of fermentation products and VOCs on dry matter intake found that silage with higher levels of biogenic amines (histamine, tyramine, tryptamine, phenylethylamine) and aldehydes (formaldehyde, acetaldehyde, propanal, butyraldehyde) was less favored by ruminants [16]. Additionally, when formic acid and histamine were added to the feed with formaldehyde, cows completely rejected it. In a study, DMI in a 600 kg cow decreased by 0.48 kg/d as the AcA concentration in the diet increased from 10 to 30 g/kg DM [17]. In conclusion, silage VOCs can have negative, neutral, or positive effects on feed selection and intake by ruminants. The herbage utilization method affected the composition and sensory characteristics of milk and cheese, even when the forage’s botanical composition and plot origin were the same. Therefore, the distinct flavor of silage has sparked growing interest in studying its chemical composition, driving further research into the VOCs that influence silage quality.
In food flavor chemistry, gas chromatography–mass spectrometry (GC-MS) is commonly used to compare the retention index (RI) and odor characteristics of target compounds with those of control compounds (real compounds) [18]. This method provides an effective initial identification of compounds, enabling the identification or extraction of target compounds from complex mixtures [19]. Gas chromatography–ion migration spectrometry (GC-IMS) [20] and electronic nose (E-nose) technology [21] have gained popularity in recent years due to their advantages of simple operation, high sensitivity (detecting compounds at the ppb level), and the ability to perform real-time and rapid analysis. The E-nose has a broad detection range and can detect and analyze various gases in real time. With the continuous advancement of identification technology, the accuracy of classifying different VOC gases has reached 97.5% [22]. Furthermore, these new applications continue to emerge and remain an attractive area of research, with significant utilization in various food studies [23], including tea flavors [24], evaluation of milk [25], investigation of meat [26], and assessment of fermented alcoholic beverages [27]. As small molecules, VOCs are the first to be perceived (by smell or taste) when silage is presented to animals. These compounds are produced, released, or retained from the feed substrate and play a role in stimulating the appetite of ruminants [28]. The type, concentration, and formation of VOCs are crucial for assessing odor characteristics, classification, and identification and guiding the production, processing, and utilization of raw materials [23]. However, current research on whole corn silage mainly focuses on its nutritional value, fermentation quality, and microbial content, with relatively little attention given to analyzing its overall characteristic odor. Therefore, utilizing various flavor-omic analytical instruments to study the aroma of silage samples allows for the extraction and detection of VOCs from different silage substrates, providing a more comprehensive dataset for understanding the odor profile of silage.
This study employed advanced analytical methods to identify VOCs in silage, such as HS-SPME-GC-MS, HS-GC-IMS, and E-nose techniques. The objective was to analyze the differences between abnormal silage with a bitter almond taste and normal silage from the same silo and to trace the specific compounds responsible for this unique smell. While odor changes due to microbial activity or production techniques are inevitable, effective management can minimize these odors and enhance silage quality. The study aims to deepen the understanding of silage odor diversity, address challenges posed by VOCs, and develop management strategies to control them, ultimately improving silage quality and supporting the sustainable development of dairy farming.
2. Materials and Methods
2.1. Materials
Figure 1 illustrates an above-ground silo with approximate dimensions of 12 m in width and 3 m in height. The structure is constructed from masonry with flat, impermeable walls. Following compaction, the silage section assumes an inverted U-shape, with the central region elevated relative to the sides, resulting in a curved profile. The silage is sealed with a transparent film, which is subsequently covered by a black-and-white film, with the white side facing outward. After sealing, tire compaction is applied, with at least one tire per square meter, ensuring uniform distribution and full coverage. In the region shown in the figure, within approximately 2.5 m of the left wall, extending from the cellar entrance to the area near the bottom of the wall, the silage exhibits a strong bitter almond odor (left). With this criterion, two types of silage were identified: abnormal silage with a strong bitter almond odor (YCQZ) and normal silage with a sour aroma but no bitter almond taste (ZCQZ). No significant differences in appearance were observed between the two types of silage. The sampling method was to draw a circle with a radius from the center of the circle to 50 cm from the pit wall. Samples were collected at 5 points, that is, the 4 points where the circumference intersects with the diameter vertically and the center point of the circle, and the sampling depth at each point was greater than 15 cm. The silage samples were mixed, reduced to 1 kg by the quartering method, packed into polyethylene bags for silage, and vacuum-sealed by a vacuum packaging machine. The sampling method was to draw a circle with a radius from the center of the circle to 50 cm from the pit wall. Samples were collected at 5 points, that is, the 4 points where the circumference intersects with the diameter vertically and the center point of the circle, and the sampling depth at each point was greater than 15 cm. The silage samples were mixed, reduced to 1 kg by the quartering method, packed into polyethylene bags for silage, and vacuum-sealed by a vacuum packaging machine. The silage samples of the two odors were respectively collected once a day for 3 consecutive days. The electronic nose analysis was conducted on the same day, while the remaining samples were stored at −80 °C for subsequent index testing.
2.2. Instrument and Equipment
Flavor Spec® gas chromatography–ion mobility spectrometry system (G.A.S. Gesellschaft für analytische Sensorsysteme mbH, Dortmund, Germany); gas chromatography–mass spectrometry (Agilent 7000B triple quadrupole, Agilent Technologies Inc., Santa Clara, CA, USA); portable electronic nose (PEN 3, Air Sense Analytics, Schwerin, Germany).
2.3. E-Nose Measurements
Table 1 shows the characteristics of the 10 metal-oxide sensor array in the Air Sense PEN. Twenty grams of feed sample was placed into a conical bottle, and 180 mL of deionized water was added to soak for 4 h. The sample was filtered through four layers of gauze. Next, 7.0 mL of the silage filtrate was placed into a 20 mL glass bottle, 1.0 g NaCl was added and mixed, and the bottles were sealed with silicon septum caps and incubated in a thermostat-controlled bath at 40 °C for 5 min. The air injection needle and suction needle set were inserted into the top of the vial at the same time, and the volatiles were extracted at a constant rate. The experiment lasted 120 s, and the recovery time for flushing the sensors was 240 s. Three samples were taken from each of the two groups, and three parallel repetitions were performed for each sample. Data were recorded every second. For data pattern construction, the mean G/G0 values of each sensor response were calculated from measurements in the 117–119 s range when the sensors were stable, using Win Muster v.1.6 software.
2.4. HS-GC-IMS Measurement
The testing was performed using the Flavor Spec® GC-IMS. One gram of ground feed was placed into a 20 mL headspace vial and sealed. The two groups of silage, each with three samples, were tested once. An automatic headspace injection method was used, with a sample volume of 200 µL. The ion mobility spectrometry (IMS) column was an MXT-5 (15 m × 0.53 mm × 1 µm) (Restek, Westport, CT, USA). The samples were incubated for 20 min at 65 °C. After incubation, 200 µL of sample was drawn using a syringe heated to 45 °C and injected into the GC injector at 85 °C in splitless mode. Nitrogen gas with a purity of 99.99% served as the carrier gas for the GC, with the flow rate programmed according to the following sequence: 2 mL/min for 2 min, 10 mL/min for 8 min, 100 mL/min for 10 min, and 150 mL/min for 10 min. The drift tube of the IMS system was 9.8 cm in length and operated at a voltage of 500 V/cm and a temperature of 45 °C, with a nitrogen flow rate of 150 mL/min.
2.5. HS-SPME-GC-MS Measurement
An Agilent 8890 gas chromatograph was coupled with an Agilent 7000B triple quadrupole (QqQ) mass spectrometer. Headspace solid-phase microextraction (HS-SPME) was performed in the automatic sampler (Agilent PAL 3, RSI 120), which featured an injection needle (8010-0265, 2.5 mL, 65 mm, PTFE). A 7 mL sample was placed into a 20 mL headspace vial for the detection of volatile substances. The SPME conditions were set as follows: the sample vial was incubated at 55 °C for 20 min. A 120 µm DVB/CAR/PDMS extraction fiber was used for the insert SPME process, enriching flavor compounds at 55 °C for 40 min before desorption at 250 °C for 3 min. The fibers were conditioned for 10 min at 270 °C in the injector port. For separation, Agilent HP-5 columns (both 30 m × 0.25 mm in i.d. and 0.25 mm film thickness; J&W Scientific, Folsom, CA, USA) were adopted. The carrier gas was helium with a purity greater than 99.999%, and the flow rate was set to 1.0 mL/min. The no-split mode was used. The injector temperature was maintained at 250 °C. The GC column temperature program was as follows: hold at 40 °C for 2 min, ramp to 230 °C at 4 °C/min, and hold at 230 °C for an additional 5 min. The mass spectrometer acquired spectra using an ionization energy of 70 eV in full-scan mode, covering a mass range of 30 to 400 m/z. The interface temperature, ion source temperature, and quadrupole temperature were set to 250 °C, 280 °C, and 150 °C, respectively.
2.6. Statistical Analysis Methodology
Data standardization and one-way analysis of variance (ANOVA) were performed using SPSSAU website (
3. Results
3.1. Analysis of the Overall Flavor Profiles by E-Nose
The PCA results of the E-nose response values are presented in Figure 2a, displaying the 95% confidence region. The cumulative variance contribution rates of the first principal component (91.1%) and the second principal component (8.3%) sum to 99.4%, indicating that they capture most of the information in the original data. The distance between the confidence ellipses indicates that the data points of the two types of silage can be distinguished, suggesting that the electronic nose is capable of distinguishing between normal (ZCQZ) and abnormal (YCQZ) silage by reflecting the overall flavor profile of the samples. To further assess the contribution degree of each sensor in the electronic nose, the PLS-DA discriminant results are illustrated in Figure 2b. A VIP value exceeding 1.0 is typically regarded as indicative of significant importance in the PLS-DA discriminant process. The electronic nose sensors with VIP values above 1.0 include W5S, highlighting their crucial roles in differentiating the odors of normal and abnormal silage (p < 0.05). The results of the significance analysis are shown in Table 2. The five sensors—W5S, W1C, W1W, W3C, and W2W—show significant differences in distinguishing between the two types of silage samples (p < 0.01).
3.2. Analysis of HS-GC-IMS Test Results
3.2.1. Comparison of Differences in VOCs of Silage Based on HS-GC-IMS
Using the Reporter plugin, a comparison of the spectral differences between the two groups of silage feed samples is shown in Figure 3. The 3D spectrum (Figure 3a) illustrates the ion migration time (normalized), gas chromatography retention time (s), and ion peak intensity along the X-, Y-, and Z-axes, respectively. However, due to the high degree of similarity between the VOCs in the two groups, visually comparing the differences using the 3D spectrum is challenging. Therefore, a two-dimensional plane-view spectrum was obtained through dimensionality reduction. The two-dimensional plane-view spectrum (Figure 3b) normalizes the ion migration time and reaction ion peak (RIP) position. The entire spectrum background is colored blue, while the red vertical line at 1.0 on the horizontal axis represents the RIP peak. The horizontal axis represents the ion migration time, and the vertical axis represents the gas chromatography retention time (in seconds). Each point on the spectrum corresponds to a VOC, with its intensity representing the VOC content. The color of each point indicates the intensity, where white represents low intensity, red signifies high intensity, and deeper colors indicate higher ion peak intensities. Figure 3c displays a GC-IMS two-dimensional plane-view difference spectrum created by selecting the normal silage spectrum as the reference and subtracting the signal peaks of the abnormal silage spectrum. This approach provides a clearer comparison of the differences between the two VOC profiles. If the VOCs in both groups are consistent, the background after subtraction will appear white. Conversely, if the VOC concentration in the abnormal group is higher than that of the reference group, the resulting background will be red, and vice versa. The GC-IMS spectrum results indicate that a significant number of VOCs are concentrated within the retention time range of 1000 s in the feed. The spectrum clearly illustrates the differences in the properties and signal strength of the VOCs between the two groups. Most VOC ions in abnormal silage exhibit longer migration times and significantly higher concentrations than those in normal silage. Figure 3c shows the differences in both the quantity and composition of flavor compounds in the abnormal group compared to the normal group.
3.2.2. Comparison of VOCs Fingerprints in Silage Based on HS-GC-IMS
The VOC fingerprint spectra for the silage samples were generated using the Gallery Plot plugin, as shown in Figure 4. The development of GC-IMS fingerprint chromatograms provides a clearer understanding of the differences in VOCs among the samples. Table 3 presents detailed information on the identified compounds. In the complete fingerprint spectrum, each row represents the concentration value detected in a single sample, while each column corresponds to the same VOC across different samples. The content of each VOC is visualized, with the brightness of each square indicating the concentration level. Brighter squares represent higher concentrations of VOCs, while darker squares indicate lower concentrations. By comparing the differences between the fingerprints, a total of 32 VOCs were qualitatively identified, including 13 esters, 7 alcohols, 5 aldehydes, and 1 heterocyclic compound. The HS-GC-IMS results indicate that both samples contain distinct characteristic VOCs, with the abnormal sample exhibiting significantly more specific compounds than the normal one. This difference may account for the variation in their odors and the presence of a bitter almond taste. In the area labeled with a red frame, substances such as 3-methyl-1-pentanol, ethyl isovalerate, 1-pentane- 3-ol, benzaldehyde, 1-hexanol, hexyl acetate, 2-pentyl furan, and ethyl acetate were found in higher concentrations in the normal group samples. By contrast, the abnormal sample features elevated concentrations of compounds such as 2-pentyl-3-hexenal, 3-hexenoic acid ethyl ester, methyl salicylate, 2-propenyl-2-pentenoate, octanoic acid ethyl ester, 3-pentanone, 1-butanol, 1-hexanol, ethyl propionate, and 1-propanol, as indicated in the yellow frame.
According to the PCA analysis chart of VOCs detected by GC-IMS, the cumulative variance contribution of PCA1 (98.2%) and PCA2 (1%) was 99.2%, effectively distinguishing the samples between different groups. As shown in Figure 5, the GC-IMS results align with the E-nose analysis results, and the considerable distance between the two sample groups indicates that the VOCs in the two types of silage are significantly different.
3.3. Analysis of HS-SPME-GC-MS Results
HS-SPME-GC-MS analysis was performed on the two groups of silage (Supplemental Table S1;
Esters and acids are the most abundant compounds identified. Esters were detected in both samples. The identified esters in this sample include 2-methylbutyl hexanoate, benzoyl isothiocyanate, isobutyl acetate, methylazoxymethanol acetate, propane-2-isocyanate-, 1,2-ethanediol phenyl cyclic sulfite, hydrogen isocyanate, tetramethylene diisocyanate, butanoic acid, 2-hydroxy methyl ester, methoxy acetic acid, 3-methyl butyl ester, benzyl (1,2,3-thiadiazol-4-y) carbamate, and propane-1-isocyanate, totaling 13 esters identified. By contrast, abnormal silage feed contains 10 compounds, including 3-ethyl-3-pentyl methylphosphonofluoridate, hydrogen isocyanate, methyl nitrite, benzoyl isothiocyanate, boronic acid dimethyl ester, ethane, isothiocyanate, phosphonoacetic acid, and tetramethyl butyric acid neopentyl ester. Notably, 3-ethyl-3-pentyl methylphosphonofluoridate is the most abundant VOC in abnormal feed and is exclusively detected in abnormal fermentation. These high concentrations of esters highlight their unique influence on the flavor characteristics of the bitter almond-flavored silage. In general, most silage has a vinegar-like taste, which is attributed to their higher acetic acid content. Two acids, acetic acid-d3 (2,2,2-D3) and octanoic acid, silver(1+) salt, were detected in abnormal corn silage. Particularly, benzaldehyde was identified in abnormal corn silage, further defining the unique olfactory signature of bitter almonds. Various compounds are also present in abnormal silage, with compounds like formamidoxime, pyridinium, and 1-amino- and chloride 1-amino chloropyridine adding to the pungent odor complexity.
4. Discussion
4.1. Analysis of Volatile Profiles by E-Nose
The E-nose data show that each gas sensor in the electronic nose array has different sensitivities to the measured gases. The differential contribution values indicate that W5S, W1S, and W2S are the key substances distinguishing the two feedstuffs. This suggests that nitrogen oxides, alcohols, aldehydes, ketones, and methane are key compounds that differentiate the two silage groups, with nitrogen oxides likely serving as the primary odor source responsible for distinguishing the differences in feed quality between normal and abnormal silage. Overall, the results demonstrate the effectiveness of the E-nose in distinguishing odors based on compound type and concentration, providing accurate qualitative results with acceptable cost and efficiency.
4.2. Characteristic VOCs of Silage Based on HS-GC–IMS
Using GC-IMS technology to obtain three-dimensional, two-dimensional, and color difference spectra revealed differences in the VOCs between normal and abnormal silage under different flavor conditions. Based on the fingerprint spectrum results, the silage samples exhibit distinct characteristic VOCs.
Compounds such as 3-hexenyl acetate and salicylic acid methyl ester in abnormal silage demonstrate stronger signal intensities in the GC-IMS spectrum. In many plants, methyl salicylate (MeSal) is a naturally emitted compound during stress events, making it a key marker for early plant disease detection [29]. T. K. Jayasekara et al. identified salicylic acid methyl ester as the primary volatile component in Securidaca long pedunculate (Polygalaceae) through GC/MS analysis [30]. MeSal functions as a volatile signaling molecule and is recognized for its aromatic qualities, reminiscent of the scent of wintergreen [31]. This indicates that these compounds are produced by several different pathways compared to the metabolites typically found in normal silage fermentation. Additionally, common VOCs such as ethanol, ethyl ethanoate, ethyl formate, isopentyl isovalerate, ethyl 2-methyl-2-pentenoate, and 2-methyl-2-pentenoate were also identified. Some VOCs appear as monomers and dimers, including ethyl heptanoate, ethyl hexanoate, propanol, ethyl pentanoate, butyl iso pentanoate, and 3-hydroxy hexanoate, due to varying concentrations and characteristics. Regarding the metabolic pathways leading to the biosynthesis of volatile compounds, aldehydes can convert into their corresponding alcohols, such as 1-butanol, 1-hexanol, and 1-propanol. Acids (hexanoic acid, 3-hexenyl acid) will produce corresponding ethers (3-hexenyl acetate, ethyl acetate) during fermentation, and they are also known to play a significant role in dehydration during the wine-making process to produce wines with elevated aromatic profiles [32]. The presence of substantial amounts of organic acids in silage can rapidly lower the pH of the environment, promoting the production of various esters in conjunction with aldehydes [33]. For example, ethanoic acid, which is the second most abundant acid in silage, can react with 1-hexanol to generate ethyl hexanoate and ethyl acetate when combined with ethanol. The literature indicates that esters exhibit high odor intensity ratios (OIRs) and are among the most active compounds in the fermentation process [34]. Commonly reported esters include butyric acid ethyl ester, hexanoic acid ethyl ester, and octanoic acid ethyl ester [35], which are recognized as potent odorants that impart sweet, floral, and fruity fragrances.
4.3. Identification and Analysis of VOCs in Silage by HS-SPME-GC-MS
4.3.1. Esters
The aroma compounds in different flavor types of silage vary somewhat, both in type and content. Abnormal silage exhibits considerably greater diversity and a higher concentration of volatile compounds than the normal silage.
Esters were detected in both samples. Notably, 3-ethyl-3-pentyl methylphosphonofluoridate is the most abundant VOC in abnormal feed and is exclusively detected in abnormal fermentation. It is a phosphate derivative where an alkyl group replaces the hydrogen atom in the phosphate molecule. Organophosphate esters (OPEs) are widely reported to exist in the environment [36] as well as in various plant and animal bodies [37]. They play an important role in controlling harmful bacterial or insect populations on corn plants by functioning as organophosphate pesticides. Typically, OPEs degrade rapidly through the action of biological enzymes or microorganisms [38]. Diisopropyl fluorophosphate (DFP) can be degraded and metabolized by enzymes isolated from environmental microbes [39]. Isothiocyanates (ITCs) and related derivatives, such as benzyl isothiocyanate (BITC) and isocyanates, are key flavor contributors to both types of silage feeds. These compounds are secondary metabolites found primarily in cruciferous plants and are typically the main substances responsible for their characteristic odor [40]. When plant tissues are damaged by cutting or chewing, benzoyl isothiocyanate hydrolyzes to produce degradation products such as isothiocyanates, indoles, and thiocyanates. These compounds decompose slowly in the air, imparting a unique pungency and bitterness, which creates a distinctive sensory experience for animals consuming these plants [41]. As a naturally occurring organic isothiocyanate, benzoyl isothiocyanate is also believed to be a potential new antibacterial agent [42]. Research [43] has shown that certain isothiocyanate derivatives are plant chemicals found in the seeds of Sinapis alba L. (white mustard) that inhibit harmful gut bacteria like Clostridium and Escherichia coli while also exhibiting antioxidant, anti-cancer, and anti-inflammatory properties [44]. Isothiocyanates are known to produce unpleasant odors. The characteristic pungent odor of the abnormal silage stored on the side of the silo may result from the frequent use of bleach or disinfectants for cleaning. Previous studies have reported that environments frequently cleaned with bleach exhibit higher levels of HNCO, organic isocyanates, HCN, and amides [45]. Within one day of cleaning with bleach, HNCO and other nitrogen-containing substances, including isocyanates, cyanogen chloride, and carbon chloride, are formed [46]. In industrial water, swimming pool disinfection, and other applications, use of chloroisothiocyanuric acid esters as disinfectants produces byproducts like cyanuric acid, which has a slightly bitter taste and decomposes into a strongly acetone-like cyanide odor, resembling almond flavor upon heating [47]. This compound is also utilized as a disinfectant in agriculture and silkworm rearing. Isothiocyanates play a crucial role as intermediate organic compounds in pesticide production and readily react with alcohol to form aminocarbonyl compounds, which are essential for producing various insecticides, fungicides, and herbicides. Consequently, residues of isothiocyanate derivatives, such as phenyl isothiocyanate, may accumulate in silage corn due to extensive use of aminocarbonyl insecticides during corn’s growth stage. Additionally, isocyanates can be found in furniture. For example, the -NCO functional group has been detected in bedding covers and foam [48]. The most widely used and highest-yielding organic isocyanate is toluene diisocyanate (TDI), known for its strong and pungent odor, typically produced through spray-coating polyurethane. It remains unclear whether these sources directly release isocyanates or if they chemically or microbiologically degrade to form HNCO. Isocyanate ethyl propionate was identified in abnormal corn silage, formed by esterifying isocyanic acid with ethyl acetate, which also exhibits a pungent odor. This ester may contribute to the bitter almond smell in abnormal corn silage.
4.3.2. Acids
Two acids, acetic acid-d3 (2,2,2-D3) and octanoic acid, silver(1+) salt, were detected in abnormal corn silage. Organic acids are commonly used to assess the fermentation quality and aerobic stability of silage, as they represent typical fermentation products produced by microorganisms during the silage process. Among these, octanoic acid has a foul odor characterized as acidic and sweaty, which can negatively impact silage. While most silage exhibits a sour balsamic taste due to higher concentrations of acetic acid, propionic acid, and butyric acid—commonly produced by heterofermentative lactobacilli, Enterobacter, or Clostridium—abnormal silage tends to have a more pronounced sour and sweaty odor. This stronger odor may be linked to the presence of acetic acid and caprylic acid.
4.3.3. Aldehydes
Aldehydes are formed through the further oxidation of lipids in plants by the action of lipoxygenase and often contribute to flavor due to their low threshold [49]. Benzaldehyde, with its distinctive bitter almond odor, has been reported as a major VOC in various types of meat, including raw beef [50]. Its presence is thought to be linked to the grass-fed plants consumed by cattle [51] and is commonly found in the plant kingdom, mainly as glycosides in the stems, leaves, or seeds of plants [52]. Plant parts such as almond skins, cherries, bay leaves, and peach stones are also used as livestock feed [53], contributing to the characteristic almond aroma. This suggests that benzaldehyde may be one of the primary VOCs responsible for the bitter almond flavor. In addition, benzaldehyde has been identified as an intermediate of the herbicide wild Yanquat and the plant growth regulator antimyramine [54]. The synthesis of the organophosphorus insecticide 2-chloroethyl isocyanate via an amidation reaction, followed by a reaction with cypermethrin, has been linked to the isocyanate compounds detected in abnormal silage. Up to now, insufficient attention has been given to the impact of the development of the bitter almond flavor in silage. Therefore, further studies and comparisons are needed better to understand its contribution to the flavor of abnormal silage.
4.3.4. Others
Particularly, a variety of pungent odor compounds was significantly detected in abnormal silage, including formamidoxime, pyridinium, 1-amino- and chloride 1-amino chloropyridine. Formamide contains two active functional groups—carbonyl and amide groups—making it susceptible to reactions that yield various nitrogen-containing heterocyclic compounds. Under the influence of a strong dehydrating agent, hydrogen cyanide can also be formed. Propylene glycol monomethyl ether is characterized by low volatility and a distinct odor. It is commonly used in fuel antifreeze and cleaning agents. N-methyl morpholine N-oxide (NMO), another organic compound with a characteristic odor, is used in the production of its upstream and downstream products, N-methyl morpholine and oxidized 4-methyl morpholine monohydrate. These compounds serve as important organic chemical intermediates and are widely used in pesticides, herbicides, metal corrosion inhibitors, fiber treatments, and solvents. 1-Aminochloropyridine, a member of the pyridine class, is used in agriculture as a herbicide and synergist. Amiclopyridinium chloride, known for its chlorine-like odor, is a post-emergence translocation herbicide used to treat the stems and leaves of various crops worldwide. These crops include cereals, oilseed rape, maize, sugarcane, forest land, pastures, grasslands, and non-cultivated land, where it helps to control broadleaf weeds (both annual and perennial) and shrubs. These VOCs contribute to unique olfactory signatures, enhancing the complexity of the bitter almond odor.
5. Conclusions
In this study, the volatile flavor profiles of two different types of WPCS stored in the same silo were analyzed using HS-SPME-GC-MS, HS-GC-IMS, and E-nose. A total of 32 and 101 VOCs were identified by HS-GC-IMS and HS-SPME-GC-MS, respectively, including aldehydes, alcohols, esters, ketones, and hydrocarbons. The analysis of abnormal silage via HS-SPME-GC-MS revealed three major classes of characteristic VOCs associated with a bitter almond taste, including benzaldehyde, cyanides, and isocyanates. The action of organophosphate ester-related pesticides was found to increase specific volatile compounds, such as cyanides and isocyanates, through biological enzymatic or microbial biochemical processes. This suggests that these compounds are unique volatile byproducts and precursors of the bitter almond flavor in abnormal silage. The presence of benzaldehyde may be linked to herbicides or plant growth regulators applied to corn, highlighting the unique bitter almond flavor characteristics imparted by the silage fermentation process. E-nose differential contribution values indicated that W5S (nitrogen oxides) was the key substance distinguishing the two feedstuffs. These findings are consistent with GC-IMS and GC-MS results, which identified 34 nitrogen-containing heterocyclic compounds in the abnormal silage sample, including 1H-tetrazole-1,5-diamine, which is noted for its high nitrogen content. In summary, the VOC data revealed distinct combinations and concentrations of compounds in the feedstuffs, each with different odor profiles, suggesting that these compounds can interact with one another to regulate further flavor characteristics, which are critical for feed quality. The comprehensive use of HS-SPME-GC-MS and HS-GC-IMS techniques provided a detailed understanding of the impact of VOCs on silage odor. The odor of silage is likely influenced by environmental factors, such as temperature and microorganisms, and thus warrants further investigation in the future. This study enhances our understanding of the chemical processes involved in silage odor formation and provides a scientific basis for addressing bitter almond odors and improving silage quality.
Conceptualization, Y.C. and L.W.; methodology, Y.C. and Y.Z. (Yawei Zhang); software, Y.C. and Y.Z. (Yangdong Zhang); validation, Y.C. and N.Z.; formal analysis, Y.C. and Y.Z. (Yangdong Zhang); investigation, Y.C. and Y.Z. (Yangdong Zhang); resources, Y.Z. (Yuanqing Zhang) and Y.Z. (Yangdong Zhang); data curation, Y.C.; writing—original draft preparation, Y.C.; writing—review and editing, Y.G and L.W.; visualization, Y.C. and Y.Z. (Yawei Zhang); supervision, Y.Z. (Yangdong Zhang) and Y.Z. (Yawei Zhang); project administration, N.Z. and Y.Z. (Yuanqing Zhang); funding acquisition, Y.Z. (Yuanqing Zhang) and Y.Z. (Yangdong Zhang). All authors have read and agreed to the published version of the manuscript.
Not applicable.
All data supporting the results of this study are included in this manuscript and its
The authors declare no conflicts of interest.
Footnotes
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Enlarge this image.Figure 2. Statistical analysis of volatile profiles of two samples based on E-nose. (a) The scores plot of PCA based on E-nose. (b) Important VOCs (VIP greater than 1.0) identified by PLS-DA based on E-nose. (c) The violin plot based on E-nose.
Enlarge this image.Figure 3. Three-dimensional (a) and 2D top view (b) HS-GC-IMS map of VOCs of silage. (c) HS-GC-IMS 2D plane-view difference spectrum.
Enlarge this image.Figure 4. VOC fingerprint comparisons of two varieties of silage samples detected by HS-GC-IMS. Each row represents all the signal peaks selected from a silage sample. The substances within the red box in Figure 4 have higher content in ZCQZ, while the substances within the yellow box have higher content in YCQZ.
Enlarge this image.Figure 6. The total ion chromatograms identified by HS-SPME-GC-MS for (a) normal and (b) abnormal silage groups. Abscissa: drift times. Ordinate: reactant ion peak.
Characteristics of the 10 metal-oxide semiconductor sensors of E-nose.
| Array No. | Sensor | Specificity |
|---|---|---|
| S1 | W1C | Aromatic |
| S2 | W5S | Broad range, nitrogen oxides |
| S3 | W3C | Aromatic compounds |
| S4 | W6S | Hydrogen |
| S5 | W5C | Arom-aliph, alkenes |
| S6 | W1S | Broad range, methane |
| S7 | W1W | Sulfur-organic |
| S8 | W2S | Broad range, alcohols |
| S9 | W2W | Sulf-chlor |
| S10 | W3S | Methane-aliph |
Analysis of variance (ANOVA) was conducted on the 10 sensors of E-nose for two sets of samples.
| Sensor | Group (Mean ± SD) | p-Value | |
|---|---|---|---|
| YCQZ (n = 9) | ZCQZ (n = 9) | ||
| W1C | 3.36 ± 0.54 | 2.17 ± 0.71 | 0.001 ** |
| W5S | 43.71 ± 23.40 | 7.54 ± 7.19 | 0.000 ** |
| W3C | 3.51 ± 0.59 | 2.37 ± 0.88 | 0.005 ** |
| W6S | 1.75 ± 0.24 | 1.35 ± 0.45 | 0.032 * |
| W5C | 3.51 ± 0.62 | 2.49 ± 1.02 | 0.020 * |
| W1S | 5.14 ± 1.38 | 4.61 ± 3.30 | 0.665 |
| W1W | 2.96 ± 0.80 | 1.57 ± 0.65 | 0.001 ** |
| W2S | 4.37 ± 1.12 | 4.26 ± 3.25 | 0.926 |
| W2W | 3.83 ± 0.76 | 2.36 ± 1.17 | 0.006 ** |
| W3S | 1.10 ± 0.09 | 1.10 ± 0.10 | 0.970 |
* p < 0.05 ** p < 0.01.
Information of VOCs in silage detected by HS-GC-IMS.
| Count | Compounds | CAS# | Formula | RI | Rt/s | Dt/ms |
|---|---|---|---|---|---|---|
| 1 | Ethyl heptanoate-M | C106309 | C9H18O2 | 1099.9 | 763.593 | 1.39905 |
| 2 | Ethyl heptanoate-D | C106309 | C9H18O2 | 1097.4 | 757.933 | 1.9266 |
| 3 | Ethyl hexanoate-M | C123660 | C8H16O2 | 1010.1 | 585.026 | 1.33876 |
| 4 | Ethyl hexanoate-D | C123660 | C8H16O2 | 1010.9 | 586.434 | 1.79998 |
| 5 | Benzaldehyde | C100527 | C7H6O | 975.8 | 519.405 | 1.46871 |
| 6 | Isoamyl acetate | C123922 | C7H14O2 | 873.2 | 354.345 | 1.74719 |
| 7 | 2-Methyl-2-pentenal | C623369 | C6H10O | 833.6 | 308.942 | 1.50419 |
| 8 | 1-Penten-3-ol | C616251 | C5H10O | 697.8 | 184.419 | 0.94559 |
| 9 | propionic acid propyl ester | C106365 | C6H12O2 | 801.8 | 276.671 | 1.59744 |
| 10 | 1-hexanal | C66251 | C6H12O | 791.1 | 266.58 | 1.55781 |
| 11 | 3-Methyl-2-butenal | C107868 | C5H8O | 745.6 | 223.109 | 1.35592 |
| 12 | 2-Methylpentanal | C123159 | C6H12O | 756.4 | 232.975 | 1.53422 |
| 13 | Acetic acid propyl ester | C109604 | C5H10O2 | 713.4 | 196.287 | 1.48611 |
| 14 | Ethyl acetate | C141786 | C4H8O2 | 616.8 | 146.897 | 1.33928 |
| 15 | 3-Pentanone | C96220 | C5H10O | 692.7 | 180.722 | 1.36617 |
| 16 | 1-Butanol | C71363 | C4H10O | 666.3 | 167.553 | 1.37839 |
| 17 | 1-Propanol-M | C71238 | C3H8O | 569.0 | 129.339 | 1.27328 |
| 18 | 1-Propanol-D | C71238 | C3H8O | 571.9 | 130.372 | 1.43829 |
| 19 | Acetic acid methyl ester | C79209 | C3H6O2 | 540.9 | 120.043 | 1.19872 |
| 20 | Ethanol | C64175 | C2H6O | 501.7 | 108.166 | 1.14127 |
| 21 | 2-Pentyl furan | C3777693 | C9H14O | 995.9 | 560.553 | 1.25404 |
| 22 | Ethyl pentanoate-M | C539822 | C7H14O2 | 900.1 | 389.987 | 1.26823 |
Rt: Represents the retention time in the capillary GC column. RI: Represents the retention index of adjacent n-alkanes before and after the peak corresponding to the target metabolite. Dt: Represents the drift time in the drift tube. M, D: Represent the monomer, dimer of the substance, respectively.
Supplementary Materials
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References
1. Ferrero, F.; Tabacco, E.; Piano, S.; Casale, M.; Borreani, G. Temperature during conservation in laboratory silos affects fermentation profile and aerobic stability of corn silage treated with Lactobacillus buchneri, Lactobacillus hilgardii, and their combination. J. Dairy Sci.; 2021; 104, pp. 1696-1713. [DOI: https://dx.doi.org/10.3168/jds.2020-18733] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/33309357]
2. Liu, Y.; Li, Y.; Lu, Q.; Sun, L.; Du, S.; Liu, T.; Hou, M.; Ge, G.; Wang, Z.; Jia, Y. Effects of Lactic Acid Bacteria Additives on the Quality, Volatile Chemicals and Microbial Community of Leymus chinensis Silage During Aerobic Exposure. Front. Microbiol.; 2022; 13, 938153. [DOI: https://dx.doi.org/10.3389/fmicb.2022.938153] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/36118219]
3. Ferraretto, L.F.; Shaver, R.D.; Luck, B.D. Silage review: Recent advances and future technologies for whole-plant and fractionated corn silage harvesting. J. Dairy Sci.; 2018; 101, pp. 3937-3951. [DOI: https://dx.doi.org/10.3168/jds.2017-13728]
4. Kmiecik, L.L.; Zimmermann, G.G.; Jasper, S.P.; Savi, D.; Strapasson Neto, L.; Sobenko, L.R. Whole-plant corn silage harvesting modalities: Energy efficiency and operational performance. An. Acad. Bras. Ciências; 2023; 95, e20220312. [DOI: https://dx.doi.org/10.1590/0001-3765202320220312] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/37909543]
5. National Academies of Sciences, Engineering, and Medicine. Nutrient Requirements of Dairy Cattle: Eighth Revised Edition; The National Academies Press: Washington, DC, USA, 2021; 502.
6. Zhang, K.; Zhang, T.-T.; Guo, R.-R.; Ye, Q.; Zhao, H.-L.; Huang, X.-H. The regulation of key flavor of traditional fermented food by microbial metabolism: A review. Food Chem. X; 2023; 19, 100871. [DOI: https://dx.doi.org/10.1016/j.fochx.2023.100871]
7. da Silva, N.C.; Nascimento, C.F.; Nascimento, F.A.; de Resende, F.D.; Daniel, J.L.P.; Siqueira, G.R. Fermentation and aerobic stability of rehydrated corn grain silage treated with different doses of Lactobacillus buchneri or a combination of Lactobacillus plantarum and Pediococcus acidilactici. J. Dairy Sci.; 2018; 101, pp. 4158-4167. [DOI: https://dx.doi.org/10.3168/jds.2017-13797] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/29454688]
8. Li, H.; Han, X.; Liu, H.; Hao, J.; Jiang, W.; Li, S. Silage Fermentation on Sweet Sorghum Whole Plant for Fen-Flavor Baijiu. Foods; 2021; 10, 1477. [DOI: https://dx.doi.org/10.3390/foods10071477] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/34202182]
9. Irawan, A.; Sofyan, A.; Ridwan, R.; Hassim, H.A.; Respati, A.N.; Wardani, W.W.; Sadarman,; Astuti, W.D.; Jayanegara, A. Effects of different lactic acid bacteria groups and fibrolytic enzymes as additives on silage quality: A meta-analysis. Bioresour. Technol. Rep.; 2021; 14, 100654. [DOI: https://dx.doi.org/10.1016/j.biteb.2021.100654]
10. Robinson, P.H.; Swanepoel, N. Impacts of a polyethylene silage pile underlay plastic with or without enhanced oxygen barrier (EOB) characteristics on preservation of whole crop maize silage, as well as a short investigation of peripheral deterioration on exposed silage faces. Anim. Feed Sci. Technol.; 2016; 215, pp. 13-24. [DOI: https://dx.doi.org/10.1016/j.anifeedsci.2016.02.001]
11. Manzocchi, E.; Martin, B.; Bord, C.; Verdier-Metz, I.; Bouchon, M.; De Marchi, M.; Constant, I.; Giller, K.; Kreuzer, M.; Berard, J. et al. Feeding cows with hay, silage, or fresh herbage on pasture or indoors affects sensory properties and chemical composition of milk and cheese. J. Dairy Sci.; 2021; 104, pp. 5285-5302. [DOI: https://dx.doi.org/10.3168/jds.2020-19738]
12. Santamarina-García, G.; Amores, G.; Hernández, I.; Morán, L.; Barrón, L.J.R.; Virto, M. Relationship between the dynamics of volatile aroma compounds and microbial succession during the ripening of raw ewe milk-derived Idiazabal cheese. Curr. Res. Food Sci.; 2023; 6, 100425. [DOI: https://dx.doi.org/10.1016/j.crfs.2022.100425]
13. Vargas-Bello-Pérez, E.; Khushvakov, J.; Ye, Y.; Pedersen, N.C.; Hansen, H.H.; Ahrné, L.; Khakimov, B. Goat Milk Foodomics. Dietary Supplementation of Sunflower Oil and Rapeseed Oil Modify Milk Amino Acid and Organic Acid Profiles in Dairy Goats. Front. Vet. Sci.; 2022; 9, 837229. [DOI: https://dx.doi.org/10.3389/fvets.2022.837229] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/35400103]
14. Kilcawley, K.N.; Faulkner, H.; Clarke, H.J.; O’Sullivan, M.G.; Kerry, J.P. Factors Influencing the Flavour of Bovine Milk and Cheese from Grass Based versus Non-Grass Based Milk Production Systems. Foods; 2018; 7, 37. [DOI: https://dx.doi.org/10.3390/foods7030037] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/29534042]
15. Serrapica, F.; Uzun, P.; Masucci, F.; Napolitano, F.; Braghieri, A.; Genovese, A.; Sacchi, R.; Romano, R.; Barone, C.M.A.; Di Francia, A. Hay or silage? How the forage preservation method changes the volatile compounds and sensory properties of Caciocavallo cheese. J. Dairy Sci.; 2020; 103, pp. 1391-1403. [DOI: https://dx.doi.org/10.3168/jds.2019-17155] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/31785866]
16. Scherer, R.; Gerlach, K.; Südekum, K.H. Biogenic amines and gamma-amino butyric acid in silages: Formation, occurrence and influence on dry matter intake and ruminant production. Anim. Feed Sci. Technol.; 2015; 210, pp. 1-16. [DOI: https://dx.doi.org/10.1016/j.anifeedsci.2015.10.001]
17. Gerlach, K.; Daniel, J.L.P.; Jobim, C.C.; Nussio, L.G. A data analysis on the effect of acetic acid on dry matter intake in dairy cattle. Anim. Feed Sci. Technol.; 2021; 272, 114782. [DOI: https://dx.doi.org/10.1016/j.anifeedsci.2020.114782]
18. Song, H.; Liu, J. GC-O-MS technique and its applications in food flavor analysis. Food Res. Int.; 2018; 114, pp. 187-198. [DOI: https://dx.doi.org/10.1016/j.foodres.2018.07.037]
19. d’Acampora Zellner, B.; Dugo, P.; Dugo, G.; Mondello, L. Gas chromatography–olfactometry in food flavour analysis. J. Chromatogr. A; 2008; 1186, pp. 123-143. [DOI: https://dx.doi.org/10.1016/j.chroma.2007.09.006]
20. Wang, S.; Chen, H.; Sun, B. Recent progress in food flavor analysis using gas chromatography–ion mobility spectrometry (GC–IMS). Food Chem.; 2020; 315, 126158. [DOI: https://dx.doi.org/10.1016/j.foodchem.2019.126158]
21. Wei, G.; Dan, M.; Zhao, G.; Wang, D. Recent advances in chromatography-mass spectrometry and electronic nose technology in food flavor analysis and detection. Food Chem.; 2023; 405, 134814. [DOI: https://dx.doi.org/10.1016/j.foodchem.2022.134814]
22. Shooshtari, M.; Salehi, A. An electronic nose based on carbon nanotube-titanium dioxide hybrid nanostructures for detection and discrimination of volatile organic compounds. Sens. Actuators B Chem.; 2022; 357, 131418. [DOI: https://dx.doi.org/10.1016/j.snb.2022.131418]
23. Yu, M.; Yang, P.; Song, H.; Guan, X. Research progress in comprehensive two-dimensional gas chromatography-mass spectrometry and its combination with olfactometry systems in the flavor analysis field. J. Food Compos. Anal.; 2022; 114, 104790. [DOI: https://dx.doi.org/10.1016/j.jfca.2022.104790]
24. Fan, X.; Jiao, X.; Liu, J.; Jia, M.; Blanchard, C.; Zhou, Z. Characterizing the volatile compounds of different sorghum cultivars by both GC-MS and HS-GC-IMS. Food Res. Int.; 2021; 140, 109975. [DOI: https://dx.doi.org/10.1016/j.foodres.2020.109975]
25. Li, C.; Al-Dalali, S.; Wang, Z.; Xu, B.; Zhou, H. Investigation of volatile flavor compounds and characterization of aroma-active compounds of water-boiled salted duck using GC–MS–O, GC–IMS, and E-nose. Food Chem.; 2022; 386, 132728. [DOI: https://dx.doi.org/10.1016/j.foodchem.2022.132728] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/35509168]
26. Chen, Y.; Zhang, X.; Liu, X.; Liu, Y.; Hou, A.; Wang, Y.; Li, L.; Peng, X.; Xiao, Y. Discrimination and characterization of volatile organic compounds and nutritional values of three varieties of chopped pepper seeds. Food Chem. X; 2024; 21, 101150. [DOI: https://dx.doi.org/10.1016/j.fochx.2024.101150] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/38312485]
27. Hu, X.; Lu, L.; Guo, Z.; Zhu, Z. Volatile compounds, affecting factors and evaluation methods for rice aroma: A review. Trends Food Sci. Technol.; 2020; 97, pp. 136-146.
28. Krizsan, S.J.; Westad, F.; Ådnøy, T.; Odden, E.; Aakre, S.E.; Randby, Å.T. Effect of volatile compounds in grass silage on voluntary intake by growing cattle. Animal; 2007; 1, pp. 283-292. [DOI: https://dx.doi.org/10.1017/S1751731107683773] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/22444294]
29. Ashrafi, A.M.; Bytešníková, Z.; Cané, C.; Richtera, L.; Vallejos, S. New trends in methyl salicylate sensing and their implications in agriculture. Biosens. Bioelectron.; 2023; 223, 115008. [DOI: https://dx.doi.org/10.1016/j.bios.2022.115008]
30. Jayasekara, T.K.; Stevenson, P.C.; Belmain, S.R.; Farman, D.I.; Hall, D.R. Identification of methyl salicylate as the principal volatile component in the methanol extract of root bark of Securidaca longepedunculata Fers. J. Mass Spectrom.; 2002; 37, pp. 577-580. [DOI: https://dx.doi.org/10.1002/jms.314]
31. Singewar, K.; Fladung, M.; Robischon, M. Methyl salicylate as a signaling compound that contributes to forest ecosystem stability. Trees; 2021; 35, pp. 1755-1769. [DOI: https://dx.doi.org/10.1007/s00468-021-02191-y]
32. Nievierowski, T.H.; Veras, F.F.; Silveira, R.D.; Dachery, B.; Hernandes, K.C.; Lopes, F.C.; Scortegagna, E.; Zini, C.A.; Welke, J.E. Role of partial dehydration in a naturally ventilated room on the mycobiota, ochratoxins, volatile profile and phenolic composition of Merlot grapes intended for wine production. Food Res. Int.; 2021; 141, 110145. [DOI: https://dx.doi.org/10.1016/j.foodres.2021.110145]
33. Wang, D.; Javed, H.U.; Shi, Y.; Naz, S.; Ali, S.; Duan, C.-Q. Impact of Drying Method on the Evaluation of Fatty Acids and Their Derived Volatile Compounds in ‘Thompson Seedless’ Raisins. Molecules; 2020; 25, 608. [DOI: https://dx.doi.org/10.3390/molecules25030608]
34. Wang, J.; Yang, Z.J.; Xu, L.Y.; Wang, B.; Zhang, J.H.; Li, B.Z.; Cao, Y.P.; Tan, L. Key aroma compounds identified in Cheddar cheese with different ripening times by aroma extract dilution analysis, odor activity value, aroma recombination, and omission. J. Dairy Sci.; 2021; 104, pp. 1576-1590. [DOI: https://dx.doi.org/10.3168/jds.2020-18757]
35. Bertuzzi, A.S.; McSweeney, P.L.H.; Rea, M.C.; Kilcawley, K.N. Detection of Volatile Compounds of Cheese and Their Contribution to the Flavor Profile of Surface-Ripened Cheese. Compr. Rev. Food Sci. Food Saf.; 2018; 17, pp. 371-390. [DOI: https://dx.doi.org/10.1111/1541-4337.12332]
36. Luo, Q.; Gu, L.; Wu, Z.; Shan, Y.; Wang, H.; Sun, L.-N. Distribution, source apportionment and ecological risks of organophosphate esters in surface sediments from the Liao River, Northeast China. Chemosphere; 2020; 250, 126297. [DOI: https://dx.doi.org/10.1016/j.chemosphere.2020.126297] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/32113093]
37. Liu, M.; Wang, M.; Zhao, A.; Zhu, L.; Wang, C.; Wei, C.; Liu, W.; Xu, J. Determination of organophosphate diesters in facility vegetable soils using ultra-high performance liquid chromatography-electrostatic field orbitrap high resolution mass spectrometry. Se Pu Chin. J. Chromatogr.; 2023; 41, pp. 58-65. [DOI: https://dx.doi.org/10.3724/SP.J.1123.2022.05002]
38. Ma, Z.-Y.; Xiao, M.; Li, C.-K.; Shoberu, A.; Zou, J.-P. S-(1,3-Dioxoisoindolin-2-yl)O, O-diethyl phosphorothioate (SDDP): A practical electrophilic reagent for the phosphorothiolation of electron-rich compounds. Chin. Chem. Lett.; 2024; 35, 109076. [DOI: https://dx.doi.org/10.1016/j.cclet.2023.109076]
39. Kumar, S.; Kaushik, G.; Dar, M.A.; Nimesh, S.; LÓPez-Chuken, U.J.; Villarreal-Chiu, J.F. Microbial Degradation of Organophosphate Pesticides: A Review. Pedosphere; 2018; 28, pp. 190-208. [DOI: https://dx.doi.org/10.1016/S1002-0160(18)60017-7]
40. Palliyaguru, D.L.; Yuan, J.-M.; Kensler, T.W.; Fahey, J.W. Isothiocyanates: Translating the Power of Plants to People. Mol. Nutr. Food Res.; 2018; 62, e1700965. [DOI: https://dx.doi.org/10.1002/mnfr.201700965]
41. Bell, L.; Oloyede, O.O.; Lignou, S.; Wagstaff, C.; Methven, L. Taste and Flavor Perceptions of Glucosinolates, Isothiocyanates, and Related Compounds. Mol. Nutr. Food Res.; 2018; 62, 1700990. [DOI: https://dx.doi.org/10.1002/mnfr.201700990] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/29578640]
42. Suh, S.-J.; Moon, S.-K.; Kim, C.-H. Raphanus sativus and its isothiocyanates inhibit vascular smooth muscle cells proliferation and induce G1 cell cycle arrest. Int. Immunopharmacol.; 2006; 6, pp. 854-861. [DOI: https://dx.doi.org/10.1016/j.intimp.2005.11.014] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/16546717]
43. Ildikó, S.-G.; Klára, K.A.; Marianna, T.-M.; Ágnes, B.; Zsuzsanna, M.-B.; Bálint, C. The effect of radio frequency heat treatment on nutritional and colloid-chemical properties of different white mustard (Sinapis alba L.) varieties. Innov. Food Sci. Emerg. Technol.; 2006; 7, pp. 74-79. [DOI: https://dx.doi.org/10.1016/j.ifset.2005.06.001]
44. Kim, M.G.; Lee, H.S. Growth-Inhibiting Activities of Phenethyl Isothiocyanate and Its Derivatives against Intestinal Bacteria. J. Food Sci.; 2009; 74, pp. M467-M471. [DOI: https://dx.doi.org/10.1111/j.1750-3841.2009.01333.x]
45. Mattila, J.M.; Lakey, P.S.J.; Shiraiwa, M.; Wang, C.; Abbatt, J.P.D.; Arata, C.; Goldstein, A.H.; Ampollini, L.; Katz, E.F.; DeCarlo, P.F. et al. Multiphase Chemistry Controls Inorganic Chlorinated and Nitrogenated Compounds in Indoor Air during Bleach Cleaning. Environ. Sci. Technol.; 2020; 54, pp. 1730-1739. [DOI: https://dx.doi.org/10.1021/acs.est.9b05767] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/31940195]
46. Mattila, J.M.; Arata, C.; Wang, C.; Katz, E.F.; Abeleira, A.; Zhou, Y.; Zhou, S.; Goldstein, A.H.; Abbatt, J.P.D.; DeCarlo, P.F. et al. Dark Chemistry during Bleach Cleaning Enhances Oxidation of Organics and Secondary Organic Aerosol Production Indoors. Environ. Sci. Technol. Lett.; 2020; 7, pp. 795-801. [DOI: https://dx.doi.org/10.1021/acs.estlett.0c00573]
47. Zhang, Q. Study on the Method of Rapid Field Detection of Urea and CyanuricAcid in Swimming Pool Water; Zhengzhou University: Zhengzhou, China, 2022.
48. Boor, B.E.; Liang, Y.; Crain, N.E.; Järnström, H.; Novoselac, A.; Xu, Y. Identification of Phthalate and Alternative Plasticizers, Flame Retardants, and Unreacted Isocyanates in Infant Crib Mattress Covers and Foam. Environ. Sci. Technol. Lett.; 2015; 2, pp. 89-94. [DOI: https://dx.doi.org/10.1021/acs.estlett.5b00039]
49. Noci, F.; Monahan, F.J.; Scollan, N.D.; Moloney, A.P. The fatty acid composition of muscle and adipose tissue of steers offered unwilted or wilted grass silage supplemented with sunflower oil and fishoil. Br. J. Nutr.; 2007; 97, pp. 502-513. [DOI: https://dx.doi.org/10.1017/S0007114507381361]
50. Wan, J.; Liu, Q.; Ma, C.; Muhoza, B.; Huang, Y.; Sun, M.; Song, S.; Ho, C.-T. Characteristic flavor fingerprint disclosure of dzo beef in Tibet by applying SAFE-GC-O-MS and HS-GC-IMS technology. Food Res. Int.; 2023; 166, 112581. [DOI: https://dx.doi.org/10.1016/j.foodres.2023.112581] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/36914343]
51. Yang, C.; Liu, J.; Wu, X.; Bao, P.; Long, R.; Guo, X.; Ding, X.; Yan, P. The response of gene expression associated with lipid metabolism, fat deposition and fatty acid profile in the longissimus dorsi muscle of Gannan yaks to different energy levels of diets. PLoS ONE; 2017; 12, e0187604. [DOI: https://dx.doi.org/10.1371/journal.pone.0187604] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/29121115]
52. He, X.-Y.; Wu, L.-J.; Wang, W.-X.; Xie, P.-J.; Chen, Y.-H.; Wang, F. Amygdalin—A pharmacological and toxicological review. J. Ethnopharmacol.; 2020; 254, 112717. [DOI: https://dx.doi.org/10.1016/j.jep.2020.112717] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/32114166]
53. Barral-Martinez, M.; Fraga-Corral, M.; Garcia-Perez, P.; Simal-Gandara, J.; Prieto, M.A. Almond By-Products: Valorization for Sustainability and Competitiveness of the Industry. Foods; 2021; 10, 1793. [DOI: https://dx.doi.org/10.3390/foods10081793] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/34441570]
54. Siddiqui, A.A.; Sohail, A.; Bhat, S.A.; Rehman, M.T.; Bano, B. Non-enzymatic Glycation of Almond Cystatin Leads to Conformational Changes and Altered Activity. Protein Pept. Lett.; 2015; 22, pp. 449-459. [DOI: https://dx.doi.org/10.2174/0929866522666150326105704] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/25808045]
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Details
1 College of Animal Science, Shanxi Agricultural University, Jinzhong 030108, China;
2 College of Animal Science, Shanxi Agricultural University, Jinzhong 030108, China;
3 State Key Laboratory of Animal Nutrition and Feeding, Institute of Animal Sciences, Chinese Academy of Agricultural Sciences, Beijing 100193, China;