Optimization of Jinhua Ham Classification Method Based on Volatile Flavor Substances and Determination of Key Odor Biomarkers

Jinhua ham is a traditional cured meat food in China. For a long time, its grade has mainly been evaluated by the human nose through the three-sticks method, which is highly subjective and is not conducive to establishing evaluation standards through odor markers. In this paper, we analyzed the well-graded Grade I–III hams provided by Jinzi Ham Co., Ltd. (Jinhua, China). Firstly, we used different extraction fibers, extraction temperatures, and extraction time to determine the optimal conditions for headspace solid-phase microextraction (HS-SPME). Then, the aroma components of Jinhua ham were analyzed by headspace solid-phase microextraction combined with gas chromatography–mass spectrometry (GC–MS), and OAV was calculated to screen the key aroma volatiles of three kinds of Jinhua ham. It was found that a total of 56 components were detected in the three types of ham. Among them, there are 21 kinds of key aroma volatiles. Aldehydes, alcohols, and acids are the three main components of Jinhua ham, and the content of aldehydes gradually decreases from Grade I to Grade III ham. The content of acids gradually increased, and we speculated that the increase in acid content was caused by the proliferation of microorganisms in Grade III ham. The key flavor volatiles in Grade I hams was hexanal and 2-methylbutanal. Grade I hams had a strong meat aroma, pleasant fatty, and roasted aroma without any off-flavors. In Grade II ham, the characteristic volatiles (E,E)-2,4-decadienal and ethyl isovalerate were detected. These two volatiles contribute greatly to the flavor of Grade II ham, which makes the flavor of Grade II ham have a special fruity aroma. They also may be prone to sourness and affect the flavor of the ham. Volatiles with low threshold values, such as pyrazines, furans, and sulfur-containing compounds, were relatively high in Grade III hams. This may also contribute to the poorer flavor quality of Grade III hams. This experiment provided a reliable test method and evaluation basis for the rating of Jinhua ham. These results have positive implications for the establishment of odor markers-based grading criteria.     

Decaffeination and Neuraminidase Inhibitory Activity of Arabica Green Coffee (Coffea arabica) Beans: Chlorogenic Acid as a Potential Bioactive Compound

Coffee has been studied for its health benefits, including prevention of several chronic diseases, such as type 2 diabetes mellitus, cancer, Parkinson’s, and liver diseases. Chlorogenic acid (CGA), an important component in coffee beans, was shown to possess antiviral activity against viruses. However, the presence of caffeine in coffee beans may also cause insomnia and stomach irritation, and increase heart rate and respiration rate. These unwanted effects may be reduced by decaffeination of green bean Arabica coffee (GBAC) by treatment with dichloromethane, followed by solid-phase extraction using methanol. In this study, the caffeine and chlorogenic acid (CGA) level in the coffee bean from three different areas in West Java, before and after decaffeination, was determined and validated using HPLC. The results showed that the levels of caffeine were reduced significantly, with an order as follows: Tasikmalaya (2.28% to 0.097% (97 ppm), Pangalengan (1.57% to 0.049% (495 ppm), and Garut (1.45% to 0.00002% (0.2 ppm). The CGA levels in the GBAC were also reduced as follows: Tasikmalaya (0.54% to 0.001% (118 ppm), Pangalengan (0.97% to 0.0047% (388 ppm)), and Garut (0.81% to 0.029% (282 ppm). The decaffeinated samples were then subjected to the H5N1 neuraminidase (NA) binding assay to determine its bioactivity as an anti-influenza agent. The results show that samples from Tasikmalaya, Pangalengan, and Garut possess NA inhibitory activity with IC₅₀ of 69.70, 75.23, and 55.74 μg/mL, respectively. The low level of caffeine with a higher level of CGA correlates with their higher levels of NA inhibitory, as shown in the Garut samples. Therefore, the level of caffeine and CGA influenced the level of NA inhibitory activity. This is supported by the validation of CGA-NA binding interaction via molecular docking and pharmacophore modeling; hence, CGA could potentially serve as a bioactive compound for neuraminidase activity in GBAC.     

Determination of caffeine in coffee products by dynamic complexation with 3,4-dimethoxycinnamate and separation by CZE

A method based on the formation of pi-complexes with chlorogenate-like species was proposed for the determination of caffeine in regular (nondecaffeinated) and decaffeinated coffee. Both caffeate and 3,4-dimethoxycinnamate were able to transform caffeine--a neutral species in aqueous solutions--into an anionic species. The usage of 3,4-dimethoxycinnamate in the running electrolyte is advantageous, because of its greater chemical stability and the improved resolution of the peaks of caffeine, theobromine, and theophylline. Negative peaks were registered with a capacitively coupled contactless conductivity detector when solutions of these alkylxanthines were analyzed with a BGE composed of 20 mmol/L 3,4-dimethoxycinnamic acid and pH adjusted to 8.5 with Tris. This behavior was expected, because the complex is larger and thus should move slower than the free anion. Caffeine was determined in ground and instant coffee with precision and accuracy that meet Brazilian norms about such products. The LOD was estimated as 33 mg/L, which corresponds to 0.8 and 0.3 mg of caffeine per gram of dry instant coffee and ground coffee, respectively. For the case of decaffeinated coffee, ten times preconcentration with dichloromethane was carried out to allow the quantitation of caffeine, which should not exceed the concentration of 1 mg/g in dry matter.     

Profiles of sulfur containing compounds obtained from Arabica and Robusta coffees by capillary column gas chromatography

Summary The profiles of sulfur containing compounds present in both Arabica and Robusta coffees were determined by concentrating the volatiles from dry ground roasted coffee headspace, pressed coffee oil or brewed coffee headspace onto an adsorbent (Tenax) which was then thermally desorbed into a capillary column. The volatiles were then chromatographed and detected with a flame photometric detector. Significant quantitative differences between the profiles of Arabica coffee and Robusta coffee are found for all three sample types with a few components being up to 20-times more concentrated in the Robusta coffee than in the Arabica coffee. These differences may be utilized to detect as little as 1 % Robusta coffee present in Arabica coffee.     

Antioxidant and Sensory Assessment of Innovative Coffee Blends of Reduced Caffeine Content

Considering the current trend in the global coffee market, which involves an increased demand for decaffeinated coffee, the aim of the present study was to formulate coffee blends with reduced caffeine content, but with pronounced antioxidant and attractive sensory properties. For this purpose, green and roasted Arabica and Robusta coffee beans of different origins were subjected to the screening analysis of their chemical and bioactive composition using standard AOAC, spectrophotometric and chromatographic methods. From roasted coffee beans, espresso, Turkish and filter coffees were prepared, and their sensory evaluation was performed using a 10-point hedonic scale. The results showed that Arabica coffee beans were richer in sucrose and oil, while Robusta beans were characterized by higher content of all determined bioactive parameters. Among all studied samples, the highest content of 3-O-caffeoylquinic acid (14.09 mg g⁻¹ dmb), 4-O-caffeoylquinic acid (8.23 mg g⁻¹ dmb) and 5-O-caffeoylquinic acid (4.65 mg g⁻¹ dmb), as well as caffeine (22.38 mg g⁻¹ dmb), was detected in roasted Robusta beans from the Minas Gerais region of Brazil, which were therefore used to formulate coffee blends with reduced caffeine content. Robusta brews were found to be more astringent and recognized as more sensorily attractive, while Arabica decaffeinated brews were evaluated as more bitter. The obtained results point out that coffee brews may represent a significant source of phenolic compounds, mainly caffeoylquinic acids, with potent antioxidant properties, even if they have reduced caffeine content.     

Caffeine and theobromine in coffee, tea and cola-beverages

Summary A new method based on reversed-phase ion-interaction HPLC is described for the identification and quantitation of theobromine, theophylline and caffeine in beverages. The interaction reagent used is octylamine orto-phosphate which also constitutes the mobile phase. The stationary phase is a microparticulate reversed-phase C-18 packing. With spectrophotometric detection at 274 nm, detection levels of 0.15, 0.30 and 0.40 ppm were achieved for theobromine, theophylline and caffeine, respectively.Quantitative analysis was performed by the standard addition method for theobromine and caffeine contents in tea, espresso-coffee, decaffeinated coffee, decaffeinated tea and cola-beverages.     

Characterization of the Aroma Profile and Main Key Odorants of Espresso Coffee

Espresso coffee (EC) is a common coffee preparation technique that nowadays is broadly widespread all over the globe. Its popularity is in part attributed to the intense aroma and pleasant flavor. Many researchers have studied and reviewed the aroma of the coffee, but there is a lack of specific review focused on EC aroma profile even if it is intensively investigated. Thus, the objective of the current review was to summarize the aroma profile of EC and how different preparation variables can affect EC flavor. Moreover, a collection of diverse analytical procedures for volatile analysis was also reported. The findings of this survey showed that the volatile fraction of EC is extremely complex, but just some compounds are responsible for the characteristic aroma of the coffee, such as some aldehyde, ketones, furanones, furans, sulfur compounds, pyrazines, etc. In addition, during preparation, some variables, e.g., temperature and pressure of water, granulometry of the coffee particle, and brew ratio, can also modify the aroma profile of this beverage, and therefore its quality. A better understanding of the aroma fraction of EC and how the preparation variables should be adjusted according to desired EC would assist coffee workers in obtaining a higher quality product.     

Single Origin Coffee Aroma: From Optimized Flavor Protocols and Coffee Customization to Instrumental Volatile Characterization and Chemometrics

In this study, the aroma profile of 10 single origin Arabica coffees originating from eight different growing locations, from Central America to Indonesia, was analyzed using Headspace SPME-GC-MS as the analytical method. Their roasting was performed under temperature–time conditions, customized for each sample to reach specific sensory brew characteristics in an attempt to underline the customization of roast profiles and implementation of separate roastings followed by subsequent blending as a means to tailor cup quality. A total of 138 volatile compounds were identified in all coffee samples, mainly furan (~24–41%) and pyrazine (~25–39%) derivatives, many of which are recognized as coffee key odorants, while the main formation mechanism was the Maillard reaction. Volatile compounds’ composition data were also chemometrically processed using the HCA Heatmap, PCA and HCA aiming to explore if they meet the expected aroma quality attributes and if they can be an indicator of coffee origin. The desired brew characteristics of the samples were satisfactorily captured from the volatile compounds formed, contributing to the aroma potential of each sample. Furthermore, the volatile compounds presented a strong variation with the applied roasting conditions, meaning lighter roasted samples were efficiently differentiated from darker roasted samples, while roasting degree exceeded the geographical origin of the coffee. The coffee samples were distinguished into two groups, with the first two PCs accounting for 73.66% of the total variation, attributed mainly to the presence of higher quantities of furans and pyrazines, as well as to other chemical classes (e.g., dihydrofuranone and phenol derivatives), while HCA confirmed the above results rendering roasting conditions as the underlying criterion for differentiation.     

Analysis of the headspace of foodstuffs near room temperature

Formation of volatiles in onion varieties, aging of coffee, and the quality of birch syrup were studied by using near room temperature headspace GC-MS analysis. Accurate volumes (1–500 mL) of the gaseous samples were introduced into the capillary column either by a syringe or by sucking with a pump. The standard deviation of the contents was typically 1-7% depending on the foodstuff and the compound. The ratio of 2,3-butanedione to 2-methylfuran decreased during the storage of coffee and was a typical measure of aging. Analysis of even the very polar 2,5-dimethyl-4-hydroxy-3(2H)-furanone in the headspace of birch syrup was possible. The distribution patterns of the sulfur-containing volatiles generated by crushed onions were close to each other in different varieties. The rate of the total formation of the compounds varied from cultivar to cultivar.     

Development of a sensitive non-targeted method for characterizing the wine volatile profile using headspace solid-phase microextraction comprehensive two-dimensional gas chromatography time-of-flight mass spectrometry

Future understanding of differences in the composition and sensory attributes of wines require improved analytical methods which allow the monitoring of a large number of volatiles including those present at low concentrations. This study presents the optimization and application of a headspace solid-phase microextraction (HS-SPME) method for analysis of wine volatiles by comprehensive two-dimensional gas chromatography (GC×GC) time-of-flight mass spectrometry (TOFMS). This study demonstrates an important advancement in wine volatile analysis as the method allows for the simultaneous analysis of a significantly larger number of compounds found in the wine headspace compared to other current single dimensional GC-MS methodologies. The methodology allowed for the simultaneous analysis of over 350 different tentatively identified volatile and semi-volatile compounds found in the wine headspace. These included potent aroma compound classes such as monoterpenes, norisoprenoids, sesquiterpenes, and alkyl-methoxypyrazines which have been documented to contribute to wine aroma. It is intended that wine aroma research and wine sensory research will utilize this non-targeted method to assess compositional differences in the wine volatile profile.     

Tandem Solid-Phase Extraction Columns for Simultaneous Aroma Extraction and Fractionation of Wuliangye and Other Baijiu

Wuliangye baijiu is one of the most famous baijiu in China, with a rich, harmonic aroma profile highly appreciated by consumers. Thousands of volatiles have been identified for the unique aroma profile. Among them, fatty acid esters have been identified as the main contributors to the aroma profile. In addition, many non-ester minor compounds, many of which are more polar than the esters, have been identified to contribute to the characteristic aroma unique to Wuliangye baijiu. The analysis of these minor compounds has been challenging due to the dominance of esters in the sample. Thus, it is desirable to fractionate the aroma extract into subgroups based on functional group or polarity to simplify the analysis. This study attempts a new approach to achieve simultaneous volatile extraction and fractionation using tandem LiChrolut EN and silica gel solid-phase extraction (SPE) columns. A baijiu sample (10 mL, diluted in 40 mL of water) was first passed through the LiChrolut EN (1.0 g) column. The loaded LiChrolut EN column was then dried with air and coupled with a silica gel (5.0 g) SPE column with anhydrous Na₂SO₄ (10.0 g) in between. The volatile compounds were eluted from the LiChrolut EN column and simultaneously fractionated on the silica gel column based on polarity. The simultaneous extraction and fractionation technique enabled the fractionations of all fatty acid esters into less polar fractions. Fatty acids, alcohols, pyrazines, furans, phenols, hydroxy esters, and other polar compounds were collected in more polar fractions. This technique was used to study the volatile compounds in Wuliangye, Moutai, and Fengjiu baijiu. In addition to fatty acid esters, many minor polar compounds, including 2,6-dimethylpyrazine, 2-ethyl-6-methylpyrazine, 2-ethyl-3,5-dimethylpyrazine, p-cresol, and 2-acetylpyrrole, were unequivocally identified in the samples. The procedure is fast and straightforward, with low solvent consumption.     

A high-throughput platform for low-volume high-temperature/pressure sealed vessel solvent extractions

A high-throughput platform for performing parallel solvent extractions in sealed HPLC/GC vials inside a microwave reactor is described. The system consist of a strongly microwave-absorbing silicon carbide plate with 20 cylindrical wells of appropriate dimensions to be fitted with standard HPLC/GC autosampler vials serving as extraction vessels. Due to the possibility of heating up to four heating platforms simultaneously (80 vials), efficient parallel analytical-scale solvent extractions can be performed using volumes of 0.5-1.5 mL at a maximum temperature/pressure limit of 200°C/20 bar. Since the extraction and subsequent analysis by either gas chromatography or liquid chromatography coupled with mass detection (GC-MS or LC-MS) is performed directly from the autosampler vial, errors caused by sample transfer can be minimized. The platform was evaluated for the extraction and quantification of caffeine from commercial coffee powders assessing different solvent types, extraction temperatures and times. For example, 141±11 μg caffeine (5 mg coffee powder) were extracted during a single extraction cycle using methanol as extraction solvent, whereas only 90±11 were obtained performing the extraction in methylene chloride, applying the same reaction conditions (90°C, 10 min). In multiple extraction experiments a total of ~150 μg caffeine was extracted from 5 mg commercial coffee powder. In addition to the quantitative caffeine determination, a comparative qualitative analysis of the liquid phase coffee extracts and the headspace volatiles was performed, placing special emphasis on headspace analysis using solid-phase microextraction (SPME) techniques. The miniaturized parallel extraction technique introduced herein allows solvent extractions to be performed at significantly expanded temperature/pressure limits and shortened extraction times, using standard HPLC autosampler vials as reaction vessels. Remarkable differences regarding peak pattern and main peaks were observed when low-temperature extraction (60°C) and high-temperature extraction (160°C) are compared prior to headspace-SPME-GC-MS performed in the same HPLC/GC vials.     

Volatile enrichment on automatic static headspace using a precolumn and stopping the carrier gas flow during injection

The aim of this study was to develop a technique for performing automatic static headspace analysis of volatiles without a cryogenic device. Reconcentration of solutes was accomplished using a graphitized carbon coated precolumn between two splitting points and stopping carrier gas flow during injection. Chromatographic profiles of volatile compounds from ground coffee compared with split and splitless injections provide confirmation of the good sensitivity of the technique. The analysis of a standard mixture covering a wide range of volatility confirms that the described technique might be useful to achieve enrichement of low volatile headspace compounds, even if discrimination against the various components is present.     

Developing qualitative extraction profiles of coffee aromas utilizing polymeric ionic liquid sorbent coatings in headspace solid-phase microextraction gas chromatography-mass spectrometry

Two solid-phase microextraction (SPME) sorbent coatings based on polymeric ionic liquids (PILs) have been utilized for the analysis of complex coffee aroma samples. The PIL-based SPME coatings examined, namely, poly(1-(4-vinylbenzyl)-3-hexadecylimidazolium bis[(trifluoromethyl)sulfonyl]imide) [poly(VBHDIm⁺ NTf₂⁻)], with ∼14-μm thickness, and poly(1-vinyl-3-hexylimidazolium chloride) [poly(ViHIm⁺ Cl⁻)], with ∼8-μm thickness, were employed for the headspace determination of up to 49 analytes from four different coffee beans: two French roast coffees of different brands, Sumatra coffee, and decaffeinated Sumatra coffee. The analysis was conducted using gas chromatography coupled to mass spectrometry. For comparative purposes, the commercial polyacrylate (PA, 85-μm film thickness) SPME coating was utilized under the same extraction conditions. The three SPME coatings tested behaved quite differently as a function of the families of compounds extracted. Thus, the poly(VBHDIm⁺ NTf₂⁻) coating was extremely selective for aldehydes while also exhibiting good extraction efficiencies for acids. The poly-(ViHIm⁺ Cl⁻) coating exhibited superior performance for aromatic alcohols, and the PA coating worked better for heterocyclic aromatics. Both PIL-based SPME sorbent coatings demonstrated exceptional selectivity and extraction efficiency when dealing with complex coffee aromas in spite of their small film thicknesses.     

Effect of maturation in small oak casks on the volatility of red wine aroma compounds

Wood maturation of red wine produces complex interactions between oak extracted compounds and pre-existing components in wine. Wood contributes with some aroma volatiles; but the whole volatile fraction, including the volatiles extracted from grape or produced during fermentation, could be involved in interactions with the non-volatile fraction. Samples of red wine with increasing time of wood contact, matured in 25 l casks of new American oak, were analysed on their volatile composition both in solution and headspace, and phenolic components. There was an increase of acetic acid and acetate esters as the time of wood contact increased. On the other hand, ethyl esters decreased as the time in wood increased. Some volatiles showed different behaviour when comparing their relationship between headspace and solution from two different casks, even though they were treated as replicates. This different evolution among casks was also consistent with different evolution of phenolic compounds, suggesting that there was an interaction among the volatile fraction and phenolic components.     

A preliminary study of plant aroma profile characteristics by a combination sampling method coupled with GC-MS

Plant aroma profile characteristic is an important bio-information. In this study, a sampling method in combination with headspace solid phase microextraction (HSSPME), simultaneous distillation extraction (SDE) and steam distillation (SD), followed by gas chromatography-mass spectrometry (GC-MS) detection, was used to study the aroma profile characteristics of the fresh and stale samples of common tomato, cherry tomato, durian and longan, and of the different samples of Chinese mango and Allium varieties. The typical aroma volatiles of these samples were isolated and identified according to the different degrees of certainty. The different samples showed different aroma profile characteristics when principal component analysis (PCA) was conducted to analyze the data of the aroma profile chromatograms. Then the crucial aroma volatiles contributing greatly to the clustering differences of the aroma profile characteristics of the fresh and stale common tomato, cherry tomato, durian and longan were identified by common model strategy. These compounds are potential bio-markers of plant metabolism, but further study is needed. Continuous investigations for the aroma profile characteristics of common tomato during storage and chive during growth were conducted to distill the potential bio-information from the plant metabolism processes. The saturated hexanal of common tomato increased during storage, whereas the unsaturated hexenal reduced. The accumulating trends of volatile sulfides were observed during chive growth. The preliminary results related with the corresponding bio-information could provide helpful clues to the study of plant's secondary metabolism process and benefit quality control.     

Multi-channel open tubular traps for headspace sampling, gas chromatographic fraction collection and olfactory assessment of milk volatiles

A headspace sampling method is described for concentrating milk volatiles onto a multi-channel open tubular silicone rubber trap (MCT) for thermal desorption into a GC-FID. Sections of the chromatographic profile, single peaks or combinations of compounds are recaptured with secondary MCTs during a subsequent run. The recaptured aroma is released in a controlled manner by heating the MCT in a portable heating device. An aroma release window of several minutes allows up to six people the opportunity to sniff each aroma fraction more than once. Olfactory results suggest that a synergistic combination of 2-heptanone and 2-nonanone could be responsible for a pungent cheese, sour milk-like aroma. MCTs containing single components or fractions can be desorbed into a GC-MS for compound identification.     

Evaluation of two commercial solid-phase microextraction fibres for the analysis of target aroma compounds in cooked beef meat

The aroma profile of cooked beef meat has been investigated by solid-phase microextraction (SPME) combined with gas chromatography-mass spectrometry (GC-MS). Out of more than 200 volatile compounds, 36 key odour-active compounds were selected for analysis. Several extraction times, desorption times, temperature conditions and fibre types were tested to achieve a fast and reproducible extraction, and a representative analysis of the aroma profile of cooked beef. Extraction conditions and fibre type significantly affected the majority of the target compounds. Divinylbenzene-carboxen-polydimethylsiloxane (DVB-CAR-PDMS) fibre presented a better reproducibility at all extraction times and extracted more efficiently the less volatile compounds than the carboxen-polydimethylsiloxane (CAR-PDMS) fibre. The high molecular weight compounds seemed to achieve faster the equilibrium between the headspace and DVB-CAR-PDMS fibre. The use of SPME was shown to be a simple, sensitive, selective, representative, fast, and low-cost method for the evaluation of key odour-active compounds in cooked beef meat, even if further research on quantitative analysis of volatiles using SPME on solid samples has to be done.     

New genetic insights to consider coffee waste as feedstock for fuel,feed, and chemicals

Caffeine is a natural plant product found in many drinks, including coffee, tea, soft and energy drinks. Due to caffeine’s presence in the environment, microorganisms have evolved two different mechanisms to live on caffeine. The genetic maps of the caffeine N-demethylation pathway and C-8 oxidation pathway have been discovered in Pseudomonas putida CBB5 and Pseudomonas sp. CBB1, respectively. These genes are the only characterized bacterial caffeine-degrading genes, and may be of great value in producing fine chemicals, biofuels, and animal feed from coffee and tea waste. Here, we present preliminary results for production of theobromine and 7-methylxanthine from caffeine and theobromine, respectively, by two strains of metabolically engineered E. coli. We also demonstrate complete decaffeination of tea extract by an immobilized mixed culture of Klebsiella and Rhodococcus cells. These processes provide a first level demonstration of biotechnological utilization of coffee and tea waste.      

Über den Einbau von Trichloräthylen in Inhaltsstoffe des Kaffees bei dessen Decoffeinierung

After decaffeination of green coffee beans by extraction with [¹⁴C]-trichloroethylene, the beans were radioactive and did not loose their activity during roasting. On the basis of the radioactivity uptake, it is estimated that during commercial decaffeination 0.25 to 1 g of trichloroethylene is incorporated per kg of beans. The nature of the compound or compounds formed on interaction of trichloroethylene with coffee constituents is still unknown, but attention is directed to the possibility of the formation of highly toxic S-dichlorovinyl derivatives as is the case in the extraction of soya beans with trichloroethylene.