Use of methylpyrazine ratios to monitor the coffee roasting

The formation of methylpyrazines has been determined in roasted coffee beans (Arabica and Robusta). The determination of different methylpyrazines was studied by the coupled steam distillation-microdistillator as extraction method and gas chromatography using a capillary column and a thermionic detector.

The pyrazine; 2-methyl-; 2,3-dimethyl-; 2,5-dimethyl-; 2,6-dimethyl-; trimethyl- and tetramethyl pyrazine were detected in coffee beans. The principal compound was 2-methylpyrazine. The concentration of the latter pyrazine was more than 2000 μg/100 g of coffee beans depending on the time, the temperature of roasting and the origin of the beans. The correlation of the methylpyrazine quantity with the sensory analysis of the beans has shown the possibility of monitoring the roasting process of coffee beans.     

Influence of carboxypeptidases on free amino acid,peptide and methylpyrazine contents of under‐fermented cocoa beans

A study of the action of two carboxypeptidases on free amino acids, peptides and methylpyrazines in under‐fermented cocoa beans was carried out. Carboxypeptidase B from porcine pancreas and carboxypeptidase Y from baker's yeast were used separately for digestion. Hydrophobic free amino acids (alanine, valine, isoleucine, leucine, phenylalanine and tyrosine) were predominantly produced in samples digested with both carboxypeptidase B and Y. The peptide patterns of samples digested with both carboxypeptidases were similar to that of the control. The concentration of 2,3,5,6‐tetramethylpyrazine in samples with carboxypeptidase B addition was significantly higher than those of 2,5‐dimethyl‐ and 2,3,5‐trimethylpyrazine; the concentration of 2,3,5‐trimethylpyrazine was highest (1727.86 µg per 100 g) in the sample with carboxypeptidase B addition that had been incubated for 24 h. These findings indicate that carboxypeptidase B from porcine pancreas was more prominent in the formation of cocoa‐specific aroma. © 2002 Society of Chemical Industry     

Evaluation of the effect of processing on cocoa polyphenols: antiradical activity,anthocyanins and procyanidins profiling from raw beans to chocolate

The content and composition of anthocyanins and procyanidins in fermented cocoa beans (from different geographic origins: Ecuador, Cameroon, Ivory Coast, Ghana and Nigeria), roasted nibs, cocoa mass and chocolate were determined, beside the determination of the total antiradical capacity. Concerning geographic origin, cocoa beans and processed products from Ecuador showed the highest levels of anthocyanins, followed by Nigeria and Cameroon. Generally, as cocoa beans were further processed, the levels of anthocyanins and flavan‐3‐ols decreased. The largest observed losses of phenolics occurred during roasting. A progressive decreasing trend in polyphenol concentration was observed in the other processed samples as well. Despite the original content of polyphenols in raw cocoa beans, technological processes imply a significant impact on cocoa quality, confirming the need of specific optimisation to obtain high value chocolate.     

Aroma precursors and methylpyrazines in underfermented cocoa beans induced by endogenous carboxypeptidase

Cocoa-specific aroma precursors and methylpyrazines in underfermented cocoa beans obtained from fermentation induced by indigenous carboxypeptidase have been investigated. Fermentation conditions and cocoa bean components were analyzed during 0 to 3 d of fermentation. Underfermented cocoa beans were characterized as having hydrophilic peptides and free hydrophobic amino acids much higher than unfermented ones. These 2 key components of cocoa aroma precursors may be produced from the breakdown of proteins and polypeptides by endogenous carboxypeptidase during the fermentation process. The enzyme was activated during fermentation. Polypeptides of 47, 31, and 19 kDa were observed in the samples throughout the 3-d fermentation period; however, only the first 2 polypeptides were remarkably reduced during fermentation. Since the 1st day of fermentation, underfermented cocoa beans contained methylpyrazines, a dominant group of cocoa-specific aroma. This might be due to microbial activities during fermentation, observed through a decrease of pH value and an increase of temperature of cocoa beans. The concentration of tetramethylpyrazines was significantly increased during the 3 d of fermentation. This may increase the cocoa-specific flavor to the beans.     

Tocopherols in cocoa butter obtained from cocoa bean roasted in different forms and under various process parameters

Obtaining good quality chocolate strongly depends on raw material, i.e. cocoa beans. The processing of cocoa beans consists of some important steps, including fermentation, drying fermented beans and roasting. Traditionally roasting is performed on whole beans but currently, roasting crushed cocoa beans or cocoa liquor becomes more and more popular. Many biologically active compounds may be found in the cocoa beans, including tocopherols. This work investigates the influence of the constant or variable roasting process parameters (temperature, velocity and relative humidity of roasting air) on the tocopherol concentration in cocoa butter (CB) extracted from cocoa beans originating from Togo and roasted in two different forms, namely as whole and crushed beans. Whole cocoa beans were roasted to a 2% moisture content and crushed cocoa beans were firstly partially dried which further enables easier dehusking, then ground, dehusked and roasted until their humidity decreased to around 2%. Roasting resulted in lowering the content of individual tocopherols in analyzed material. The degree of degradation of tocopherols in CBs was different, depending on the form of roasted beans from which these CBs were extracted. Higher concentrations were determined in CBs extracted from beans roasted in the form of crushed samples comparing to CBs obtained from beans roasted as a whole. The study investigates different roasting parameters of crushed beans, none of which drastically lowered the concentration of tocopherols in extracted CBs. Their concentration in CBs extracted from whole beans was, on the other hand, influenced by roasting air parameters. In case of whole beans roasted under constant parameters, application of 150 °C proved to be more favorable than roasting at 135 °C, as well as application of “dry” air and 1 m/s roasting air velocity. Discussing the variable roasting process parameters, in case of applied roasting temperature it is more favorable to change it from 150 °C to 135 °C, than the other way round. Changing the relative humidity of roasting air from 5 to 0.3%, lower degradation of tocopherols in CB occurs when the process is conducted at 150 °C. It may be further concluded that a direct dependence between the velocity of roasting air varied during the process and the concentration of tocopherols in extracted CB may not be indicated. In conclusion, it is stated that the temperature of the air applied during the roasting process has the decisive influence on the tocopherol content in CBs extracted from cocoa beans subjected to the process.     

发酵和焙烤对可可豆多酚、黄酮和风味品质的影响

本实验采用高效液相色谱仪（high performance liquid chromatography,HPLC）、色差仪、电子鼻分别检测未发酵、发酵和焙烤海南可可豆的总酚含量、总黄酮含量、色度和可可豆风味差异。结果表明：未经焙烤的未发酵豆总酚（464.03 mg/10 g）和总黄酮（126.86 mg/10 g）含量明显高于发酵豆总酚（211.86 mg/10 g）和总黄酮（61.98 mg/10 g）含量。105～145 ℃焙烤30 min,未发酵豆总酚和总黄酮含量分别为419.5～129.8 mg/10 g和77.8～16.8 mg/10 g;发酵豆总酚和总黄酮含量分别为182.53～86.25 mg/10 g和34.7～7.0 mg/10 g。其中,125 ℃焙烤20～40 min,未发酵豆总酚和黄酮含量分别为353.74～289.45 mg/10 g和42.86～32.20 mg/10 g;发酵豆总酚和黄酮含量分别为152.08～123.55 mg/10 g和25.12～21.14 mg/10 g。在105～145 ℃的温度焙烤下没食子酸含量变化显著。可可豆色度值的范围：L*值集中在40.0～47.0之间,a*值集中在5.0～6.8之间,b*值集中在4.0～8.5之间。未发酵和发酵可可豆之间,以及不同温度焙烤可可豆之间的电子感官风味分析结果差异较大。     

Application of various methods for determination of the color of cocoa beans roasted under variable process parameters

Cocoa beans of Forastero variety from Togo were subjected to roasting under either constant or variable process parameters. The variable process parameters were roasting air flow rate, temperature and relative humidity. The color of roasted cocoa beans was determined by pigment extraction under various conditions followed by either their spectrophotometric assays or CIE L*a*b measurements. Also the Maillard compounds index and total polyphenols content correlated with progress of the browning of beans were determined. It was found that an increase in the roasting air relative humidity stimulated formation of brown pigments, while elevated temperature caused worsening of color parameters of roasted cocoa beans. The most suitable method of color characterization of roasted cocoa beans was found to be pigments extraction combined with either separation of their fractions or CIE L*a*b measurements. These assays revealed that cocoa beans roasted under variable roasting air flow rate were characterized by improved color parameters. The relatively simple and inexpensive CIE L*a*b measurements ensured fast analysis of color parameters, while total polyphenols in roasted cocoa beans were quickly estimated by using F–C reagent. Furthermore, quantification of melanoid pigments in roasted cocoa beans can be based on determination of the index of nonenzymatic browning products content, which is relatively simple and inexpensive.     

Ghanaian Cocoa Bean Fermentation Characterized by Spectroscopic and Chromatographic Methods and Chemometrics

Abstract: Export of cocoa beans is of great economic importance in Ghana and several other tropical countries. Raw cocoa has an astringent, unpleasant taste, and flavor, and has to be fermented, dried, and roasted to obtain the characteristic cocoa flavor and taste. In an attempt to obtain a deeper understanding of the changes in the cocoa beans during fermentation and investigate the possibility of future development of objective methods for assessing the degree of fermentation, a novel combination of methods including cut test, colorimetry, fluorescence spectroscopy, NIR spectroscopy, and GC-MS evaluated by chemometric methods was used to examine cocoa beans sampled at different durations of fermentation and samples representing fully fermented and dried beans from all cocoa growing regions of Ghana. Using colorimetry it was found that samples moved towards higher a* and b* values as fermentation progressed. Furthermore, the degree of fermentation could, in general, be well described by the spectroscopic methods used. In addition, it was possible to link analysis of volatile compounds with predictions of fermentation time. Fermented and dried cocoa beans from the Volta and the Western regions clustered separately in the score plots based on colorimetric, fluorescence, NIR, and GC-MS indicating regional differences in the composition of Ghanaian cocoa beans. The study demonstrates the potential of colorimetry and spectroscopic methods as valuable tools for determining the fermentation degree of cocoa beans. Using GC-MS it was possible to demonstrate the formation of several important aroma compounds such 2-phenylethyl acetate, propionic acid, and acetoin and the breakdown of others like diacetyl during fermentation. Practical Application: The present study demonstrates the potential of using colorimetry and spectroscopic methods as objective methods for determining cocoa bean quality along the processing chain. Development of objective methods for determining cocoa bean quality will be of great importance for quality insurance within the fields of cocoa processing and raw material control in chocolate producing companies.     

Quantification of 3-aminopropionamide in cocoa, coffee and cereal products

Based on recent results confirming 3-aminopropionamide (3-APA) as a very effective precursor of acrylamide in the absence of further “catalysts”, this compound was quantified for the first time in cocoa masses, cocoa beans, coffee and cereal products by LC–MS–MS after derivatisation with dansyl chloride. Cocoa masses contained >3000 μg/kg of 3-APA, but varied significantly in its concentration. For the quantification of acrylamide (AA) in cocoa and coffee, an improved isolation procedure using charcoal was developed. In various samples of unroasted and roasted cocoa beans, the concentrations of AA were by a factor of >5 lower than those of 3-APA, but the concentrations of 3-APA and AA were more closely correlated as compared to the concentrations of AA and Asparagine. Experiments on authentic cocoa beans from Ghana and Sulawesi indicated that the thermal generation of 3-APA during roasting was much more pronounced as compared to its biochemical formation. By administering fermented cocoa beans with [¹³C₄ ¹⁵N₂]-asparagine before roasting, 3-APA was confirmed as transient intermediate in AA formation during cocoa roasting. Among the cereal products analysed, in particular popcorn contained quite high amounts of 3-APA, which were also well correlated with the AA concentration. Contrary, in coffee products, 3-APA was always lower than AA.     

Antioxidant Properties of Cocoa Beans (Theobroma cacao L.): Influence of Cultivar and Roasting Conditions

Effects of various roasting conditions on antioxidant properties of five Theobroma cacao L. varieties were investigated. The cocoa beans were roasted at four different temperatures (110–150°C) and three different air humidities (0.3–5.0%). The raw cocoa beans were characterized by high antioxidant activities. The antioxidant properties of the roasted cocoa beans varied markedly among the analyzed cultivars and geographical regions and were affected by roasting conditions. Generally, cocoa beans of the cv. Forastero from Brazil exhibited higher total phenolic content, free radical scavenging activity, and metal chelating ability than samples of the other analyzed cocoa varieties. Roasting at 110°C caused negligible changes in total phenolics content and antioxidant activity of cocoa beans, while almost all samples tended to have lower antioxidant potential when roasting temperature increased. The air humidity used in roasting did not affect the total phenolics content and antioxidant activity for lowest roasting temperature (110°C). Moreover, the obtained results revealed that thermal processing at the higher temperatures and elevated air humidity resulted in the higher antioxidant capacities. It was also found that the ferrous ion chelating activity of cocoa beans increased with the roasting temperature (in the range from 110 to 150°C), with the exception of cv. Trinitario from Papua New Guinea. The data showed that roasting at lower temperatures with humid air are more favorable in terms of preserving the bioactivity of roasted cocoa beans.     

Analysis and stereodifferentiation of linalool in Theobroma cacao and cocoa products using enantioselective multidimensional gas chromatography

Contents and enantiomeric distributions of linalool (3,7-dimethyl-1,6-octadien-3-ol) were investigated in raw and roasted cocoa beans (seeds of Theobroma cacao) of defined origin as well as in commercial products, such as cocoa powders or chocolates. The stereodifferentiation of linalool was achieved by enantioselective multidimensional gas chromatography using heptakis (2,3-di-O-methyl-6-O-tert-butyldimethylsilyl)-β-cyclodextrin as chiral stationary phase. Simultaneous steam distillation–extraction at pH 7 allowed sample preparation without racemization of linalool. Cocoa beans contain linalool primarily as (S)-enantiomer. Model experiments and analyses of commercial products such as cocoa powders and chocolates revealed that technological procedures employed in the manufacturing of cocoa products do not result in significant changes of the original enantiomeric distribution of linalool.     

Improved Fermentation of Cocoa Beans with Enhanced Aroma Profiles

The cocoa (Theobroma cacao) fermentation was carried out using starter consortia of Saccharomyces cerevisiae, Lactobacillus plantarum, and Acetobacter aceti with 10% inoculum. The fermented beans were soaked with sterile water (1:3 wt/v) for the duration of 1, 3, and 5 h. The treated beans were dried to the moisture content of 7 ± 1%. The samples were then roasted, ground and subjected to steam distillation. The volatiles were extracted and identified using GC-MS. The samples soaked for 1 h scored 100% cut score along with 0.85% fermentation index. The volatile and non-volatiles were associated with beans treated in water after fermentation. Volatile compounds were classified as esters, alcohols, acids, aldehydes, ketones, pyrazines, hydroperoxides, and hydrocarbons. Most of the oxygenated aliphatic compounds and aromatics were potentially derived via an acetic acid and shikimic acid pathways, respectively. Untreated cocoa beans contained 12 sesquiterpene hydrocarbons potentially derived through the mevalonic acid pathway and contributed to harsh notes. Also, these metabolites were completely absent in the treated samples indicating depletion while soaking. Thus, the treated samples (1 h) had enhanced quality with high flavor notes with 8.5 rating on sensory evaluation. Thus, these soaking treatments with fermentation lead to improvement of flavor profiles which confers improved cocoa quality.     

Effects of incubation and polyphenol oxidase enrichment on colour, fermentation index, procyanidins and astringency of unfermented and partly fermented cocoa beans

Incubation of unfermented and partly fermented cocoa beans in acetate buffer, pH 5.5, at 45 °C increased yellowness, total colour differences and fermentation index value of the cocoa bean powders and decreased cocoa procyanidins (monomers to pentamers), and their astringency. Fermentation index and (–)‐epicatechin content, equivalent to those of fully fermented beans, were reached by unfermented beans after 4–8‐h incubation, but not by partly fermented beans even after 16 h. During incubation of partly fermented cocoa beans enriched with polyphenol oxidase, yellowness and fermentation index value were increased, whilst (–)‐epicatechin was decreased. Tyrosinase had a less significant effect in yellow colour formation, but showed a significant reduction of (–)‐epicatechin and increase in fermentation index compared with crude cocoa polyphenol oxidase. However, both enzymes have similar effects on procyanidin degradation and astringent taste reduction. Incubation of cocoa beans for 16 h increased the cut test score of unfermented and partly fermented beans by 50 and 30%, respectively.     

Physicochemical properties,antioxidant content,volatile organic compounds and sensory profile of cocoa beans fermented with yeast starter cultures

This study determined the influence of selected yeast starters on the antioxidant content, volatile organic compounds and sensory qualities of fermented cocoa beans. Cocoa beans fermented with Hanseniaspora thailandica (MH979675), Pichia kudriazevii (MH979681), or a mixture of both species in fermentation (20 kg) were highly reproducible based on the physicochemical parameters (temperature, pH of nibs, fermentation index and cut test). The usage of H. thailandica improved the total polyphenols content of cocoa beans with fermentation index around 1.24 and 1.33. The correlation analyses suggested that phenolics and flavonoids compounds may not be the main constituents that led to the antioxidant activities of the fermented samples. Principal Component Analysis showed distinct differences in VOC profiles between spontaneous and fermentation using yeast. Sweet and spicy notes were only found in sample fermented with yeast. The inoculation of yeast in fermentation influenced the sensorial and antioxidant properties of cocoa beans.     

Investigating the fermentation of cocoa by correlating denaturing gradient gel electrophoresis profiles and near infrared spectra

Raw cocoa has an astringent, unpleasant taste and flavour, and has to be fermented, dried and roasted in order to obtain the characteristic cocoa flavour and taste. During the fermentation microbial activity outside the cocoa beans induces biochemical and physical changes inside the beans. The process is complex involving activity of several different groups of microorganisms which bring about numerous biochemical and physical changes inside the beans. Due to the complexity of these processes no thorough investigations of the interactions between the microbial activities on the outside of the beans and the chemical processes inside the beans have been carried out previously. Recently it has been shown that Denaturing Gradient Gel Electrophoresis (DGGE) offers an efficient tool for monitoring the microbiological changes taking place during the fermentation of cocoa. Near Infrared (NIR) spectroscopy has previously been used to determine various components in cocoa beans, offering a rapid alternative compared to traditional analytical methods for obtaining knowledge about changes in the chemical composition of the cocoa beans during fermentation. During a number of cocoa fermentations bean samples were taken with 24 h intervals to be dried and analysed by NIR. Cocoa pulp samples taken simultaneously during the same fermentations have previously been characterised using DGGE [Nielsen, D.S., Teniola, O.D., Ban-Koffi, L., Owusu, M., Andersson, T., Holzapfel, W.H. (2007). The microbiology of Ghanaian cocoa fermentations analysed using culture dependent and culture-independent methods. International Journal of Food Microbiology 114, 168-186.]. Here we report the first study where microbiological changes during the fermentation determined using DGGE are correlated to changes inside the beans determined by NIR using multivariate data analysis. Following data pre-processing (baseline correction followed by Co-shift correction or Correlation Optimised Warping) the DGGE spectra were analysed using Principal Component Analysis (PCA). A clear grouping according to fermentation time was seen demonstrating the microbial succession taking place during the fermentation. Subsequently the DGGE spectra were correlated to the NIR spectra using Partial Least Squares regression models (PLS2). Correlations of 0.87 (bacterial derived DGGE spectra) and 0.81 (yeast derived DGGE spectra) were obtained indicating the relationship between the microbial activities in the pulp and the (bio)chemical changes inside the beans. By comparing the X-block loadings of the PLS2 models and the DGGE spectra it was possible to directly link several microbial species with changes in the NIR spectra and consequently also with changes inside the beans.     

Linalool content in roasted cocoa butter as a characteristic of several flavour grade cocoas

The role of linalool content in relation to flavour of roasted cocoa butter was investigated. Cocoa butters from flavour grade beans have relatively higher contents of linalool than those from basic grade cocoa varieties. The estimation of linalool content by simultaneous steam distillation extraction proved suitable for quality classification of roasted cocoa butters.     

Activation of remaining key enzymes in dried under-fermented cocoa beans and its effect on aroma precursor formation

Incubation-activation of remaining key enzymes in dried under-fermented cocoa beans and its effect on aroma precursor formation has been studied using defatted unfermented and partly fermented cocoa bean powders. Results of the study showed that aspartic endoprotease, carboxypeptidase and invertase were significantly inactivated during fermentation and drying, and the effect of fermentation was significantly lower than that of drying. The enzyme activities remaining in these beans were still sufficient to carry out enzymatic reaction during incubation. Peptide patterns, resulting from incubation of unfermented and partly fermented beans powders, were quite similar to the well-fermented patterns. Meanwhile, free amino acid concentrations of the unfermented beans were significantly increased during the first 4 h of incubation and then remained constant; however, with partly fermented beans, the formation continued and the hydrophobic and total free amino acid concentrations reached the value of well-fermented beans after 24 h of incubation. Reducing sugar concentrations of both unfermented and partly fermented cocoa beans could reach the level of well-fermented beans by incubation.     

Effect of drying time,bean depth and temperature on free amino acid,peptide‐N,sugar and pyrazine concentrations of Malaysian cocoa beans

The effect of drying time on free amino acid, peptide‐N, sugar and pyrazine concentrations as well as the influence of bean depth and temperature on these compounds during cocoa drying was studied. Drying time, bean depth and temperature significantly decreased the concentration of free amino acids, peptide‐N, total reducing sugars and sucrose and increased the concentration of trimethyl‐, tetramethylpyrazine and total pyrazines in cocoa beans. The best drying treatment was obtained at the combination of bean depth/drying temperature of 8.3 cm/40 °C. This was based on the fact that it produced significantly high concentrations of hydrophobic free amino acids, peptide‐N and total reducing sugars and significantly low concentration of trimethyl‐, tetramethylpyrazine and total pyrazines. Drying treatment of 1.5 cm/60 °C significantly produced the lowest concentration of free amino acids, peptide‐N and total reducing sugars and the highest concentration of pyrazines. © 1999 Society of Chemical Industry     

Alkylpyrazines and Other Volatiles in Cocoa Liquors at pH 5 to 8, by Selected Ion Flow Tube-Mass Spectrometry (SIFT-MS)

ABSTRACT:  Cocoa beans were alkalized before or after roasting and made into cocoa liquor before analyzing by SIFT-MS. In both alkalized-before-roasting and alkalized-after-roasting samples, there were significantly higher concentrations of alkylpyrazines for the samples with pH above 7 than pH below 7. At pH 8, the concentrations of 2,3-, 2,5-, and 2,6-dimethylpyrazine (DMP), 2,3,5-trimethylpyrazine (TrMP), 2,3,5,6-tetramethylpyrazine (TMP), and 2,3-diethyl-5-methylpyrazine (EMP) in the samples alkalized-before-roasting were higher than those in the samples alkalized-after-roasting. Volatiles increased under conditions that promoted the Maillard reaction. The partition coefficient was not significantly affected by pH from 5.2 to 8. The ratios of TrMP/DMP and DMP/TMP increased while the ratio of TMP/TrMP decreased as the pH increased. The concentrations of Strecker aldehydes and other volatiles followed a similar pattern as that of the alkylpyrazines. High pH favors the production of alkylpyrazines and Strecker aldehydes.
Practical Applications: Cocoa beans alkalized-before-roasting had higher levels of many important chocolate aroma volatiles than those alkalized-after-roasting. Thus, alkalizing before roasting should produce a stronger cocoa aroma. The higher the pH, the higher the concentrations of these important volatiles. There was little change in volatile concentration at acid pH. Above pH 7, volatile levels increased rapidly with increasing pH. To maintain the recommended ratio of tetramethylpyrazine to trimethylpyrazine, a longer roasting time or higher temperature is needed for cocoa liquor with a pH below 6.5, while a shorter roasting time or lower temperature is needed for cocoa liquor with a pH above 7.