Occurrence of 3-chloropropane-1,2-diol and its esters in coffee

Fifteen coffee samples comprising green, roasted, decaffeinated and instant coffees were analysed for their content of free and bound 3-chloropropane-1,2-diol (3-MCPD). Green coffees contained only traces of the free 3-MCPD. 3-MCPD in roasted coffees was found at the level of 10.1–18.5 µg/kg. The highest 3-MCPD level of 18.5 µg/kg was found in one instant coffee sample and in coffees with very long time application during roasting. The final colour of the roasted coffee beans was directly linked to the 3-MCPD formed, the darker beans having the greatest concentration, and arabica coffees contained lower 3-MCPD levels than robusta coffees. The level of bound 3-MCPD varied between 6 µg/kg (soluble coffees) and 390 µg/kg (decaffeinated coffee) and exceeded the free 3-MCPD level 8–33 times. Arabica coffees contained higher levels of the bound 3-MCPD than robusta coffees. The recognised precursors of 3-MCPD (salt, glycerol, lipids) were also determined and their influence on the formation of the free and bound 3-MCPD was discussed.     

Investigation of ochratoxin a, B and citrinin contamination in various commercial foods

Ochratoxin A (OTA), ochratoxin B (OTB) and citrinin (CIT) in commercial foods were simultaneously determined and confirmed with high-performance liquid chromatography (HPLC) and liquid chromatography coupled with tandem mass spectrometry (LC/MS/MS). The samples examined were made up of cereal, fruit, coffee, and cacao products. The limits of quantification (S/N> or =10) of OTA, OTB and CIT were 0.1 microg/kg or less. Aflatoxins (AF), deoxynivalenol (DON) and fumonisins were also surveyed. Of 157 samples examined, 44 were contaminated with OTA at levels of 0.11 to 4.0 microg/kg. At least 2 positive samples were labeled as domestics. In most positive samples, the OTA level was low, less than 1 microg/kg. The highest incidence of OTA was observed in cacao powder (10/12), followed by instant coffee (5/7), cocoa (5/8) and raisin (6/13). OTB was found in fruit and cacao products containing relatively high levels of OTA. Co-occurrence of OTA, CIT and DON was found in cereal products, and co-occurrence of OTA and AF was found in cacao products. Approximately 30% of naturally contaminated OTA in roasted coffee bean moved into the extract solution when brewed with paper filter.     

Influence of roasting and brew preparation on the ochratoxin A content in coffee infusion

A study of the effect of coffee processing in the ochratoxin A (OTA) level has been carried out from the green beans to the drinking form. The analysis of OTA has been carried out by an in-house validated HPLC method with fluorescence detection. The limits of detection were 0.04 µg/kg for green and roasted coffee, and 0.01 µg/L for coffee brew. Thirty-six green coffee samples of different origin (Colombia, Costa Rica, Brazil, Vietnam, India and Uganda) were analysed. The highest concentrations of OTA were found in Vietnamese samples - Robusta species treated by dry processing - (range 0.64-8.05 µg/kg), that also showed the highest percentage of defective beans (7.6%). These contaminated samples were roasted in a process that controlled loss of weight and color, as in the industry. A mean reduction of 66.5% was obtained, but the reduction seems to be heterogeneous. Coffee brew was prepared by the three brewing processes more utilized in Europe: moka, auto-drip and espresso. A reduction of the OTA level has been attained, being greater when using a espresso coffee maker (49.8%) than when using auto-drip (14.5%) or moka brewing (32.1%). Therefore, the method of coffee brew preparation plays a key role in the final OTA human exposure.     

Ochratoxin A in Brazilian roasted and instant coffees

Thirty-four samples of roast and ground coffee, 14 samples of instant coffee and two samples of decaffeinated instant coffee were collected in markets and supermarkets in the city of Campinas, Brazil, and analysed for ochratoxin A using immunoaffinity columns for clean-up and HPLC with fluorescence detection for quantification. The limit of detection was 0.2 ng/g ochratoxin A. Twenty-three samples of ground and roast coffee were found to be contaminated with the toxin at levels ranging between 0.3 and 6.5 ng/g. The average concentration in all 34 samples was 0.9 ng/g. All samples of instant coffee contained ochratoxin A at levels ranging from 0.5 to 5.1 ng/g, with an average figure of 2.2 ng/g. Roast and ground coffee is the type of coffee most used by Brazilians for the preparation of the beverage. Considering that an average Brazilian adult takes five cups of coffee per day, which corresponds to 30 g of roast and ground coffee, the probable daily intake of ochratoxin A by a 70 kg adult would be 0.4 ng/kg bw, which is far below the current Provisional Tolerable Daily Intake of 14 ng/kg bw for ochratoxin A as set by the Codex Alimentarius. To study the transfer of ochratoxin A into coffee brew, the beverage was prepared by two methods: (a) the drip method and (b) the Brazilian country style method. No significant difference was observed between the two methods in terms of extraction of the toxin using five contaminated samples containing between 0.8 and 6.5 ng/g ochratoxin A. The drip method extracted 86 +/- 15% and the Brazilian country style 74 +/- 20% of the ochratoxin A initially present in the roast and ground coffee.     

Survey of Canadian retail coffees for ochratoxin A

One-hundred and one specimens of coffee were gathered from retail outlets across Canada and analysed for ochratoxin A. Seventy-one specimens were roasted beans or roasted ground coffee, and 30 were instant (or 'soluble') coffees. All samples were extracted with methanol-sodium bicarbonate. The extracts were cleaned up either by immunoaffinity column chromatography or by a combination of solid-phase extraction and immunoaffinity column chromatography. Ochratoxin A was quantified by liquid chromatography (LC) with fluorescence detection. The minimum quantifiable level was 0.1 ng g^-1. Ochratoxin A was present, above the minimum quantifiable level, in 42 (59%) of 71 beans and ground coffee and in 20 (67%) of 30 instant coffees. The mean ochratoxin A level in the positive samples of beans and ground coffee was 0.6 ng g^-1, and the mean level in the positive samples of instant coffee was 1.1 ng g^-1.     

Isolation, identification and toxigenic potential of ochratoxin A-producing Aspergillus species from coffee beans grown in two regions of Thailand

In 2006 and 2007, 32 Thai dried coffee bean samples (Coffea arabica) from two growing sites of Chiang Mai Province, and 32 Thai dried coffee bean samples (Coffea canephora var. robusta) from two growing sites of Chumphon Province, Thailand, were collected and assessed for the distribution of fungi with the potential to produce ochratoxin A (OTA). The overall percentage of fungal contamination in coffee was 98% and reduced to 60% after surface disinfection. There were remarkable ecological differences in the composition of ochratoxigenic species present in these two regions. Arabica coffee bean samples from the North had an average of 78% incidence of colonization with Aspergillus of section Circumdati with Aspergillus westerdijkiae and A. melleus as the predominant species. Aspergillus spp. of section Nigri were found in 75% of the samples whereas A. ochraceus was not detected. Robusta coffee beans from the South were 98-100% contaminated with predominantly A. carbonarius and A. niger. A. westerdijkiae was only found in one sample. The diversity of the fungal population was probably correlated with the geographical origin of the coffee, coffee cultivar, and processing method. Representative isolates of section Circumdati (52) and Nigri (82) were examined for their OTA production using HPLC with fluorescence detection. Aspergillus westerdijkiae (42 isolates out of 42), A. steynii (13/13), and A. carbonarius (35/35) in general produced large amounts of OTA, while one isolate of A. sclerotiorum produced intermediate amounts of OTA. 13% of the A. niger isolates produced OTA in intermediate amounts. OTA levels in coffee bean samples were analyzed using the Ridascreen OTA ELISA kits. Of the 64 coffee bean samples analyzed, 98% were contaminated with OTA in levels of <0.6-5.5 microg/kg (Arabica) and 1-27 microg/kg (Robusta). Presence of OTA in representative coffee samples was also confirmed by LC-MS/MS after ion-exchange purification.     

Development of a new method for the simultaneous determination of 21 mycotoxins in coffee beverages by liquid chromatography tandem mass spectrometry

A new method for the simultaneous detection of 21 mycotoxins (ochratoxin A, aflatoxin B₁, aflatoxin B_2, aflatoxin G₁, aflatoxin G₂, sterigmatocystin, nivalenol, deoxynivalenol, 3-acetyl deoxynivalenol, 15-acetyl deoxynivalenol, diacetoxyscirpenol, neosolaniol, HT-2 toxin, T-2 toxin, fumonisin B₁, fumonisin B₂, enniatin A, enniatin A₁, enniatin B, enniatin B₁, and beauvericin) in coffee beverages was internally validated. The method is based on liquid/liquid extraction with a mixture of ethyl acetate/formic acid (95:5 v/v) and detection using triple quadrupole (QqQ) and ion trap (IT) liquid chromatography tandem mass spectrometry. The limits of detection and quantification were 0.02 to 39.64 μg/kg, respectively, and the correlation coefficients were optimal for all mycotoxins (R² ≥ 0.992). The recovery values ranged from 72% to 97%. The developed method was demonstrated in six real samples of roasted and instant coffee, caffeinated and decaffeinated coffee, and coffee with sugar added. The analyses indicate the presence of the studied mycotoxins in coffee beverages at μg/kg concentrations. Ochratoxin A, a mycotoxin that is regulated in coffee, was detected in two samples under the maximum limit established by a European legislation (CE1881/2006).     

Simultaneous Detection of Multiple Adulterants in Ground Roasted Coffee by ATR-FTIR Spectroscopy and Data Fusion

This paper proposes a novel screening method for the simultaneous detection of four adulterants (spent coffee grounds, roasted coffee husks, roasted corn, and roasted barley) in ground roasted coffee using partial least squares discriminant analysis (PLS-DA) with mid-infrared spectroscopy. Two different acquisition modes (attenuated total reflectance, ATR, and diffuse reflectance, DR) are compared. Two recent chemometric approaches, hierarchical models (HM) and data fusion (DF), were employed in order to improve model performance. First level models provided discrimination between unadulterated and adulterated coffee samples, whereas second level models were able to identify the presence of each specific adulterant. The use of DF decreased the percentage of misclassified samples for the first level models from 19.6/14.7% (DR) and 7.5/14.5% (ATR) down to 2.5/4.5% considering the training/test sets. The percentage of misclassified samples in the second level models went as low as 0% (DF—spent coffee, training set). The proposed method is simple, fast, reliable for detecting adulteration in coffee samples, and capable of identifying these adulterants, even when in complex mixtures containing other adulterants.     

Fast simultaneous analysis of caffeine, trigonelline, nicotinic acid and sucrose in coffee by liquid chromatography–mass spectrometry

A rapid liquid chromatography–mass spectrometry method for the simultaneous quantification of caffeine, trigonelline, nicotinic acid and sucrose in coffee was developed and validated. The method involved extraction with hot water, clarification with basic lead acetate and membrane filtration, followed by chromatographic separation using a Spherisorb^® S5 ODS2, 5 μm chromatographic column and gradient elution with 0.3% aqueous formic acid/methanol at a flow rate of 0.2 mL/min. The electrospray ionization source was operated in the negative mode to generate sucrose ions and in the positive mode to generate caffeine, trigonelline and nicotinic acid ions. Ionization suppression of all analytes was found due to matrix effect. Calibrations curves prepared in green and roasted coffee extracts were linear with r² > 0.999. Roasted coffee was spiked and recoveries ranged from 93.0% to 105.1% for caffeine, from 85.2% to 116.2% for trigonelline, from 89.6% to 113.5% for nicotinic acid and from 94.1% to 109.7% for sucrose. Good repeatibilities (RSD < 5%) were found for all analytes in the matrix. The limit of detection (LOD), calculated on the basis of signal-to-noise ratios of 3:1, was 11.9, 36.4, 18.5 and 5.0 ng/mL for caffeine, trigonelline, nicotinic acid and sucrose, respectively. Analysis of 11 coffee samples (regular or decaffeinated green, ground roasted and instant) gave results in agreement with the literature. The method showed to be suitable for different types of coffee available in the market thus appearing as a fast and reliable alternative method to be used for routine coffee analysis.     

Occurrence of aflatoxins, ochratoxin A, and fumonisins in retail foods in Japan

We conducted a survey of aflatoxin B1, B2, G1, and G2, ochratoxin A, and fumonisin B1, B2, and B3 contamination in various foods on the retail market in Japan in 2004 and 2005. The mycotoxins were analyzed by high-performance liquid chromatography, liquid chromatography-mass spectrometry, or high-performance thin-layer chromatography. Aflatoxins were detected in 10 of 21 peanut butter samples; the highest concentration of aflatoxin B1 was 2.59 microg/kg. Aflatoxin contamination was not found in corn products, corn, peanuts, buckwheat flour, dried buckwheat noodles, rice, or sesame oil. Ochratoxin A was detected in oatmeal, wheat flour, rye, buckwheat flour, green coffee beans, roasted coffee beans, raisins, beer, and wine but not in rice or corn products. Ochratoxin A concentrations in contaminated samples were below 0.8 microg/kg. Fumonisins were detected in popcorn, frozen corn, corn flakes, and corn grits. The highest concentrations of fumonisins B1, B2, and B3 in these samples were 354.0, 94.0, and 64.0 microg/kg, respectively.     

ABTS radical scavenging capacity in green and roasted coffee extracts

The impact of two parameters (temperature and duration) on the radical scavenging capacity of individual compounds, and total extracts found in coffee was investigated. Phenolic coffee extracts of light (200 °C), medium (225 °C) and dark (235 °C) roasted coffees in a range of 0–30 min were analyzed by an on-line RP-HLPC-ABTS^•+ decolourization assay. This study revealed a general decrease of radical scavenging capacity related to native phenolic compounds. Processing coffee beans leads to generation of up to 10 new radical scavengers. The roasting process influences not only color and taste in coffees, but also the radical scavenging capacity of coffee as well. Phenolic content in roasted coffee and green coffee is very different. Six compounds identified as caffeoylquinic acids and dicaffeoylquinic acids, endowed with radical scavenging capacity were found in green coffee, whereas depending on the roasting process, roasted coffees can present up to 16 different radical scavengers. The compounds formed during the roast are most likely chlorogenic acids derivatives, of which 4 could be clearly identified as two feruloylquinic acids and two caffeoylquinides. In longer roasting durations, these molecules are subjected to auto-degradation, thus total radical scavenging capacity in coffee decline along with roasting (duration and temperature).     

基于UPLC-MS/MS探究烘焙程度对咖啡豆中有机酸含量影响

为探究烘焙工艺对咖啡豆口感的影响,本文研究了烘焙程度对咖啡豆中有机酸含量的影响。采用新建立超高效液质联用法（UPLC-MS/MS）测定不同烘焙条件下7种有机酸的含量。结果表明:7种有机酸化合物在0.5～20.0 mg/kg范围内线性关系良好,决定系数均大于0.990。7种有机酸的加标回收率在81.9%～104.7%,RSD为0.53%～6.64%。洪都拉斯咖啡豆样品检测结果显示随烘焙程度增加,苹果酸和柠檬酸含量下降,富马酸含量先升后降,琥珀酸、酒石酸、莽草酸和奎尼酸含量上升。轻度烘焙下苹果酸与柠檬酸总量最高,达到1201.5 mg/kg,酸度最佳;奎尼酸含量最少,为1363.7 mg/kg,涩度最低。肯尼亚、印尼、巴西、萨尔瓦多咖啡豆中有机酸在相同烘焙条件下表现相似。     

Supercritical CO2 decaffeination of unroasted coffee beans produces melanoidins with distinct NF-κB inhibitory activity

The supercritical CO(2)-decaffeination process causes unroasted coffee beans to turn brown. Therefore, we suspected that the decaffeinated beans contained melanoidins. Decaffeinated unroasted coffee extract absorbed light at 405 nm with a specific extinction coefficient, K(mix 405 nm), of 0.02. Membrane dialysis (molecular weight cut-off, 12 to 14 kDa) increased the K(mix 405 nm) value 15 fold. Gel filtration chromatography showed that the high-MW fraction (MW > 12 kDa) had an elution profile closer to that of melanoidins of medium-roast coffee than to the corresponding fraction of unroasted coffee, indicating the presence of melanoidins in decaffeinated unroasted beans. Using murine myoblast C2C12 cells with a stably transfected nuclear factor-κB (NF-κB) luciferase reporter gene, we found that the high-MW fraction of decaffeinated unroasted beans had an NF-κB inhibitory activity of IC(50) = 499 μg/mL, more potent than that of regular-roast coffee (IC(50) = 766 μg/mL). Our results indicate that melanoidins form during the supercritical CO(2)-decaffeination process and possess biological properties distinct from those formed during the regular roasting process. PRACTICAL APPLICATION: We discovered the roasting effect of decaffeination process, reporting the discovery of melanoidins in green (unroasted) decaf coffee beans. Our results indicated that melanoidins form during the supercritical CO2-decaffeination process and possess biological properties distinct from those formed during the regular roasting process. Our results offer new insights into the formation of bioactive coffee components during coffee decaffeination process.     

云南不同地区烘焙咖啡豆主要成分分析及类黑精组成成分

以云南3个咖啡主产区(普洱、怒江、德宏)的生咖啡豆为原料,通过不同程度(轻度、中度、重度)烘焙,对其主要成分进行分析,并提取类黑精,采用热裂解-气相色谱-质谱(pyrolysis-gas chromatography-mass spectrometry,Py-GC-MS)联用技术分析咖啡类黑精的化学组成。结果显示:随着烘焙程度加深,咖啡豆的蛋白质和粗脂肪含量增加,总氨基酸含量降低,其中蛋白质含量最高为重度烘焙后的普洱咖啡豆(16.3 g/100 g),氨基酸含量最高为普洱生咖啡豆(9.41%),粗脂肪含量最高为中度烘焙后的德宏咖啡豆(13.85 g/100 g),矿物质元素中含量较高的为K、Mg、P、Ca,普洱咖啡豆经重度烘焙后K含量最高(2.2 g/100 g)。Py-GC-MS分析结果表明:不同产地、不同烘焙程度咖啡豆的类黑精组成差异明显,但也存在共性特征,咖啡因和酸类相对含量最高,其次是胺类、酯类、酚类、吡咯类、呋喃类、吡啶类、醛类、醇类、酮类等。     

Influence of torrefacto roast on antioxidant and pro-oxidant activity of coffee

The addition of sugar at the end of the torrefacto roasting process may influence the antioxidant and pro-oxidant properties of coffee because sugar is one of the main precursors the Maillard reaction. The aim of the work was to study and to compare the antioxidant and pro-oxidant properties of some commercial roasted coffees which are selected to represent conventional roasted arabica coffee and arabica/robusta blends, and torrefacto roasted blends. Higher antioxidant activity was observed in Colombian coffees than in conventional roasted coffee blends. On the other hand, when the percentage of torrefacto coffee was increased, an increase of the antioxidant activity and a slight tendency to decrease the pro-oxidant activity were observed. Moreover, principal component analysis allowed separation of: (a) brands by PC1 (46.9%), characterised by colour parameters defined by roasting degree and (b) torrefacto roasted blends by PC2 (33.7%), characterised by antioxidant/pro-oxidant activity.     

烘焙度对冷萃咖啡理化指标与风味成分的影响

以亚洲咖啡豆为研究对象,分别选取浅、中、深3 种烘焙度的中国云南和印度尼西亚苏门答腊产的卡蒂姆种咖啡豆,比较分析冷萃与热萃方式对萃取浓度、萃取率、可滴定酸、总酚、总糖、咖啡因、葫芦巴碱、绿原酸、抗氧化活性与挥发性成分的差异,并进行主成分分析,从而探究烘焙度对冷萃咖啡理化指标与风味成分的影响规律。结果表明,随着烘焙度增加,冷萃咖啡的萃取浓度、萃取率均显著上升,可滴定酸、总酚、葫芦巴碱、绿原酸、抗氧化活性均显著下降（P＜0.05）。冷萃咖啡较热萃咖啡拥有更高的萃取浓度、萃取率与总糖含量（P＜0.05）,而可滴定酸、总酚含量、抗氧化活性较热萃显著偏低（P＜0.05）。经顶空固相微萃取-气相色谱-质谱检测分析发现,浅烘咖啡豆萃取液中的挥发性成分含量显著低于烘焙度高的咖啡萃取液,深烘咖啡豆萃取液中挥发性成分种类与总含量最多。进一步通过主成分分析能较好区分冷萃和热萃咖啡,两者挥发性成分贡献率具有较大差异。2-丁酮、2-丁烯醛等花香类物质对浅烘冷萃咖啡贡献率更高,而2-甲基吡嗪、糠醇等呈现烘焙坚果类香气物质对浅烘热萃咖啡贡献率更高;2,6-二乙基吡嗪、川芎嗪等烘焙坚果类香气物质对中烘冷萃和热萃咖啡具有较高的贡献率;2-乙烯基呋喃、甲基糠硫醇、2,5-二乙基吡嗪、糠基甲基硫醚等物质对深烘冷萃咖啡有较高贡献率,二甲基二硫、对甲酚、1-甲基吡咯等物质对深烘热萃咖啡贡献率更高。相对于热萃咖啡,烘焙度对冷萃咖啡抗氧化能力与挥发性成分的影响更大。     

Effect of operating conditions on ochratoxin A extraction from roasted coffee

Operating conditions affect ochratoxin A (OTA) extraction from roasted coffee. The OTA content found in the beverage can thus be greater than that found in the roasted coffee used to prepare it. Three extraction parameters were studied for roasted coffee: type of extraction solvent (alkaline, neutral, acid), temperature (ambient temperature/23°C, 60°C and 85°C), and extraction time (5, 20, 30, 40, 50, 60 and 80 min). The alkaline solvent used in the method recommended by the European Union extracted OTA better, but a maximum content was obtained at 60°C after 50 min. At least a 100% improvement in extraction was obtained when compared with the European Union usual quantification method that is carried out at ambient temperature. It turned out that the OTA extraction parameters for roasted coffee, as defined by that method, were not optimum and needed to be modified. These results were verified in double-extraction experiments showing that OTA is not completely extracted by this method. Confirmation was obtained by comparison of extraction methods on several commercial samples of roasted coffee.     

Species,roasting degree and decaffeination influence the antibacterial activity of coffee against Streptococcus mutans

Coffee beverage has been associated with antibacterial activity against Streptococcus mutans, a cariogenic bacterium. This study aimed at identifying natural compounds in coffee that contribute to such activity and investigate the influence of species, roasting and decaffeination on it. Coffee chemical compounds and aqueous extracts of green and roasted regular and decaffeinated Coffea arabica and Coffea canephora beans were tested. MIC, biofilm inhibition and biofilm reduction results were correlated with the concentration of coffee compounds in the extracts. 5-Caffeoylquinic acid, trigonelline and caffeic acid solutions showed bacteriostatic activity (MIC = 0.8 mg/mL). Lighter and regular extracts showed higher inhibitory activity than darker and decaffeinated extracts, with an inverse correlation between bacterial colony-forming units and roasting degree. Only regular C. canephora extracts showed biofilm formation inhibition. The joint effect of chlorogenic acids, trigonelline and caffeine or other compounds removed by decaffeination seems to be one of the causes for coffee antibacterial activity against S. mutans.     

From the green bean to the cup of coffee: investigating coffee roasting by on-line monitoring of volatiles

Volatile organic compounds (VOCs), emitted from green coffee beans, during coffee roasting and from a cup of coffee, were all analysed by proton-transfer-reaction mass spectrometry. Firstly, the headspace (HS) of green beans was investigated. Alcohols dominate the HS, but aldehydes, hydrocarbons and organic acids were also abundant. Secondly, we roasted coffee under two different conditions and monitored on-line the VOCs emitted during the process. In a first roasting series, a batch of beans was roasted. After an initial drying phase, dominated by evaporation of water and methanol, the HS concentrations of VOCs such as acetic acid, acetaldehyde, pyridine and methylbutanal rapidly increased and went through a maximum at medium roast level. In a second series, just six beans were roasted. We observed sporadic bursts of some volatiles (furans, butanal, 2,3-pentanedione), coinciding with popping sounds. Other VOCs showed smooth time-intensity profiles (pyridine, pyrazine). These experiments gave a real-time insight into the complex processes taking place during roasting. Finally, the HS of coffee extracts, prepared from beans roasted to different roast levels, were analysed. Most VOCs showed a maximum concentration at medium roast level (e.g. pentanedione, furfural, 5-methyl furfural), while others showed a gradual increase (e.g. pyrrol) or decrease (e.g. methanol).     

Caffeine Content of Retail Market Coffee in Portugal

How much caffeine does one ingest when drinking a simple cup of coffee in Portugal? The study presented herein tried to answer this question through the assessment of caffeine content of commercially available espresso coffee samples, both caffeinated and decaffeinated, using a high-performance liquid chromatography assay. Caffeine was rapidly separated from the sample matrix using a RP-18 column (250?×?4 mm i.d., 5 μm). The flow rate was 1.0 mL/min and the mobile phase consisted of water acidified with 5% of orthophosphoric acid/methanol (35:65, v/v). Caffeine was detected directly at 273 nm. The assay was validated for linearity, lower limit of quantification and limit of detection, precision, accuracy, and stability. Seventeen different brands of caffeinated coffee and six of decaffeinated coffee were analyzed. As for capsule coffee, eight caffeinated and two decaffeinated blends were analyzed. The caffeine content of caffeinated coffee varied from 53.8?±?5.9 to 141.3?±?5.3 mg/cup, whereas for caffeinated capsule coffee caffeine concentrations ranged from 45.0?±?5.3 to 60.8?±?6.2 mg/cup. As for decaffeinated coffee, caffeine concentrations ranged from 0.96?±?0.04 to 3.9?±?0.1 mg/cup and for decaffeinated capsule coffee from 0.93?±?0.04 to 1.2?±?0.1 mg/cup.