LC-MS methods combination for identification and quantification of trans-sinapoylquinic acid regioisomers in green coffee.

The present study aims to both identify and quantify trans-sinapoylquinic acid (SiQA) regioisomers in green coffee by combined UHPLC-ESI-QqTOF-MS/MS and UHPLC-ESI-QqQ-MS/MS methods. Among the various mono-acyl chlorogenic acids found in green coffee, SiQA regioisomers are the least studied despite having been indicated as unique phytochemical markers of Coffea canephora (known as Robusta). The lack of commercially available authentic standards has been bypassed by resorting to the advantages offered by high-resolution LC-MS as far as the identification is concerned. SiQA regioisomers have been identified in several samples of Robusta and Coffea arabica (known as Arabica) commercial lots from different geographical origin and, for the first time, in different samples of coffee wild species (Coffea liberica and Coffea pseudozanguebariae). Quantification (total SiQA ranging from 3 to 5 mg/100 g) let to reconsider these chlorogenic acids as unique phytochemical markers of Robusta being present in the same quantity and distribution in C. liberica as well. Gardeniae Fructus samples (fruits of Gardenia jasminoides) have additionally been characterized as this matrix is recognized as one of the few naturally occurring SiQA sources. The SiQA regioisomer content (total SiQA about 80 mg/100 mg) fully supports the proposal to use this matrix as a surrogate standard for further studies.

16-O-Methylated diterpenes in green Coffea arabica: ultra-high-performance liquid chromatography-tandem mass spectrometry method optimization and validation.

Coffee diterpenes are the main constituents of the coffee oil unsaponifiable fraction. The three most important diterpenes are cafestol, kahweol, and 16-O-methylcafestol (16-OMC), and they are produced, except for cafestol, only by plants of the Coffea genus. Recently, in addition to these three major diterpenes, another 16-O-methylated diterpene (16-O-methylkahweol: 16-OMK) has been identified and quantified, for the first time, in Robusta coffee. For many years, 16-OMC has been considered present exclusively in Robusta, and so it has been reputed an excellent authenticity marker for the presence of Robusta in coffee products. For its quantification, nuclear magnetic resonance (NMR) has proved very useful when compared with other methods. Quite recently, the detection of very low levels of the two 16-O-methylated diterpenes (16-OMD) 16-OMC and 16-OMK in roasted Arabica was reported. This finding makes the use of NMR methods in 16-OMD quantification in Arabica coffee particularly challenging in view of both the trace amounts of 16-OMD and the impossibility to discriminate between 16-OMC and 16-OMK. The ultra-high performance liquid chromatography mass spectrometry (UHPLC-MS) method, already used to detect 16-OMC and 16-OMK in Arabica roasted coffee, is then more suitable for quantitative analyses. Up to now however, no quantification of coffee 16-OMD via ultra-high performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) has been carried out; this largely stimulated the present study. For the first time, a simple procedure for the quantitative detection of 16-OMD in Arabica coffee has been developed, and as far as 16-OMC is concerned, fully validated in terms of specificity, linearity, concentration range, limit of detection (LOD), limit of quantification (LOQ), and repeatability following the criteria specified in the EU Commission Decision 2002/675/EC. This method proved to be very specific and sensitive. In order to avoid the chemical complexity generated by the roasting process, the method was optimized and validated on several green Arabica samples from different geographical origins.

Distribution of p-coumaroylquinic acids in commercial Coffea spp. of different geographical origin and in other wild coffee species.

Quantitative analyses of mono-p-coumaroylquinic acids (pCoQAs) and total chlorogenic acids (CGAs) in green coffee commercial lots of C. arabica, C. canephora and C. liberica from different geographical origins and eight wild Coffea species were carried out. Among the commercial lots, pCoQAs average content of C. arabica (0.67 mg/g) is higher than that of C. canephora (0.40 mg/g) being C. liberica intermediate (0.58 mg/g). As far as the analyzed wild Coffea species is concerned, C. pseudozanguebariae is characterized by the lower pCoQAs content (0.12 mg/g) whereas C. sessiliflora is by far the richest source of pCoQAs (2.18 mg/g). Effect of the roasting process on the mono-p-coumaroylquinic acids profile was evaluated for the economically exploited species C. arabica and C. canephora. For the first time distribution of mono-p-coumaroylquinic acid isomers in wild coffee species by fast and accurate UHPLC-DAD analyses using authentic standards previously synthetized, is reported.

Aqueous extracts of walnut (Juglans regia L.) leaves: quantitative analyses of hydroxycinnamic and chlorogenic acids.

Identification of both hydroxycinnamic and chlorogenic acids present in aqueous extracts of walnut leaves (Juglans regia L.) were carried out by using, for the first time, standard compounds not commercially available for qualitative identification. In particular, in addition to caffeic, ferulic, p-coumaric and sinapic acids, cis and trans mono-caffeoylquinic, dicaffeoylquinic, mono-feruloylquinic and cis and trans mono-p-coumaroylquinic acid isomers were detected and quantified by Ultra High Pressure Liquid Chromatography and the seasonal variations of these secondary metabolites were investigated.

Preliminary Characterization of Monofloral Coffea spp. Honey: Correlation between Potential Biomarkers and Pollen Content.

To determine the botanical origin of Coffea honey, a new method using proton nuclear magnetic resonance ((1)H NMR) is proposed. Integration of the aromatic region of the NMR spectrum of Coffea honey diluted in deuterated water allowed us to simultaneously quantify caffeine, theobromine, and trigonelline, as well as other compounds. The amounts of the three markers listed are significantly higher than those previously reported for Citrus spp. honey: caffeine ranged from 15 to 98 mg/kg, theobromine from 25 to 160 mg/kg, and trigonelline from 23 to 86 mg/kg. The concurrent presence of these three substances is proposed as an indicator of the botanical origin of Coffea honey. Excellent correlation was found between these markers and the relative amounts of Coffea pollen measured in the same samples.

A new method for the preparative isolation of chlorogenic acid lactones from coffee and model roasts of 5-caffeoylquinic acid.

Chlorogenic acid lactones (CQL) are known to contribute to the bitter taste of roasted coffee. CQL might also have beneficial biological activities. Until now, there is a lack of pure standard compounds for quantification and biological testing. Using high-speed countercurrent chromatography, milligram amounts of lactones could be isolated. The structures of 3-O-caffeoyl-γ-quinide, 4-O-caffeoyl-muco-γ-quinide, and 5-O-caffeoyl-epi-δ-quinide were confirmed by 1D and 2D NMR spectroscopy including (13)C NMR data, which were previously not available from the literature. An UHPLC method was developed that enabled the separation of the lactones from roasted coffee in significantly shorter time than conventional HPLC.