ReviewAnalysis of the bioactive alkaloids tetrahydro-β-carboline and β-carboline in food
Introduction
Tetrahydro-β-carbolines (THβCs) and β-carbolines (βCs) are a group of naturally occurring alkaloids that possess a common tricyclic pyrido[3,4-b]indole ring structure. Compounds of this family have been reported in plant systems [1], cigarette smoke, roasted foodstuffs and mammalian tissues. In the last few years, we have shown the occurrence of THβCs in many commercial foods, and suggested that it may contribute to their ultimate presence in human biological tissues and fluids [2], [3], [4]. This also means that β-carbolines are naturally occurring substances in foods chemically produced during food production, processing and storage.
Research done in the last decades has pointed out the occurrence of THβCs and βCs under physiological conditions in biological tissues and fluids [5], [6], [7], [8], [9]. THβCs and βCs might function as neuromodulators via effects on monoamine oxidase, monoamine uptake and benzodiazepine receptors binding [5], [10]. Simultaneously, THβCs and βCs have been increasingly studied in relation to alcoholism [7], [11], [12], [13], [14]. Collins and co-workers have reported that N-methylated THβCs and βCs are endogenous neurotoxins [15], [16]. 1-Methyl-1,2,3,4-tetrahydro-β-carboline-3-carboxylic acid (MTCA), a tryptophan–acetaldehyde condensation THβC, is a precursor of mutagenic N-nitroso compounds [17], shows cytogenetic effects [18], and may cause neuronal cell death in vitro [19]. Furthermore, βCs produced during food cooking may exhibit comutagenic and genotoxic potential [20]. Taking all those reports together, we should conclude that a full delineation of the biological activity and possible toxicity of THβCs and βCs is desirable and still needed.
The analysis of THβCs and βCs in complex matrices involves their isolation and further identification prior to quantitative analysis. Given the relatively low concentration of these substances in most foods and biological samples, along with the very complex matrices involved, it is often necessary a previous clean-up for purification and trace enrichment. Analytical techniques with high efficiency and selectivity, mainly based on capillary gas chromatography (GC) and high-performance liquid chromatography (HPLC), are required to avoid coeluting and interfering peaks. Although UV absorption, and electrochemical detection systems are applicable, fluorescence and mass spectrometry are preferred for selective and sensitive detection. For identification purposes, mass spectrometry (MS) coupled with high-performance chromatographic techniques is the best on-line system due to its high selectivity and specificity. These aromatic heterocycles provide good mass spectra for detection following GC or HPLC.
Section snippets
Chemistry of tetrahydro-β-carbolines and β-carbolines
Those tricyclic compounds share a common name of 9H-pyrido[3,4-b]indole. Their chemical nomenclature, however, is sometimes confusing. It is adopted to call compounds with aromatic pyrido rings βCs and compounds with a reduced pyrido ring THβCs (see Fig. 1). βCs have a pyrido nitrogen in position 2 of the ring (N-2) compared to α- or γ-carbolines which have the nitrogen in positions 1 and 3, respectively. Those compounds usually contain several substituents both in the pyrido ring and/or the
Sample preparation and clean-up
Separation of THβCs and βCs by chromatographic means is generally preceded by clean-up procedures in order to isolate them from complex foods and biological matrices. Those include liquid–liquid extraction, and/or solid-phase extraction (SPE) by predominantly using ionic exchange and/or reversed-phase mechanisms. Very often, there is a chemical derivatization step prior to chromatographic analysis.
Chromatographic and spectral analysis
HPLC in combination with fluorometry and/or MS, and GC in combination with MS are the techniques predominantly used for separation and quantitation of THβCs and βCs. Some disadvantages of GC, such as the need of chemical derivatization to make volatile β-carbolines, possible artifact formation during derivatization or, even, difficulty in trace analysis are currently being solved by HPLC and mainly HPLC–MS of underivatized β-carbolines.
Tetrahydro-β-carbolines and β-carbolines in foods
As mentioned above, simple THβCs are alkaloids that occur naturally in foods as a chemical condensation between indoleamines and aldehydes or α-keto acids [2], [3], [4], [21], [28] (see Fig. 1). This reaction may occur during food production, processing and storage. In addition to the type of food involved, many chemical and technological factors, such as: amount of precursors available, pH, temperature, storage time, oxidants, antioxidants, preservatives, yeasts, processing conditions (e.g.,
Overview and conclusion
THβCs and BCs are naturally occurring alkaloids in foods produced by Pictet–Spengler chemical condensation. This suggests that foods are an exogenous source of possible THβCs and βCs ultimately present in human biological tissues and fluids. Since those compounds are biologically relevant, their analysis in foods should be recommended. Clean-up procedures include liquid–liquid extraction and/or SPE (using both C18 and cation-exchange mechanisms). Acylating reagents such as TFAA, HFBA, PFPA, MCF
Acknowledgements
The author is grateful to CICYT (Spanish Government), projects ALI970630 and ALI97-0396-CO2-O2 for financial support.
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