Biosynthesis of chlorogenic acids in growing and ripening fruits of Coffea arabica and Coffea canephora plants.

Chlorogenic acids are major secondary metabolites found in coffee seeds. The accumulation of chlorogenic acids and free quinic acids was studied in Coffea arabica cv. Tall Mokka and Coffea canephora seeds. Growth stages are specified from I to V, corresponding to rapid expansion and pericarp growth (I), endosperm formation (II), mature (green) (III), ripening (pink) (IV), and fully ripened (red) (V) stages. We detected monocaffeoylquinic acids (3CQA, 4CQA and 5CQA), dicaffeoylquinic acids (3,4diCQA, 3,5diCQA and 4,5diCQA) and a monoferuloylquinic acid (5FQA) in whole fruits (stage I), pericarps and seeds. The most abundant chlorogenic acid was 5CQA, which comprised 50-60% of the total of C. arabica and 45-50% of C. canephora seeds. The content of dicaffeoylquinic acid, mainly 3,5-diCQA, was high in C. canephora. A high content of 5FQA was found in seeds of stages III to V, especially in C. canephora. Total chlorogenic acids were accumulated up to 14 mg per fruit in C. arabica and 17 mg in C. canephora, respectively. In contrast, free quinic acid varied from 0.4-2.0 mg (C. arabica) and 0.2-4.0 mg (C. canephora) per fruit during growth. High biosynthetic activity of 5CQA, which was estimated via the incorporation of [U-14C]phenylalanine into chlorogenic acids, was found in young fruits (perisperm and pericarp) in stage I, and in developing seeds (endosperm) in stages II and III. The biosynthetic activity of chlorogenic acids was clearly reduced in ripening and ripe seeds, especially in C canephora. Transcripts of PAL1, C3'H and CCoAMT, three genes related to the chlorogenic acid biosynthesis, were detected in every stagè of growth, although the amounts were significantly less in stage V. Of these genes, CCoAMT, a gene for FQA biosynthesis, was expressed more weakly in stage I. The transcript level of CCoAMT was higher in seeds than in pericarp, but the reverse was found in PAL1. The pattern of expression of genes for the CQA and FQA synthesis is roughly related to the estimated biosynthetic activity, and to the accumulation pattern of chlorogenic acids.

Determination of kavalactones in dried kava (Piper methysticum) powder using near-infrared reflectance spectroscopy and partial least-squares regression.

Kava (Piper methysticum Forst F.), or àwa in the Hawaiian language, has been used for thousands of years by the people of the South Pacific Islands, in particular Fiji, Vanuatu, Tonga, and Samoa, for social and ceremonial occasions. Kava has the unique ability to promote a state of relaxation without the loss of mental alertness. Kava recently became part of the herbal pharmacopoeia throughout the United States and Europe because of its anxiolytic properties. The active compounds are collectively called kavalactones (or kava pyrones). The need for a less time-consuming and costly method to determine the concentration of kavalactones in dried kava is urgent. The combination of near-infrared reflectance spectroscopy (NIRS) and partial least-squares (PLS) methods has been found to be a convenient, versatile, and rapid analytical tool for determination of kavalactones in dried kava powder. Calibration equations were developed based on the analyses of 110 samples with variable physical and chemical properties collected over time from Hawaii kava growers and validated by analyses of a set of 12 samples with unknown kavalactones concentration. All six major kavalactones and the total kavalactones were measured using NIRS with accuracy acceptable for commercial use. The NIRS measurements are reproducible and have a repeatability on a par with HPLC methods.

Profiles of phenolic compounds and purine alkaloids during the development of seeds of Theobroma cacao cv. Trinitario.

Changes occurring in phenolic compounds and purine alkaloids, during the growth of seeds of cacao (Theobroma cacao) cv. Trinitario, were investigated using HPLC-MS/MS. Extracts of seeds with a fresh weight of 125, 700, 1550, and 2050 mg (stages 1-4, respectively) were analyzed. The phenolic compounds present in highest concentrations in developing and mature seeds (stages 3 and 4) were flavonols and flavan-3-ols. Flavan-3-ols existed as monomers of epicatechin and catechin and as procyanidins. Type B procyanidins were major components and varied from dimers to pentadecamer. Two anthocyanins, cyanidin-3-O-arabinoside and cyanidin-3-O-galactoside, along with the N-phenylpropernoyl-l-amino acids, N-caffeoyl-l-aspartate, N-coumaroyl-l-aspartate, N-coumaroyl-3-hydroxytyrosine (clovamide), and N-coumaroyltyrosine (deoxyclovamide), and the purine alkaloids theobromine and caffeine, were present in stage 3 and 4 seeds. Other purine alkaloids, such as theophylline and additional methylxanthines, did not occur in detectable quantities. Flavan-3-ols were the only components to accumulate in detectable quantities in young seeds at developmental stages 1 and 2.

Expression of the grapevine stilbene synthase gene VST1 in papaya provides increased resistance against diseases caused by Phytophthora palmivora.

The phytoalexin resveratrol (trans-3,5,4'-trihydroxy-stilbene), a natural component of resistance to fungal diseases in many plants, is synthesized by the enzyme trihydroxystilbene synthase (stilbene synthase, EC 2.3.1.95), which appears to be deficient or lacking in susceptible plants. Earlier workers isolated a stilbene synthase gene (Vst1) from grapevine (Vitis vinifera L.), which has subsequently been introduced as a transgene into a range of species to increase resistance of hosts to pathogens to which they were originally susceptible. Papaya (Carica papaya L.) is susceptible to a variety of fungal diseases, including root, stem, and fruit rot caused by the pathogen Phytophthora palmivora. Since resveratrol at 1.0 mM inhibited mycelium growth of P. palmivora in vitro, we hypothesized that papaya resistance to this pathogen might be increased by transformation with the grapevine stilbene synthase construct pVst1, containing the Vst1 gene and its pathogen-inducible promoter. Multiple transformed lines were produced, clonally propagated, and evaluated with a leaf disk bioassay and whole plant response to inoculation with P. palmivora. RNA transcripts of stilbene synthase and resveratrol glycoside were induced in plant lines transformed with the grapevine pVst1 construct shortly after pathogen inoculation, and the transformed papaya lines exhibited increased resistance to P. palmivora. The immature transformed plants appear normal and will be advanced to field trials to evaluate their utility.