Bioconversion of rebaudioside I from rebaudioside A.
ABSTRACT: To supply the increasing demand of natural high potency sweeteners to reduce the calories in food and beverages, we have looked to steviol glycosides. In this work we report the bioconversion of rebaudioside A to rebaudioside I using a glucosyltransferase enzyme. This bioconversion reaction adds one sugar unit with a 1?3 linkage. We utilized 1D and 2D NMR spectroscopy (1H, 13C, COSY, HSQC-DEPT, HMBC, 1D TOCSY and NOESY) and mass spectral data to fully characterize rebaudioside I.
Project description:A minor product, rebaudioside M2 (2), from the bioconversion reaction of rebaudioside A (4) to rebaudioside D (3), was isolated and the complete structure of the novel steviol glycoside was determined. Rebaudioside M2 (2) is considered an isomer of rebaudioside M (1) and contains a relatively rare 1?6 sugar linkage. It was isolated and characterized with NMR (1H, 13C, COSY, HSQC-DEPT, HMBC, 1D-TOCSY, and NOESY) and mass spectral data. Additionally, we emphasize the importance of 1D and 2D NMR techniques when identifying complex steviol glycosides. Numerous NMR spectroscopy studies of rebaudioside M (1), rebaudioside D (3), and mixture of 1 and 3 led to the discovery that SG17 which was previously reported in literature, is a mixture of rebaudioside D (3), rebaudioside M (1), and possibly other related steviol glycosides.
Project description:Following our interest in new diterpene glycosides with better taste profiles than that of Rebaudioside M, we have recently isolated and characterized Rebaudioside IX-a novel steviol glycoside-from a commercially-supplied extract of Stevia rebaudiana Bertoni. This molecule contains a hexasaccharide group attached at C-13 of the central diterpene core, and contains three additional glucose units when compared with Rebaudioside M. Here we report the complete structure elucidation-based on extensive Nuclear Magnetic Resonance (NMR) analysis (1H, 13C, Correlation Spectroscopy (COSY), Heteronuclear Single Quantum Coherence-Distortionless Enhancement Polarization Transfer (HSQC-DEPT), Heteronuclear Multiple Bond Correlation (HMBC), 1D Total Correlation Spectroscopy (TOCSY), Nuclear Overhauser Effect Spectroscopy (NOESY)) and mass spectral data-of this novel diterpene glycoside with nine sugar moieties and containing a relatively rare 1?6 ?-linked glycoside. A steviol glycoside bearing nine glucose units is unprecedented in the literature, and could have an impact on the natural sweetener catalog.
Project description:Glucosylation of the 21-hydroxyl group of glucocorticoid changes its solubility into hydrophilicity from hydrophobicity and, as with glucocorticoid glucuronides as a moving object in vivo, it is conceivable that it exhibits the same behavior. Therefore, glucosylation to the 21-hydroxyl group while maintaining the 11<i>?</i>-hydroxyl group is particularly important, and glucosylation of corticosterone was confirmed by high-resolution mass spectrometry and 1D (¹H and <sup>13</sup>C) and 2D (COSY, ROESY, HSQC-DEPT and HMBC) NMR. Moreover, the difference in bioactivity between corticosterone and corticosterone 21-glucoside was investigated in vitro. Corticosterone 21-glucoside showed greater neuroprotective effects against H?O?-induced cell death and reactive oxygen species (ROS) compared with corticosterone. These results for the first time demonstrate that bioconversion of corticosterone through the region-selective glucosylation of a novel compound can present structural potential for developing new neuroprotective agents.
Project description:The worldwide heparin contamination crisis in 2008 led health authorities to take fundamental steps to better control heparin manufacture, including implementing appropriate analytical and bio-analytical methods to ensure production and release of high quality heparin sodium product. Consequently, there is an increased interest in the identification and structural elucidation of unusually modified structures that may be present in heparin. Our study focuses on the structural elucidation of species that give rise to a signal observed at 2.10 ppm in the N-acetyl region of the 1H NMR spectrum of some pharmaceutical grade heparin preparations. Structural elucidation experiments were carried out using homonuclear (COSY, TOSCY and NOESY) and heteronuclear (HSQC, HSQC-DEPT, HMQC-COSY, HSQC-TOCSY, and HMBC) 2D NMR spectroscopy on both heparin as well as heparin-like model compounds. Our results identify a novel type of oxidative modification of the heparin chain that results from a specific step in the manufacturing process used to prepare heparin.
Project description:A simple and robust solvent suppression technique that enables acquisition of high-quality 1D 1 H nuclear magnetic resonance (NMR) spectra of alcoholic beverages on cryoprobe instruments was developed and applied to acquire NMR spectra of Scotch Whisky. The method uses 3 channels to suppress signals of water and ethanol, including those of 13 C satellites of ethanol. It is executed in automation allowing high throughput investigations of alcoholic beverages. On the basis of the well-established 1D nuclear Overhauser spectroscopy (NOESY) solvent suppression technique, this method suppresses the solvent at the beginning of the pulse sequence, producing pure phase signals minimally affected by the relaxation. The developed solvent suppression procedure was integrated into several homocorrelated and heterocorrelated 2D NMR experiments, including 2D correlation spectroscopy (COSY), 2D total correlation spectroscopy (TOCSY), 2D band-selective TOCSY, 2D J-resolved spectroscopy, 2D 1 H, 13 C heteronuclear single-quantum correlation spectroscopy (HSQC), 2D 1 H, 13 C HSQC-TOCSY, and 2D 1 H, 13 C heteronuclear multiple-bond correlation spectroscopy (HMBC). A 1D chemical-shift-selective TOCSY experiments was also modified. The wealth of information obtained by these experiments will assist in NMR structure elucidation of Scotch Whisky congeners and generally the composition of alcoholic beverages at the molecular level.
Project description:Gaiadandendron punctatum G.Don. (violeta de campo) is a plant used in traditional medicine by the Saraguro people, an ancient indigenous group that lives in southern Ecuador. From samples collected in the region, six glycoside flavonoids, five with quercetin and one with kaempferol as aglycon, were isolated and characterized from hydroalcoholic extracts of leaves and flowers. Rutin (2) was found in flowers and leaves, nicotiflorin (1) was found in flowers, artabotryside A (3) was found in leaves, and three novel quercetin flavonoid glycosides were isolated, elucidated, and characterized via 1D and 2D NMR experiments (1H, 13C, COSY, DEPT, HMBC, HSQC, TOCSY, NOESY, ROESY), acid hydrolysis-derivatization-GC-MS analysis, HPLC-MS, IR, UV, and optical rotation. The new quercetin flavonoid glycosides were named hecpatrin (4) (isolated from leaves), gaiadendrin (5) (isolated from leaves), and puchikrin (6) (isolated from flowers). The hydroalcoholic extracts of the leaves presented antimicrobial activity against Micrococcus luteus, Staphylococcus aureus, and Enterococcus faecalis and the hydroalcoholic extract of the flowers was active against Micrococcus luteus. However, glycoside flavonoids presented scarce antimicrobial activity against bacteria. Hydroalcoholic extracts from leaves and flowers and their secondary metabolites showed inhibition against the ?-glucosidase enzyme at different concentrations. Rutin, gaiadendrin, and nicotiflorin showed competitive ?-glucosidase inhibition, while hecpatrin presented non-competitive inhibition.
Project description:The Melastomataceae family, the seventh largest flowering plants, has been studied in several fronts of natural product chemistry, including terpenoids, simple phenolics, flavonoids, quinones, lignans and their glycosides, as well as a vast range of tannins or polyphenols. This review concerns the phenolic and polyphenolic metabolites described in the literature for several genera of this family, the mode of isolation and purification, and the structure elucidation of these new natural products that has been achieved by extensive spectral analyses, including ESI-MS, ¹H-, (13)C-NMR spectra and two-dimensional experiments, COSY, TOCSY, J-resolved, NOESY, HMQC, DEPT, and HMBC, as well as chemical and enzymatic degradations and the chemotaxonomic meaning. Finally, a general biogenetic pathway map for ellagitannins is proposed on the bases of the most plausible free radical C-O oxidative coupling.
Project description:The seven pyridine alkaloids 1-7, the flavonoid acacetin (8), and L-proline anhydride (9) have been isolated from the aerial parts of the Mongolian medicinal plant Caryopteris mongolica Bunge. The structures of the natural products 1-9 have been assigned by MS, as well as IR, 1D NMR (<sup>1</sup>H, <sup>13</sup>C, DEPT), and 2D NMR (COSY, HSQC, HMBC, NOESY) spectroscopic methods. The compounds 2 and 4-7 represent new chemical structures. Acacetin (8) and L-proline anhydride (9) have been obtained from C. mongolica for the first time.
Project description:Zero-calorie high-intensity sweeteners from natural sources perform very well in the market place. This has encouraged food scientists to continue the effort to search for novel natural ingredients to satisfy consumer demand. Rebaudioside C (reb C) is the third most prevalent steviol glycoside in the leaves of the <i>Stevia rebaudiana</i> Bertoni plant, but has limited applications in food and beverage products due to its low sweetness and high lingering bitterness compared to other major steviol glycosides, such as rebaudioside A (reb A). Here we present a new enzyme modification strategy to improve the taste profile of reb C by using Cargill's propriety enzyme and sucrose as a glucose donor. A novel ?-1?6-glucosyl reb C derivative was produced and its structure was elucidated by mass spectrometry and NMR spectroscopy. Sensory analysis demonstrated that this new reb C derivative has improved sweetness, reduced bitterness, and enhanced solubility in water.
Project description:Ten new bis-spirolabdane diterpenoids, leonepetaefolins A-E (1, 3, 5, 7, 9) and 15-epi-leonepetaefolins A-E (2, 4, 6, 8, 10), together with eight known labdane diterpenoids (11-18) as well as two known flavonoids, apigenin and cirsiliol, were isolated from the leaves of Leonotis nepetaefolia. The structures of the new compounds were determined on the basis of 1D- and 2D-NMR experiments including (1)H, (13)C, DEPT, (1)H-(1)H COSY, HSQC, HMBC, and NOESY. The absolute configuration of an epimeric mixture of 1 and 2 was determined by X-ray crystallographic analysis. The compounds isolated were evaluated for their binding propensity in several CNS G-protein-coupled receptor assays in vitro.