Chemical composition of abaca (Musa textilis) leaf fibers used for manufacturing of high quality paper pulps

The chemical composition of leaf fibers of abaca (Musa textilis), which are commonly used for high-quality paper pulp production, was thoroughly studied. The results revealed that the lignin content was 13.2% of the total fiber. The analysis of abaca fibers by pyrolysis coupled to gas chromatography-mass spectrometry (Py-GC/MS) released predominantly compounds arising from lignin and p-hydroxycinnamic acids, with high amounts of 4-vinylphenol. The latter compound was demonstrated to arise from p-coumaric acid by pyrolysis of abaca fibers in the presence of tetramethylammonium hydroxide, which released high amounts of p-coumaric acid (as the methyl derivative). Products from p-hydroxyphenyl (H), guaiacyl (G), and syringyl (S) propanoid units, with a predominance of the latter (H:G:S molar ratio of 1.5:1:4.9), were also released after Py-GC/MS of abaca fibers. Sinapyl and coniferyl acetates, which are thought to be lignin monomer precursors, were also found in abaca. The extractives content of the abaca fiber (0.4%) was low, and the most predominant compounds were free sterols (24% of total extract) and fatty acids (24% of total extract). Additionally, significant amounts of steroid ketones (10%), triglycerides (6%), omega-hydroxyfatty acids (6%), monoglycerides (4%), fatty alcohols (4%), and a series of p-hydroxycinnamyl (p-coumaric and ferulic acids) esterified with long chain alcohols and omega-hydroxyfatty acids were also found, together with minor amounts of steroid hydrocarbons, diglycerides, alpha-hydroxyfatty acids, sterol esters, and sterol glycosides.     

Isolation, Characterization, and NO Inhibitory Activities of Sesquiterpenes from Blumea balsamifera

Blumea balsamifera belongs to the family Compositae, and its leaves have been used as a flavoring ingredient and a tea. A phytochemical investigation of the aerial parts of B. balsamifera led to the isolation of 10 new (1-10) and 1 known (11) sesquiterpenes. Their structures were elucidated on the basis of extensive one- and two-dimensional nuclear magnetic resonance (heteronuclear multiple-quantum coherence, heteronuclear multiple-bond correlation, (1)H-(1)H correlation spectroscopy, and nuclear Overhauser effect spectrometry) spectroscopic data analyses, and the structure of compound 1 was confirmed by X-ray crystallography. The inhibitory activities on lipopolysaccharide-induced NO production in murine microglial BV-2 cells of these sesquiterpenes were evaluated, and all of the compounds showed inhibitory effects.     

Antimicrobial properties of Quercus ilex L. proanthocyanidin dimers and simple phenolics: evaluation of their synergistic activity with conventional antimicrobials and prediction of their pharmacokinetic profile

The antibacterial and antifungal activities of an ample number of phenolic compounds isolated from Quercus ilex leaves, belonging to the classes of flavonoids, proanthocyanidins, and phenolic acids, are discussed. The isolation of A type proanthocyanidin, (+)-epigallocatechin-(2β→O→7, 4β→8)-(+)-catechin is reported for the first time. Its structure was established by means of highfield NMR (correlation spectroscopy, heteronuclear single quantum correlation, heteronuclear multiple bond correlation, and rotating frame Overhauser effect spectroscopy) and MS spectral analyses, while its absolute configuration was determined by circular dichroism measurements. The isolated compounds were tested for their antimicrobial effects against eight human bacterial species and 14 fungal species. In a second step, the most potent compounds were tested in combination with the conventional fungicides, bifonazole and ketoconazole, to evaluate possible synergistic effects. Results showed that proanthocyanidins 3 and 4 when combined with bifonazole and ketoconazole increase the activity of both of these conventional fungicides. Moreover, the pharmacokinetic profile of the isolated compounds was investigated using computational methods.     

Megastigmane and phenolic components from Laurus nobilis L. leaves and their inhibitory effects on nitric oxide production

Laurus nobilis L. leaves are widely used in cooking and in folk medicine. Five new megastigmane glucosides (2-4, 7, and 9) named laurosides A-E and a new phenolic glucoside 12 were isolated from the methanolic extract of L. nobilis L. leaves, along with 10 known components: megastigmane (5), megastigmane glucosides (1, 6, 8, 10, and 11), aromatic compounds (13 and 14), and flavonoids (15 and 16). The structures and relative stereochemistry have been elucidated by one- and two-dimensional nuclear magnetic resonance experiments ((1)H and (13)C NMR, DEPT, correlation spectroscopy, heteronuclear multiple quantum correlation, heteronuclear multiple bond correlation, and nuclear Overhauser enhancement spectroscopy) and by chemical derivatization. The effect of isolated compounds on nitric oxide production in lipopolysaccharide-activated murine macrophages were examined.     

Sesamolinol glucoside, disaminyl ether, and other lignans from sesame seeds

The application of a procedure based on XAD-4 adsorption resin permitted the obtainment of an enriched polyphenolic extract from Sesamum indicum seeds. Chemical analysis of the obtained extract led to the identification of 12 lignans. Among them, 2 lignans, (+)-sesamolinol-4'-O-β-D-glucoside and disaminyl ether, are reported for the first time as natural compounds. Their structure has been determined by spectroscopic methods, mainly by the application of one-dimensional (1D) and two-dimensional (2D) nuclear magnetic resonance (NMR) techniques [heteronuclear multiple-quantum coherence (HMQC), heteronuclear multiple-bond correlation (HMBC), and nuclear Overhauser effect spectrometry (NOESY)] and mass spectroscopy. The isolated compounds were evaluated for their antimutagenic activity. Among the tested lignans, the most active lignan was found to be sesamolin, followed by sesamolinol and samin, against H(2)O(2). Additionally, some of the tested lignans showed desmutagenic activity against benzo[a]pyrene (BaP).     

Studies on the constituents of Cyclanthera pedata fruits: isolation and structure elucidation of new triterpenoid saponins.

The isolation of nine triterpenoid saponins (1-9), among them six new natural compounds (1-6), from the MeOH extract of the fruits of Cyclanthera pedata is reported. All of the structures were elucidated by spectroscopic methods, including the concerted application of one-dimensional (1)H-(1)H total correlation spectroscopy, (1)H-(1)H nuclear Overhauser effect spectroscopy), and (13)C-(13)C DEPT-NMR and two-dimensional NMR techniques (double-quantum filtered correlated spectroscopy, rotating-frame Overhauser enhancement spectroscopy, heteronuclear single quantum coherence, and heteronuclear multiple bond correlation). A comparative study of seeds and fruits has been also carried out.     

Novel protoporphyrinogen oxidase inhibitors: 3H-pyrazolo[3,4-d][1,2,3]triazin-4-one derivatives

A series of 3 H-pyrazolo[3,4-d][1,2,3]triazin-4-one derivatives were synthesized as candidate herbicides by diazotization of different 5(3)-amino- N-phenyl-1 H-pyrazole-4-carboxamide derivatives prepared by the reaction of substituted 5(3)-amino-pyrazole-4-carbonyl chloride with a substituted aniline. Their structures were identified by (1)H NMR and elemental analyses. The isomers D and E were isolated, and their structures were identified by two-dimensional NMR analyses (heteronuclear single quantum coherence and heteronuclear multiple-bond correlation) and single-crystal X-ray diffraction analysis. The bioassay results showed that some of the title compounds exhibited both excellent herbicidal activity at a dose of 93.75 g/ha and strong inhibition against protoporphyrinogen oxidase activity in vitro. The structure-activity relationship showed that D16 possessed the highest activities both in vivo and in vitro when the N-substituted group of the pyrazole ring was allyl and the N-substituted group of benzooxazinone was propargyl.     

Characterization of sulfated quercetin and epicatechin metabolites

Different monosulfates of quercetin and epicatechin with metabolic interest were obtained by hemisynthesis and characterized regarding their chromatographic behavior and absorption and mass spectra. Three of these compounds were further isolated, and their structures were elucidated by mass spectrometry and (1)H and (13)C nuclear magnetic resonance using one- and two-dimensional techniques (heteronuclear single-quantum coherence and heteronuclear multiple-bond correlation). The calculation of the proton and carbon shifts caused by sulfation allowed for the assignment of the position of the sulfate group in the flavonoids, so that the compounds were identified as quercetin-3'-O-sulfate, quercetin 4'-O-sulfate, and epicatechin 4'-O-sulfate. It was found that sulfation at position 3' induced a large upfield shift in the carbon bearing the sulfate group and downfield displacements of the adjacent carbons, whereas no significant upfield or downfield shifts were observed with respect to the parent flavonoid when sulfation was produced at position 4'.     

Highly acylated (acetylated and/or p-coumaroylated) native lignins from diverse herbaceous plants

The structure of lignins isolated from the herbaceous plants sisal ( Agave sisalana), kenaf ( Hibiscus cannabinus), abaca ( Musa textilis) and curaua ( Ananas erectifolius) has been studied upon spectroscopic (2D-NMR) and chemical degradative (derivatization followed by reductive cleavage) methods. The analyses demonstrate that the structure of the lignins from these plants is highly remarkable, being extensively acylated at the gamma-carbon of the lignin side chain (up to 80% acylation) with acetate and/or p-coumarate groups and preferentially over syringyl units. Whereas the lignins from sisal and kenaf are gamma-acylated exclusively with acetate groups, the lignins from abaca and curaua are esterified with acetate and p-coumarate groups. The structures of all these highly acylated lignins are characterized by a very high syringyl/guaiacyl ratio, a large predominance of beta- O-4' linkages (up to 94% of all linkages), and a strikingly low proportion of traditional beta-beta' linkages, which indeed are completely absent in the lignins from abaca and curaua. The occurrence of beta-beta' homocoupling and cross-coupling products of sinapyl acetate in the lignins from sisal and kenaf indicates that sinapyl alcohol is acetylated at the monomer stage and that, therefore, sinapyl acetate should be considered as a real monolignol involved in the lignification reactions.     

Analysis of plant complex matrices by use of nuclear magnetic resonance spectroscopy: St. John's wort extract

The efficiency of two-dimensional homonuclear (1)H--(1)H correlated spectroscopy and two-dimensional reverse heteronuclear shift correlation spectroscopy (i.e., heteronuclear multiple quantum correlation) in characterizing and evaluating the relative content of herbal extract constituents is demonstrated. These experiments are able to fully assign the proton and carbon resonances of all three classes of constituents present in dried commercial extract of St. John's wort, that is, flavonols, phloroglucinols, and naphthodianthrones, with particular regard to the very unstable phloroglucinols. In addition, shikimic and chlorogenic acids, sucrose, lipids, polyphenols, and traces of solvents of the extractive process (methanol) were also identified. These experiments can be considered to be a very simple and fast analytical method for determining the quality and stability of the titled commercial extract. They represent a generally applicable technique for a rapid screening and a specific measurement of other commercial phytochemicals or, in selected cases, an alternative to the classical analytical techniques such as high-performance thin-layer chromatography, high-performance liquid chromatography, capillary gas chromatography, and electrophoresis.     

Purification and properties of a novel phenolic antioxidant from Radix astragali fermented by Aspergillus oryzae M29

The Chinese herb Radix astragalus (RA) has been widely used as a dietary supplement in Asia, and there are numerous reports on its bioactivities. However, there are no reports to date regarding the use of Aspergillus spp. in the culture medium of the RA plant for the production of phenolic antioxidants. In this study, utilizing the fungus Aspergillus to ferment the native RA has successfully resulted in a significant increase in the phenolic contents of RA, and the fermented RA also revealed much better antioxidant activity toward 2,2'-azinobis (3-ethylbenzothiazoline-6-sulfonic acid (ABTS) radicals, hydroxyl radical, superoxide radical and peroxyl radical than those of unfermented RA. Among these phenolics, a potent novel antioxidant was isolated and identified as 3,4-di(4'-hydroxyphenyl) isobutyric acid with a molecular weight of 272, by ESI-MS (electrospray ionization mass), 1H NMR (nuclear magnetic resonance), 13C NMR, DEPT (distortionless enhancement by polarization transfer)-NMR, HMQC (heteronuclear multiple quantum coherence), and HMBC (heteronuclear multiple bond correlation) spectra. These data demonstrated that the solid-state bioprocessing strategy could be an innovative approach to enhance the antioxidant activity of RA.     

Epicatechin carbonyl-trapping reactions in aqueous maillard systems: Identification and structural elucidation

Recently, our group reported via labeling experiments that epicatechin in Maillard reaction aqueous glucose-glycine model systems formed adduct reaction products with C2, C3, and C4 sugar fragments. In the current study, we investigated the identity of the sugar fragment precursors responsible for adduct generation by directly comparing the liquid chromatography-mass spectrometry properties of these reported epicatechin (EC)-sugar fragments adducts with those generated from reactions consisting of only EC and well-known Maillard-generated glucose fragments (i.e., glyoxal, glycolaldehyde, methylglyoxal, glyceraldehyde, etc.). The structural properties of an EC-methylglyoxal adduct reaction product were also analyzed by NMR. The most likely precursors for the C2, C3, and C4 sugar moiety of the EC-sugar fragment adducts were identified as glyoxal, hydroxyacetone, and erythrose, respectively. 1H NMR analysis of the EC-methylglyoxal indicated that the analyte underwent rapid conformational/constitutional exchange. Using cold temperature (-25 degrees C) two-dimensional NMR analyses (heteronuclear multiple bond coherence, heteronuclear multiple quantum coherence, and 1H-(1)H correlation spectroscopy), the structure of one of the isomers was reported to consist of a covalent linkage between the C1 position of the methylglyoxal and either the C6 or the C8 position of the EC A ring, presumably generated by hydroxyalkylation and aromatic substitution reactions.     

Chemical structures of corn stover and its residue after dilute acid prehydrolysis and enzymatic hydrolysis: insight into factors limiting enzymatic hydrolysis

Advanced solid-state NMR techniques and wet chemical analyses were applied to investigate untreated corn stover (UCS) and its residues after dilute acid prehydrolysis (DAP) and enzymatic hydrolysis (RES) to provide evidence for the limitations to the effectiveness of enzyme hydrolysis. Advanced solid-state NMR spectral-editing techniques as well as 1H-13C two-dimensional heteronuclear correlation NMR (2D HETCOR) were employed. Our results indicated that dilute acid prehydrolysis selectively removed amorphous carbohydrates, increased aromatic CH/other protonated -C═C- and enriched alkyl CH and CH2 components. Cinnamic acids were increased, and proteinaceous materials and N-containing degradation or condensation compounds were absorbed or coprecipitated in RES. 2D HETCOR experiments indicated a close association between lignin and the residual carbohydrates. Ketones/aldehydes were not detected in the DAP, in contrast to a report in which an appreciable amount of ketones/aldehydes was generated from the acid pretreatment of a purified cellulose in the literature. This suggested that acid pretreatment may modify the structure of purified cellulose more than biomass and that biomass may be a better substrate than model biopolymers and compounds for assessing structural changes that occur with industrial processing. On the basis of NMR and wet chemical analyses, we found the following factors could cause the limitations to the effectiveness of enzymatic hydrolysis: (1) chemical modification of carbohydrates limited the biologically degradable carbohydrates available; (2) cinnamic acids in the residue accumulated; (3) accessibility was potentially limited due to the close association of carbohydrates with lignin; and (4) proteinaceous materials and N-containing degradation or condensation compounds were absorbed or coprecipitated.     

Identification and quantification of caffeic and rosmarinic acid in complex plant extracts by the use of variable-temperature two-dimensional nuclear magnetic resonance spectroscopy

A combination of advanced nuclear magnetic resonance (NMR) methodologies for the analysis of complex phenolic mixtures that occur in natural products is described, with particular emphasis on caffeic acid and its ester derivative, rosmarinic acid. The combination of variable-temperature two-dimensional proton-proton double quantum filter correlation spectroscopy (1H-1H DQF COSY) and proton-carbon heteronuclear multiple quantum coherence (1H-13C HMQC) gradient NMR spectroscopy allows the identification and tentative quantification of caffeic and rosmarinic acids at 243 K in extracts from plants of the Lamiaceae family, without resorting to previous chromatographic separation of the components. The use of proton-carbon heteronuclear multiple bond correlation (1H-13C HMBC) gradient NMR spectroscopy leads to the complete assignment of the correlations of the spins of H2a and H3a with the ester and carboxyl carbons of rosmarinic and caffeic acid, even at room temperature, and confirms the results of the above methodology Quantitative results are in reasonable agreement with reverse phase HPLC measurements.     

NMR and molecular mechanics study of pyrethrins I and II.

Bioassay-directed fractionation of the organic extract of the Kenyan pyrethrum flowers (Chrysanthemum cinerariaefolium Vissiani) resulted in the isolation of two natural pyrethrin esters, pyrethrin I (PI) and pyrethrin II (PII) as the major constituents. These esters elicited inhibition of the multiple drug resistant (MDR) Mycobacterium tuberculosis. The high-field (1)H and (13)C nuclear magnetic resonance (NMR) chemical shifts of PI and PII were unequivocally assigned using modern two-dimensional (2D) proton-detected heteronuclear multiple-quantum coherence (HMQC) and heteronuclear multiple-bond correlation (HMBC) experiments. The conformations of both esters were deduced from (1)H-(1)H vicinal coupling constants and confirmed by 2D nuclear Overhauser effect spectroscopy (NOESY). Computer molecular modeling (MM) studies revealed that PI and PII molecules adopt a "love-seat" conformation in chloroform (CDCl(3)) solution.     

Characterization of oligomeric procyanidins and identification of quercetin glucuronide from lotus ( Nelumbo nucifera Gaertn.) seedpod

Procyanidins are a class of polyphenols in the plant kingdom. Lotus ( Nelumbo nucifera Gaertn.) seedpods, the inedible part of lotus and a byproduct during the production of lotus seeds, were found to be a new source rich in procyanidins. Detailed information about oligomeric procyanidins in lotus seedpods remains unknown. In this study, lotus seedpods were extracted using 60% aqueous methanol and characterized with phloroglucinolysis and liquid chromatography (mass spectrometry with an electrospray ionization source). The results indicate that the oligomeric and polymeric fraction had a mean degree of polymerization of 3.2 and 15.4, respectively, and consisted of (+)-catechin (m/z 289), gallocatechin or epigallocatechin (m/z 305), quercetin glycoside (m/z 463), quercetin glucuronide (m/z 477), procyanidin dimers (m/z 577.1), proanthocyanidin dimer gallate (m/z 593.3), prodelphinidin dimers (m/z 609.1), procyanidin trimers (m/z 865.1), etc. Quercetin glucuronide was further purified using flash chromatography and identified as quercetin-3-O-β-glucuronide by determining its exact mass using ion-trap time-of-flight mass spectrometry and 1H and 13C nuclear magnetic resonance, 1H-detected heteronuclear single-quantum coherence, and 1H-detected heteronuclear multiple-bond correlation analyses.     

Solid‐Phase Microextraction Method for Headspace Analysis of Volatile Compounds in Bread Crumb

A solid‐phase microextraction method was developed to analyze volatile compounds in bread crumb. Three different fibers usually used to determine volatile compounds in foodstuffs were tested, and Carboxen/ Polydimethylsiloxane showed the best extraction efficiency. This method can determine ≈65 compounds, although only the main compounds responsible for the crumb aroma profile (6 alcohols, 5 aldehydes, 5 acids, and 2 ketones) were evaluated quantitatively with a relative standard deviation <20% and limits of detection were 1–18 mg/kg. All the compounds showed a good linearity in concentration ranges of interest with acceptable correlation coefficients (r > 0.99) and recoveries close to 100%. The optimized solid‐phase microextraction (SPME) method was applied to determine aromatic compounds in some precooked frozen breads marketed in Spain (baguettes and ciabattas).     

A comprehensive approach for quantitative lignin characterization by NMR spectroscopy

A detailed approach for the quantification of different lignin structures in milled wood lignin (MWL) has been suggested using a combination of NMR techniques. 1H-13C heteronuclear multiple quantum coherence and quantitative 13C NMR of nonacetylated and acetylated spruce MWL have been found to have a synergetic effect, resulting in significant progress in the characterization of lignin moieties by NMR. About 80% of side chain moieties, such as different beta-O-4, dibenzodioxocin, phenylcoumaran, pinoresinol, and others, have been identified on the structural level. The presence of appreciable amounts of alpha-O-alkyl and gamma-O-alkyl ethers has been suggested. Although the quantification of various condensed moieties was less precise than for side chain structures, reliable information can be obtained. Comparison of the calculated results with known databases on spruce MWL structure shows that the suggested approach is rather informative and comparable with the information obtained from the combination of various wet chemistry methods. Discrepancies between the results obtained in this study and those previously published are discussed.     

Lignin modification during Eucalyptus globulus kraft pulping followed by totally chlorine-free bleaching: a two-dimensional nuclear magnetic resonance, Fourier transform infrared, and pyrolysis-gas chromatography/mass spectrometry study

Chemical modification of eucalypt lignin was investigated during kraft pulping and chlorine-free bleaching by comparing milled wood lignin, kraft lignin, and pulp enzymatic residual lignins. The syringyl-to-guaiacyl ratio (S/G) from analytical pyrolysis slightly changed during pulping and bleaching (S/G, 3-4) but was higher in the kraft lignin. Semiquantitative heteronuclear single quantum correlation (HSQC) nuclear magnetic resonance (NMR) showed that the relative amount of beta-O-4' (around 80% side chains) and resinol type substructures (15%) was slightly modified during pulping and oxygen delignification. However, a decrease of resinol substructures (to only 6%) was found after alkaline peroxide bleaching. The relative amount of surviving linkages in the highly phenolic kraft lignin was dramatically modified; resinols were predominant. Oxygen delignification did not change interunit linkages, but a relative increase of oxidized units was found in the HSQC aromatic region, in agreement with the small increase of pyrolysis markers with oxidized side chains. NMR heteronuclear multiple bond correlations showed that the oxidized units after oxygen delignification bore conjugated ketone groups.     

Antifungal activity and biotransformation of diisophorone by Botrytis cinerea

Diisophorone (1) was tested against two strains of the necrotrophic plant pathogen Botrytis cinerea.Fungal sensitivity varied according to the strain. B. cinera 2100 was more sensitive than B. cinereaUCA992: its mycelial growth was significantly inhibited at concentrations of 50 ppm and above. Although diisophorone (1) showed an effective control of B. cinerea, a detoxification mechanism was present. The detoxification of racemic diisophorone (1) by B. cinerea was investigated. Incubation with two strains of B. cinerea gave one and four biotransformation products (2-5), respectively. Their structures were established as the known 8beta-hydroxydiisophorone (2), 6alpha-hydroxydiisophorone (3), 6beta-hydroxydiisophorone (4) and 8beta,14beta-dihydroxydiisophorone (5) on the basis of their spectroscopic data, including two-dimensional NMR analysis [heteronuclear multiple quantum coherence (HMQC), heteronuclear multiple bond correlation (HMBC), and nuclear Overhauser enhancement spectroscopy (NOESY)] and an X-ray crystallographic study.