Cupric Oxidation Research Articles

The composition of lignin in estuarine suspended particulates and the distribution of particulate lignin in estuaries as determined by capillary gas chromatography of cupric oxide oxidation products

The contribution of lignin to the estuarine suspended particulate fraction is described in terms of its distribution over a spring tide and a neap tide in the Tamar Estuary, Southwest England and the Mersey Estuary, Northwest England. Suspended particulate samples are treated with CuONaOH at 170°C to yield simple, lignin-derived phenolic compounds, which are separated, derived and quantified by capillary gas chromatography with flame ionization detection (GC-FID). Estuarine distributions of particulate lignin are described and the sources and compositional changes of the lignin identified. Comparison of the lignin concentrations in the suspended material with those in the underlying sediment reveals that lignin is preferentially enriched in the suspended material.

Degradation of polysaccharides and lignin by ruminal bacteria and fungi.

Bermudagrass (Cynodon dactylon) leaf blades and whole cordgrass (Spartina alterniflora) fiber were evaluated for degradation of cell walls by microbial groups in ruminal fluid. The groups were selected by the addition of antibiotics to the inoculum as follows: (i) whole ruminal fluid (WRF), no antibiotics; (ii) cycloheximide (C) to inhibit fungi, thus showing potential bacterial activity; (iii) streptomycin and penicillin (S,P) to inhibit fiber-degrading bacteria, showing potential fungal activity; (iv) streptomycin, penicillin, and chloramphenicol (S,P,CAM) to inhibit all bacteria including methanogens; (v) streptomycin, penicillin, and cycloheximide (S,P,C) to inhibit all microbial activity as a control; and (vi) autoclaved ruminal fluid (ARF) to inhibit all biological activity as a second control. Scanning electron microscopy of tissue degradation indicated that tissues not giving a positive histological reaction for lignin were more readily degraded. Cordgrass was more highly lignified, with more tissues resisting degradation than in bermudagrass. Patterns of degradation due to treatment resulted in three distinct groups of data based on the extent of fiber or component losses: WRF and C greater than S,P and S,P,CAM greater than S,P,C and ARF. Therefore, bacterial activity was responsible for most of the fiber loss. Fiber degradation by anaerobic fungi was significantly less (P = 0.05). Cupric oxide oxidation of undigested and digested bermudagrass fiber indicated that phenolic constituents differed in their order of resistance to removal or solubilization. Vanillyl and syringyl components of lignin were the most resistant to decomposition, whereas ferulic acid was readily solubilized from fiber in the absence of microbial activity.(ABSTRACT TRUNCATED AT 250 WORDS)

Biogeochemical cycling in an organic-rich coastal marine basin: 9. Sources and accumulation rates of vascular plant-derived organic material

The sources, degradation and burial of vascular plant debris deposited over the past several decades in the lagoonal sediments of Cape Lookout Bight, North Carolina, are quantified using alkaline cupric oxide lignin oxidation product (LOP) analysis. Non-woody angiosperms, accounting for 92 ± 32% of the recognizable sedimentary vascular plant debris, are calculated to contribute 23 ± 17% of the total organic carbon buried over the past decade (upper meter of sediment column). When combined with a previously established sedimentary organic carbon budget for this site (Martens and Klump, 1984; Martens et al., 1987, in preparation) a vascular plant derived carbon burial rate of 26 ± 20 mole C m−2 yr−1 is calculated for this same time interval. The refractory nature and invariant depth distributions of the lignin oxidation products (LOP), when coupled with evidence for constant degradation rates of metabolizable materials, indicate that sediment accumulation at this site has been a steady state process with respect to source and burial of organic carbon since its conversion from an inner-continental shelf to a lagoonal environment during the late 1960's. Thus systematic down-core decreases in labile organic matter result from early diagenetic processes rather than input rate variations.

Aliphatic structures in peat fulvic acids

Fulvic acids, extracted from strongly humified milled peat, were studied through 13C-NMR measurements and cupric oxide oxidations. The 13C-NMR spectrum shows signals caused by the carboxylic carbons, aromatic or olefinic carbons, acetal carbons, aliphatic alcohol carbons and methylene and methyl carbons. In cupric oxide oxidation three fractions were obtained: an ethyl acetate soluble fraction, yielding 11 %, a base and acid soluble fraction (“oxidized fulvic acids”), yielding 25 % and a base soluble,acid insoluble fraction yielding 4 %.

THE DISTRIBUTION OF LIGNIN DERIVATIVES IN FOSSIL PLANTS.

Lignin derivatives have been obtained from a range of sub-fossil woods and fossil plant compressions by cupric oxide oxidation. Fossil woods yield essentially the same products as wood from living spermatophytes. Although some demethoxylation of the phenolic aldehydes occurs during fossilization, the overall difference between conifer and angiosperm wood is maintained. Compressions of varied plant organs yield recognizable phenolic aldehydes, though often in small amounts.

Lignin geochemistry of sediments from the Narragansett Bay Estuary

Cupric oxide oxidation has been employed to characterize the lignin geochemistry of Narragansett Bay sediments. Lignin concentrations throughout the estuary are low when expressed on a carbon-normalized basis, but can be characterized as enriched when expressed on a mass-normalized basis. This implies substantial dilution of the sedimentary lignin by inputs of lignin-poor carbon. Lignin concentrations do not correlate with the 13C isotopic composition of the sedimentary organic matter. These results are consistent with a sediment lignin component consisting of varying amounts of vascular plant debris and lignin-depleted organic matter, the latter originating from both marine (planktonic) and terrestrial (uncharacterized) sources. Compositional plots of lignin-derived phenols show that sediments in the upper estuary are influenced to a greater extent by gymnosperm lignin sources than those in the mid-and lower estuary. Given the extent to which the upper estuary is affected by pollution sources, inputs from anthropogenic discharges are the most likely cause of these compositional differences. However, an evaluation of processed paper products as an “anthropogenic” lignin source indicates that the lignin content of these materials is insufficient to account for the levels found in the sediments. Subsurface lignin compositions at an upper estuary site reveal that lignin originating from the inferred anthropogenic sources disappears at a depth shallower than that which would be expected based on the distribution of other trace organic pollutants (hydrocarbons and several synthetic organic compounds). We speculate that differences in either the depositional history or the degree of preservation of these two compound classes are responsible for the observed trends.

Evaluation of lignin and cellulose contributions to low-rank coal formation by alkaline cupric oxide oxidation

Oxidation with alkaline cupric oxide has been combined with solid-state 13C-n.m.r. spectroscopy to evaluate the cellulose and lignin input to the formation of low rank coals. Model studies carried out on lignin-cellulose mixtures, carbonified cellulose, synthetic coals and synthetic humic acids (melanoidins) have established m-hydroxybenzoic acid as a source indicator for cellulose. This product has been found specifically in the oxidation of bio- and geo-macromolecules containing cellulose and/or thermally altered cellulosic material and is chemically distinct from the characteristic oxidation products of lignin-derived materials. Systematic changes in the distribution of the major oxidation products from lignin provide a chemical fingerprint which, in general, can be utilized to trace the thermal history of the sample. Analysis of two lignite coals from the northern hemisphere has demonstrated their predominantly lignitic origin. On the other hand, a Victorian brown coal (pale lithotype) sample is shown to contain a significant amount of highly transformed carbohydrate materials which are presumably incorporated into the macromolecular structure as humic acid derivatives.

Chemical evidence of differential particle dispersal in the southern washington coastal environment

Aliphatic hydrocarbons, cupric oxide oxidation products of lignin and polycyclic aromatic hydrocarbons (PAH) were analyzed by capillary gas chromatography in sediments from the southern Washington continental shelf and slope. The concentration of diploptene relative to plantwax n- alkanes increased systematically in surface sediments with distance offshore along east-west transects of the study area and remained constant in surface sediments along the midshelf silt deposit. Analogous trends were also observed for the concentration of cinnamyl phenols relative to vanillyl phenols and total methylphenanthrenes relative to phenanthrene. These changes in sedimentary composition are evidence that diploptene from some terrestrial source, lignin characteristic of non-woody vascular plant tissue and a fossil organic material contained within weathered rock debris disperse across the Washington continental shelf and slope in geographic patterns distinct from that for other river-derived, chemically related materials. The compositional variations are explained by the particulate associations of the land-derived chemicals and differential hydraulic dispersion of their respective carrier particles after discharge at the mouth of the Columbia River.

Association of epidermal lignification with nonhost resistance of cucurbits to fungi

Etiolated seedlings of cucumber, muskmelon, watermelon, pumpkin, and squash were inoculated with Colletotrichum lagenarium (Pass.) Ell. & Halst. race 1, Colletotrichum atramentarium (Berk. & Br.) Taub., or Helminthosporium carbonum Ull. The fungi germinated and produced appressoria equally well on all hosts. Penetrations into host cells, however, were routinely observed only in the interactions of C. lagenarium with cucumber, muskmelon, and watermelon. Histochemical staining of the other host–pathogen interactions (nonhost interactions) revealed that the lack of penetration by fungi into nonhost tissue was associated with the deposition of lignin in the upper and lateral host epidermal cell walls around appressoria. Treatment of the lignified tissues with 0.5 M NaOH or hot ethanol did not alter staining reactions. Treatment with hot aminoethanol, a delignifying agent, did eliminate stainings. Cupric oxide oxidation of lignified epidermal cell walls from each host revealed that each host produced a p-coumaryl-rich, guaiacyl-poor ligninlike material. These results suggest that lignification may be a general nonhost resistance response in the Cucurbitaceae.

Determination of natural and wound-induced potato tuber suberin phenolics by thioglycolic acid derivatization and cupric oxide oxidation

Application of two lignin chemistry techniques, CuO oxidation and thioglycolic acid derivatization, has confirmed that natural and wound-induced suberin does have a lignin-like component. The time course of suberin phenolic deposition could be monitored by using the thioglycolic acid technique. The thioglycolic acid procedure is easily performed, can be adapted readily to multiple samples, and thus could be an effective tool for studying rates of wound-induced suberization.

Characterization of lignin in forest humus layers by high-performance liquid chromatography of cupric oxide oxidation products

A rapid method is described to prepare samples, separate and quantify phenolic lignin degradation products obtained from the alkaline cupric oxide oxidation of forest humus layers. The whole sample is treated with CuO-NaOH at 170°C to produce simple lignin-derived phenols, which are extracted with a commercially-available disposable column system. The extracted phenolic compounds are separated by reverse-phase high-performance liquid chromatography, based on the differential polarity of benzoic acids, aldehydes and cinnamic acids. Results from the oxidation of gymnosperm and angiosperm litter are presented.

Lignin Signature of Aquatic Humic Substances

Lignin-derived phenols dominate the cupric oxide oxidation products of dissolved humic substances from river and lake waters. The relative distributions of these phenols suggest the presence of intact, though oxidized, lignin, which is indicative of the locally dominant vascular plant vegetation. Recognizable lignin is present mostly in humic acid as opposed to fulvic acid fractions. This lignin component represents a source-specific and process-dependent tracer that can uniquely characterize dissolved organic matter.

Rapid deposition of lignin in potato tuber tissue as a response to fungi non-pathogenic on potato

Potato tuber slices were inoculated with a pathogen ( Fusarium roseum f.sp. sambucinum) and with pathogens of other plant species ( Cladosporium cucumerinum and F. roseum). Growth of all three fungi on the tuber tissue surface was similar through the first 12 h; after this time, the pathogen, but not the non-pathogens, began to penetrate into the tissue and caused a gradual softening and darkening of the tissue. The non-pathogens caused a yellow discoloration in the host cell walls, first detected 8–10h after inoculation. These yellow areas of the cell walls stained positively for lignin. Positive staining for lignin was not observed in the pathogen-challenged tissues until 18–20 h. Tissues inoculated with the non-pathogens and treated with thioglycolic acid gave a ligninthioglycolate in much greater yields than did control tissues or tissues inoculated with the pathogen. Ligninthioglycolic acid yields increased in the non-pathogen-challenged tissues by 9 h and in pathogen-challenged tissues by 18 h after inoculation. Cupric oxide oxidation of alkali pre-treated tissues also provided evidence for the formation of lignin. Alkali pre-treatment had no effect on ligninthioglycolic acid yields or on histochemical staining, which is further evidence for the production of lignin. Pre-treatment of the tissue with α-aminooxyacetic acid, a competitive inhibitor of phenylalanine ammonia lyase, or abscisic acid induced a state of “susceptibility” in the tissue to C. cucumerinum, and this was associated with a lack of lignin deposition.

High-Performance Liquid Chromatography of Preparations of Ribonucleic Acid Inactivator(s) from Cupric Ion and Hydroquinone Before and After Treatment with Histidine

□ Preparations of viral RNA inactivator(s) produced during the cupric ion-catalyzed oxidation of hydroquinone were analyzed by high-performance liquid chromatography (HPLC) using UV and electrochemical (EC) detectors. In addition to hydroquinone and the main oxidation product (p-benzoquinone), which is known not to be the inactivator(s), the analysis showed three unidentified components (I–III). Partial UV absorption spectra of I–III were determined by HPLC with the UV detector set at various wavelengths. Components II and III, but not I, were highly unstable in the presence of L-histidine, which is an excellent chelator of cupric ion and can promptly stop ongoing viral RNA inactivation by the inactivator(s). The product p-benzoquinone was also highly unstable in the presence of L-histidine; the reaction between these two compounds (with or without copper) resulted in a cascade of products. The possibility that the inactivator(s) is II or III, or both, is discussed.

Characterization of lignin by gas capillary chromatography of cupric oxide oxidation products

A sensitive, reproducible method is described for the characterization of lignin in untreated plant and geochemical samples containing 10 mg or less organic matter. The whole sample is treated with alkaline cupric oxide at 170 degrees to produce simple lignin- derived phenols that are extracted with ethyl oxide and analyzed by gas capillary chromatography on fused silica columns. A pattern of 11 or less phenols is produced that reflects the relative concentration and plant tissue sources of lignins present in the sample.

A stopped-flow study of the cupric ion oxidation of adult-human haemoglobin.

The addition of excess Cu2+ to adult human haemoglobin leads to the production of alpha 2(2+) beta 2(3+), in both the oxy and deoxy forms of the protein. Stopped-flow studies of the oxidation process yields apparent second-order rate constants of 196M-1 X S-1 and 41M-1 X S-1 for the deoxy and oxy forms respectively. The rate of the deoxy-form oxidation is linearly dependent on [Cu2+], whereas that of the oxy form is rate-limited above 2 mM to 0.11 S-1. Arrhenius activation energies of the two processes are almost identical at 91 kJ X mol-1, as are the activation enthalpies of 89 kJ X mol-1. The activation entropies show small differences, being 31 entropy units and 48 entropy units for the oxy and deoxy forms respectively. ATP and glycerate 2,3-bisphosphate at saturating concentrations do not affect the rate of oxidation of the oxy form, but halve the rate found for the deoxy form. These data are discussed in terms of the previously proposed mechanism of oxidation in which slow Cu2+ binding is followed by rapid electron transfer.

The characterization of plant tissues by their lignin oxidation products

The cupric oxide oxidation products of 23 vascular and nonvascular plant tissues have been measured. Compositional data for vanillyl, syringyl, and cinnamyl phenols are presented in the form of five lignin parameters which are related to plant variety, lignin concentration, and tissue type. On the basis of these parameters, the 23 plant tissue samples are resolved into distinct compositional regions corresponding to: 1. (a) nonvascular plants, 2. (b) gymnosperm woods, 3. (c) non woody gymnosperm tissues, 4. (d) angiosperm woods, and 5. (e) nonwoody angiosperm tissues. The same five parameters also can be determined for organic materials in soils and sediments and used either to discriminate between compositionally different organic mixtures or to estimate the relative amounts of each of the above types of plant materials in the deposits.

ALKALINE CUPRIC OXIDE OXIDATION OF ROOTS AND ALKALINE-EXTRACTABLE ORGANIC MATTER OF CHERNOZEMIC SOILS

Alkaline cupric oxide oxidation was performed on roots of three grass species, on humic substances from three Chernozemic soils, and on three known compounds. The three grass species were Stipa comata Trin. & Rupr., Stipa spartea Trin. var. curtiseta Hitchc, and Festuca scabrella Torr., the three Chernozemic soils were Brown, Dark Brown and Black, and the three known compounds were vanillin, p-hydroxybenzaldehyde, and syringaldehyde. The intent of the study was to obtain a qualitative overview of the set of ethyl ether-soluble products. The gas chromatographic patterns were, therefore, qualitatively assessed and only some of the major peaks were tentatively identified. The ethyl ether-soluble fraction represented only up to 5.7% of the total root C. Most of the compounds identified in the root digests were also present in digests of the alkaline-soluble humic substances. There were quantitative differences for each chromatographic peak among root species, among time of sampling and growth location, and among extraction procedures of humic substances. The oxidation of the three known compounds in turn led to a number of compounds. The CuO-NaOH oxidation may thus cause alterations in similar compounds released from the original structure which will lead to either inflated or deflated yields of compounds that were part of the original structure.

Selective Photometric Determination of Betamethasone Benzoate and Other 21-Hydroxycorticosteroids

Condensation of the glyoxal obtained by cupric acetate oxidation of 21-hydroxycorticosteroids with acetous phenylhydrazine reagent affords a near UV chromophore. All of the tested corticosteroids, including triamcinolone acetonide, which gives low color yields in the Porter-Silber reaction and its Lewbart-Mattox modification, gave similar absorption maxima (362−370nm) and molar absorptivities (ε=17,000−20,500). Since corticosteroid 21-esters and oxidation products do not undergo the reaction, the assay method based on it is stability indicating for betamethasone benzoate and the other test compounds. Procedures are described for the assay of two topical betamethasone benzoate preparations and hydrocortisone and prednisolone tablets; recovery and precision data are given.

New syntheses of pterin based on ethylenediamine and ethanolamine

1. Pterin was obtained by heating 2,6-diamino-4-hydroxy-5-nitrosopyrimidine with ethanolamine in ethylene glycol, and it is also formed by reacting 2,6-diamino-4-hydroxypyrimidine with ethylenediamine in ethylene glycol in the presence of cupric acetate and subsequent oxidation with alkaline hydrogen peroxide. 2. When 2,6-diamino-4-hydroxypyrimidine is reacted with benzylamine or ethanolamine in ethylene glycol in the presence of cupric acetate it, respectively, gives 6-benzylamino- or 6-(β-hydroxyethylamino)-2-amino-4-hydroxypyrimidine.