The biological activity of 4-chloromethylbiphenyl, benzyl chloride and 4-hydroxymethylbiphenyl in 4 short-term tests for carcinogenicity. A report of an individual study in the UKEMS genotoxicity trial 1981

4-Chloromethylbiphenyl (4CMB), benzyl chloride (BC) and 4-hydroxymethyl-biphenyl (4HMB) were tested for biological activity in the following assays: (i) the Salmonella/microsome assay; (ii) a bacterial 'fluctuation' assays; (iii) a DNA repair assay in Hela cells, and (iv) a mouse lymphoma mutation assay. 4CMB was active in assays (i), (ii) and (iii) but not in (iv); BC was active in assays (i), (ii), (iii) but not in (iv) while 4HMB was inactive in all assays. Where biological activity was seen this did not require addition of a liver S9 preparation. 4CMB was more active than BC in all the test systems in which a positive response was obtained. The implication of these results for a test battery approach to in vitro testing is discussed.     

The biological activity of 4-chloromethylbiphenyl,benzyl chloride and 4-hydroxymethylbiphenyl in 4 short-term tests for carcinogenicity: A report of an individual study in the UKEMS genotoxity trial 1981

4-Chloromethylbiphenyl (4CMB), benzyl chloride (BC) and 4-hydroxymethyl-biphenyl (4HMB) were tested for biological activity in the following assays: (i) the Salmonella/microsome assay; (ii) a bacterial ‘fluctuation’ assays; (iii) a DNA repair assay in Hela cells, and (iv) a mouse lymphoma mutation assay. 4CMB was active in assays (i), (ii) and (iii) but not in (iv); BC was active in assays (i), (ii), (iii) but not in (iv) while 4HMB was inactive in all assays. Where biological activity was seen this did not require addition of a liver S9 preparation. 4CMB was more active than BC in all the test systems in which a positive response was obtained. The implication of these results for a test battery approach to in vitro testing is discussed.     

Development and validation of a bioreactor system for dynamic loading and mechanical characterization of whole human intervertebral discs in organ culture

Intervertebral disc (IVD) degeneration is a common cause of back pain, and attempts to develop therapies are frustrated by lack of model systems that mimic the human condition. Human IVD organ culture models can address this gap, yet current models are limited since vertebral endplates are removed to maintain cell viability, physiological loading is not applied, and mechanical behaviors are not measured. This study aimed to (i) establish a method for isolating human IVDs from autopsy with intact vertebral endplates, and (ii) develop and validate an organ culture loading system for human or bovine IVDs. Human IVDs with intact endplates were isolated from cadavers within 48 h of death and cultured for up to 21 days. IVDs remained viable with ~80% cell viability in nucleus and annulus regions. A dynamic loading system was designed and built with the capacity to culture 9 bovine or 6 human IVDs simultaneously while applying simulated physiologic loads (maximum force: 4 kN) and measuring IVD mechanical behaviors. The loading system accurately applied dynamic loading regimes (RMS error <2.5 N and total harmonic distortion <2.45%), and precisely evaluated mechanical behavior of rubber and bovine IVDs. Bovine IVDs maintained their mechanical behavior and retained >85% viable cells throughout the 3 week culture period. This organ culture loading system can closely mimic physiological conditions and be used to investigate response of living human and bovine IVDs to mechanical and chemical challenges and to screen therapeutic repair techniques.     

TNFα Transport Induced by Dynamic Loading Alters Biomechanics of Intact Intervertebral Discs

Objective

Intervertebral disc (IVD) degeneration is an important contributor to the development of back pain, and a key factor relating pain and degeneration are the presence of pro-inflammatory cytokines and IVD motion. There is surprisingly limited understanding of how mechanics and inflammation interact in the IVD. This study investigated interactions between mechanical loading and pro-inflammatory cytokines in a large animal organ culture model to address fundamental questions regarding (i.) how inflammatory mediators arise within the IVD, (ii.) how long inflammatory mediators persist, and (iii.) how inflammatory mediators influence IVD biomechanics.

Methods

Bovine caudal IVDs were cultured for 6 or 20-days under static &amp; dynamic loading with or without exogenous TNFα in the culture medium, simulating a consequence of inflammation of the surrounding spinal tissues. TNFα transport within the IVD was assessed via immunohistochemistry. Changes in IVD structural integrity (dimensions, histology &amp; aggrecan degradation), biomechanical behavior (Creep, Recovery &amp; Dynamic stiffness) and pro-inflammatory cytokines in the culture medium (ELISA) were assessed.

Results

TNFα was able to penetrate intact IVDs when subjected to dynamic loading but not static loading. Once transported within the IVD, pro-inflammatory mediators persisted for 4–8 days after TNFα removal. TNFα exposure induced changes in IVD biomechanics (reduced diurnal displacements &amp; increased dynamic stiffness).

Discussion

This study demonstrated that exposure to TNFα, as might occur from injured surrounding tissues, can penetrate healthy intact IVDs, induce expression of additional pro-inflammatory cytokines and alter IVD mechanical behavior. We conclude that exposure to pro-inflammatory cytokine may be an initiating event in the progression of IVD degeneration in addition to being a consequence of disease.     

The Value of In Vitro Diagnostic Testing in Medical Practice: A Status Report

BackgroundIn vitro diagnostic (IVD) investigations are indispensable for routine patient management. Appropriate testing allows early-stage interventions, reducing late-stage healthcare expenditure (HCE).AimTo investigate HCE on IVDs in two developed markets and to assess the perceived value of IVDs on clinical decision-making. Physician-perceived HCE on IVD was evaluated, as well as desired features of new diagnostic markers.MethodsPast and current HCE on IVD was calculated for the US and Germany. A total of 79 US/German oncologists and cardiologists were interviewed to assess the number of cases where: physicians ask for IVDs; IVDs are used for initial diagnosis, treatment monitoring, or post-treatment; and decision-making is based on an IVD test result. A sample of 201 US and German oncologists and cardiologists was questioned regarding the proportion of HCE they believed to be attributable to IVD testing. After disclosing the actual IVD HCE, the physician’s perception of the appropriateness of the amount was captured. Finally, the association between physician-rated impact of IVD on decision-making and perceived contribution of IVD expenditure on overall HCE was assessed.ResultsIVD costs account for 2.3% and 1.4% of total HCE in the US and Germany. Most physicians (81%) believed that the actual HCE on IVDs was >5%; 19% rated the spending correctly (0–4%, p<0.001). When informed of the actual amount, 64% of physicians rated this as appropriate (p<0.0001); 66% of decision-making was based on IVD. Significantly, more physicians asked for either additional clinical or combined clinical/health economic data than for the product (test/platform) alone (p<0.0001).ConclusionsOur results indicate a poor awareness of actual HCE on IVD, but a high attributable value of diagnostic procedures for patient management. New markers should deliver actionable and medically relevant information, to guide decision-making and foster improved patient outcomes.     

Catabolic cytokine expression in degenerate and herniated human intervertebral discs: IL-1beta and TNFalpha expression profile

Low back pain is a common and debilitating disorder. Current evidence implicates intervertebral disc (IVD) degeneration and herniation as major causes, although the pathogenesis is poorly understood. While several cytokines have been implicated in the process of IVD degeneration and herniation, investigations have predominately focused on Interleukin 1 (IL-1) and tumor necrosis factor alpha (TNFalpha). However, to date no studies have investigated the expression of these cytokines simultaneously in IVD degeneration or herniation, or determined which may be the predominant cytokine associated with these disease states. Using quantitative real time PCR and immunohistochemistry we investigated gene and protein expression for IL-1beta, TNFalpha and their receptors in non-degenerate, degenerate and herniated human IVDs. IL-1beta gene expression was observed in a greater proportion of IVDs than TNFalpha (79% versus 59%). Degenerate and herniated IVDs displayed higher levels of both cytokines than non-degenerate IVDs, although in degenerate IVDs higher levels of IL-1beta gene expression (1,300 copies/100 ng cDNA) were observed compared to those of TNFalpha (250 copies of TNFalpha/100 ng cDNA). Degenerate IVDs showed ten-fold higher IL-1 receptor gene expression compared to non-degenerate IVDs. In addition, 80% of degenerate IVD cells displayed IL-1 receptor immunopositivity compared to only 30% of cells in non-degenerate IVDs. However, no increase in TNF receptor I gene or protein expression was observed in degenerate or herniated IVDs compared to non-degenerate IVDs. We have demonstrated that although both cytokines are produced by human IVD cells, IL-1beta is expressed at higher levels and in more IVDs, particularly in more degenerate IVDs (grades 4 to 12). Importantly, this study has highlighted an increase in gene and protein production for the IL-1 receptor type I but not the TNF receptor type I in degenerate IVDs. The data thus suggest that although both cytokines may be involved in the pathogenesis of IVD degeneration, IL-1 may have a more significant role than TNFalpha, and thus may be a better target for therapeutic intervention.     

Law, evolution and the brain: applications and open questions

This paper discusses several issues at the intersection of law and brain science. It focuses principally on ways in which an improved understanding of how evolutionary processes affect brain function and human behaviour may improve law's ability to regulate behaviour. It explores sample uses of such 'evolutionary analysis in law' and also raises questions about how that analysis might be improved in the future. Among the discussed uses are: (i) clarifying cost-benefit analyses; (ii) providing theoretical foundation and potential predictive power; (iii) assessing comparative effectiveness of legal strategies; and (iv) revealing deep patterns in legal architecture. Throughout, the paper emphasizes the extent to which effective law requires: (i) building effective behavioural models; (ii) integrating life-science perspectives with social-science perspectives; (iii) considering the effects of brain biology on behaviours that law seeks to regulate; and (iv) examining the effects of evolutionary processes on brain design.     

Analysis of major repetitive DNA sequences in the dog (Canis familiaris) genome

A systematic screening and analysis of repeated DNA sequences from a dog genomic library composed of small DNA inserts enabled us to characterize abundant canine repetitive DNA families. Four main families were identified: i) a group of highly repeated tRNA-derived short interspersed repetitive DNA elements (tRNA-SINEs); ii) another type of SINE-like element that was mainly found inserted into long interspersed repetitive elements (LINEs); iii) LINEs of the L1 type; and iv) satellite or satellite-like DNA. Surprisingly, no SINEs derived from 7SL RNA were found in the dog genome. These data should help in the analysis of canine DNA sequences and in the design of canine genome mapping reagents. Received: 4 November 1998 / Accepted: 2 February 1999     

Rationally designed neuropeptide antagonists: A novel approach for generation of environmentally friendly insecticides

We report our approach for the generation of a novel type of putative insecticides based on backbone cyclic peptidomimetic antagonists of insect neuropeptides using pheromone biosynthesis activating neuropeptide (PBAN) as a model. This approach, called the backbone cyclic neuropeptide based antagonist (BBC-NBA), includes the following steps: (i) elucidation of the active sequence of the chosen insect neuropeptide; (ii) disclosure of a lead antagonist based on the sequence found in step (i); (iii) design and synthesis of backbone cyclic peptide libraries (cycloscan) based on the sequence of the lead antagonist; and (iv) design and synthesis of a peptidomimetic prototype insecticide. The BBC-NBA approach was applied to PBAN and led to the discovery of a potent linear lead antagonist and a potent backbone cyclic antagonist devoid of agnoistic activity which inhibited sex pheromone biosynthesis inHeliothis peltigera female moths.     

Cloning of genomic and cDNA for mouse isovaleryl-CoA dehydrogenase (IVD) and evolutionary comparison to other known IVDs

Isovaleryl-CoA dehydrogenase (IVD) is an intramitochondrial homotetrameric flavoenzyme that catalyzes the conversion of isovaleryl-CoA to 3-methylcrotonyl-CoA in the leucine catabolism pathway. Deficiency of IVD in humans causes isovaleric acidemia, which shows tremendous clinical variability for reasons that are unknown. To help better understand this disorder, we have cloned and sequenced the mouse IVD genomic and cDNAs. The mouse IVD gene spans approximately 17 kb and contains 12 coding exons organized identically to the human gene. It maps to mouse chromosome 2 in the area of band 2E4-E5, corresponding to the syntenic region of human chromosome 15. Mouse IVD predicted amino acid sequences are 95.8 and 89.6% identical to that of the rat and human sequences, respectively, with conservation of key functional residues. We have now identified IVD sequences from seven species. Comparison of these sequences shows that the rat and mouse proteins are the most closely related, both of which, in turn, share highest homology to human. All of the mammalian enzymes appear to be more closely related than any of the IVDs on other branches of the phylogram, while the fly and worm IVDs are the most divergent. The invertebrate IVDs are more closely related to the mammalian enzymes than to those from two plant species.     

The use of the alkaline comet assay with lymphocytes in human biomonitoring studies

We reviewed the data of 45 alkaline comet assay studies with lymphocytes published during the last three years with the objective of monitoring human exposure to genotoxic agents as a result of occupation, drug treatment, diseases or environmental pollution. The strengths of the studies were that: (i) a lot of data could be obtained within a relatively short period of time in a cost-effective manner, (ii) lymphocytes could be easily collected in a non-invasive way and proved to be good surrogate cells in that they picked up effects caused by agents with different cancer target organs and (iii) a remarkable concordance between comet assay and cytogenetic assay data was proved. However, our analysis revealed some shortcomings of the studies such as: (i) the inclusion of low number of study participants and bias in the number and gender of subjects between control and exposed groups, (ii) lack of qualitative and quantitative exposure data, (iii) lack of consideration of differences in physical activity and diet between control and exposed groups, (iv) difficulty in comparison of the studies due to lack of uniformity in the comet assay procedures such as duration of alkali unwinding and electrophoresis, slide scoring method and the metrics used to assess the extent of DNA damage and (v) controversy in the sensitivity of comet assay since it picked up DNA damage caused by agents such as wood dust, pesticides and hormone preparations which were found to be weak genotoxins or non-genotoxins in other tests, but gave inconsistent results with known mutagens/carcinogens such as tobacco smoke. We feel that for the alkaline comet assay to be an important tool in human biomonitoring studies, serious consideration should be given to the flaws in the design and performance of the assay.     

Using anticipatory life cycle assessment to enable future sustainable construction

The built environment is the largest single emitter of CO₂ and an important consumer of energy. Much research has gone into the improved efficiency of building operation and construction products. Life Cycle Assessment (LCA) is commonly used to assess existing buildings or building products. Classic LCA, however, is not suited for evaluating the environmental performance of developing technologies. A new approach, anticipatory LCA (a‐LCA), promises various advantages and can be used as a design constraint during the product development stage. It helps overcome four challenges: (i) data availability, (ii) stakeholder inclusion, (iii) risk assessment, and (iv) multi‐criteria problems. This article's contribution to the line of research is twofold: first, it adapts the a‐LCA approach for construction‐specific purposes in theoretical terms for the four challenges. Second, it applies the method to an innovative prefabricated modular envelope system, the CleanTechBlock (CTB), focusing on challenge (i). Thirty‐six CTB designs are tested and compared to conventional walls. Inclusion of technology foresight is achieved through structured scenario analysis. Moreover, challenge (iv) is tackled through the analysis of different environmental impact categories, transport‐related impacts, and thickness of the wall assemblies of the CTB. The case study results show that optimized material choice and product design is needed to reach the lowest environmental impact. Methodological findings highlight the importance of context‐specific solutions and the need for benchmarking new products.     

Reactivity of Indole Derivatives Towards Oxygenated Radicals

The reactivity of a series of indole derivatives was assessed in the following systems: (i) oxidation of the indole derivatives induced by the thermolysis of 2,2′-azobis-(2-amidinopropane) (ABAP); (ii) oxidation of cumene induced by the thermolysis of 2,2′-azobis-(2-methyl propionitrile) (AIBN); (iii) lysozyme inacti vation induced by the thermolysis of ABAP and (iv) brain homogenate autoxidation.

In systems (ii) to (iv), addition of the indole derivatives decreases the rate of the process. The data obtained indicate that common factors (i.e., oxidation potential and presence of N-H bonds) control the reactivity of the indole derivatives in the four systems considered. However, in the brain homogenate autoxidation, hydrophobicity is an additional factor that affects the efficiency of antioxidants, as illustrated by Q_1/2 values (the concentration of additive required to decrease the autoxidation rate to one half that observed in the absence of additive) of 0.1 mM and ? 8 mM for 3-methylindole and tryptophan, respectively.     

Genotoxicity and carcinogenicity testing of 1,2-dibromopropane and 1,1,3-tribromopropane in comparison to 1,2-dibromo-3-chloropropane

The activities of 1,2-dibromopropane (DBP) and 1,1,3-tribromopropane (TBP) were studied in seven genotoxicity assays, (i) SOS-induction inE. coli, (ii) DNA repair in primary rat hepatocyte culture, (iii) theSalmonella/microsome assay, (iv) a host-mediated assay usingSalmonella, (v) the somatic mutation and recombination assay inDrosophila melanogaster, (vi) HGPRT-mutagenesis assay in ARL 18 cells, and (vii) micronucleus formation assay in mouse polychromatophylic erythrocytes (PCE), forestomach (FS), glandular stomach (GS), duodenum (D), jejunum (J), cecum (C) and liver (L). The halopropanes were also tested for tumor formation in the fishDanio rerio. DBP was active in assays (ii), (v), (vii FS) and (vii L). TBP was positive in assays (ii) and (iii), strongly positive in (vii L) and borderline positive in (iv). However, neither DBP nor TBP induced tumors in fish, in contrast to the carcinogenic 1,2-dibromo-3-chloropropane. The genotoxicity and potential carcinogenicity of DBP and TBP in mammals is discussed.Abbreviations 2-AA 2-aminoanthracene - DBCP 1,2-dibromo-3-chloropropane - DBP 1,2-dibromopropane - HGPRT hypoxanthine-guanine phosphoribosyl transferase - i.p. intraperitoneal(ly) - NQO 4-nitro-quinoline-1-oxide - PCE polychromatic erythrocytes - TBP 1,1,3-tribromopropane - WME Williams' medium E     

Oxime-based linker libraries as a general approach for the rapid generation and screening of multidentate inhibitors

The described oxime-based library protocol provides detailed procedures for the linkage of aminooxy functionality with aldehyde building blocks that result in the generation of libraries of multidentate inhibitors. Synthesis of inhibitors for protein tyrosine phosphatases (PTPs) and antagonists directed against the human tumor susceptibility gene 101 (TSG101) are shown as examples. Three steps are involved: (i) the design and synthesis of aminooxy platforms; (ii) tethering with aldehydes to form oxime-based linkages with sufficient purity; and (iii) direct in vitro biological evaluation of oxime products without purification. Each coupling reaction is (i) performed in capped microtubes at room temperature (20-23 °C); (ii) diluted for inhibitory evaluation; and (iii) screened with targets in microplates to provide IC(50) or K(d) values. The synthesis of the aminooxy platforms takes 3-5 d; tethering with the aldehydes takes 24 h; and inhibition assay of enzymes and protein-protein interactions takes 30 min and 2 h, respectively.     

The Second International Standard for somatropin (recombinant DNA-derived human growth hormone): preparation and calibration in an international collaborative study.

A preparation of somatropin (recombinant DNA-derived human growth hormone) was prepared as lyophilised ampoules according to WHO procedures for international biological standards. The candidate preparation (98/574) was evaluated in an international collaborative study (16 laboratories, nine countries), with the following aims: (i) to determine the suitability of the preparation to serve as the International Standard for somatropin by studying its performance in the current range of physico-chemical and biological assay methods employed for somatropin; (ii) to assign a content in terms of the existing (first) International Standard for somatropin, using the currently recognised assay procedure (Size Exclusion High Performance Liquid Chromatography, SE HPLC); (iii) to confirm the specific biological activity of the candidate preparation; (iv) to confirm the stability of the candidate preparation. On the basis of the collaborative study WHO agreed that: the preparation in ampoules coded 98/574 is suitable to serve as the next WHO International Standard for somatropin; the preparation in ampoules coded 98/574 should be established as the second International Standard for somatropin, with a defined ampoule content of 1.95 mg total somatropin plus somatropin-related proteins per ampoule; the specific activity of the preparation should be defined as 3.0 IU/mg somatropin.     

Interleukin-1 receptor antagonist delivered directly and by gene therapy inhibits matrix degradation in the intact degenerate human intervertebral disc: an <Emphasis Type="Italic">in situ</Emphasis> zymographic and gene therapy study

Data implicate IL-1 in the altered matrix biology that characterizes human intervertebral disc (IVD) degeneration. In the current study we investigated the enzymic mechanism by which IL-1 induces matrix degradation in degeneration of the human IVD, and whether the IL-1 inhibitor IL-1 receptor antagonist (IL-1Ra) will inhibit degradation. A combination of in situ zymography (ISZ) and immunohistochemistry was used to examine the effects of IL-1 and IL-1Ra on matrix degradation and metal-dependent protease (MDP) expression in explants of non-degenerate and degenerate human IVDs. ISZ employed three substrates (gelatin, collagen, casein) and different challenges (IL-1β, IL-1Ra and enzyme inhibitors). Immunohistochemistry was undertaken for MDPs. In addition, IL-1Ra was introduced into degenerate IVD explants using genetically engineered constructs. The novel findings from this study are: IL-1Ra delivered directly onto explants of degenerate IVDs eliminates matrix degradation as assessed by multi-substrate ISZ; there is a direct relationship between matrix degradation assessed by ISZ and MDP expression defined by immunohistochemistry; single injections of IVD cells engineered to over-express IL-1Ra significantly inhibit MDP expression for two weeks. Our findings show that IL-1 is a key cytokine driving matrix degradation in the degenerate IVD. Furthermore, IL-1Ra delivered directly or by gene therapy inhibits IVD matrix degradation. IL-1Ra could be used therapeutically to inhibit degeneration of the IVD.     

A new phylogenetic tribal classification of the grape family (Vitaceae)

Vitaceae (the grape family) consist of 16 genera and ca. 950 species primarily distributed in tropical regions. The family is well‐known for the economic importance of grapes, and is also ecologically significant with many species as dominant climbers in tropical and temperate forests. Recent phylogenetic and phylogenomic analyses of sequence data from all three genomes have supported five major clades within Vitaceae: (i) the clade of Ampelopsis, Nekemias, Rhoicissus, and Clematicissus; (ii) the Cissus clade; (iii) the clade of Cayratia, Causonis, Cyphostemma, Pseudocayratia, Tetrastigma, and an undescribed genus “Afrocayratia”; (iv) the clade of Parthenocissus and Yua; and (v) the grape genus Vitis and its close tropical relatives Ampelocissus, Pterisanthes and Nothocissus, with Nothocissus and Pterisanthes nested within Ampelocissus. Based on the phylogenetic and morphological (mostly inflorescence, floral and seed characters) evidence, the new classification places the 950 species and 16 genera into five tribes: (i) tribe Ampelopsideae J.Wen & Z.L.Nie, trib. nov. (47 species in four genera; Ampelopsis, Nekemias, Rhoicissus and Clematicissus); (ii) tribe Cisseae Rchb. (300 species in one genus; Cissus); (iii) tribe Cayratieae J.Wen & L.M.Lu, trib. nov. (370 species in seven genera; Cayratia, Causonis, “Afrocayratia”, Pseudocayratia, Acareosperma, Cyphostemma and Tetrastigma); (iv) tribe Parthenocisseae J.Wen & Z.D.Chen, trib. nov. (ca. 16 spp. in two genera; Parthenocissus and Yua); and (v) tribe Viteae Dumort. (ca. 190 species in two genera; Ampelocissus and Vitis).     

Synthesis of 6I-amino-6I-deoxy-2I-VII,3I-VII-tetradeca-O-methyl-cyclomaltoheptaose

The preparation of 6(I)-amino-6(I)-deoxy-2(I-VII),3(I-VII)-tetradeca-O-methyl-cyclomaltoheptaose is reported. Two different routes (A and B), both starting from beta-cyclodextrin (betaCD), have been examined. Route A involved: (i) synthesis of heptakis(6-O-tert-butyldimethylsilyl)-betaCD from betaCD; (ii) permethylation of the secondary hydroxyl groups with methyl iodide and sodium hydride; (iii) desilylation of the primary hydroxyls with ammonium fluoride; (iv) monotosylation at O-6 position of per-(2,3-O-methyl)-betaCD; (5) nucleophilic replacement of the tosyl group with azide anion; (v) reduction of the azido group by catalytic transfer hydrogenation using hydrazine hydrate in the presence of Pd/C in methanol/water. Route B started from the known 6(I)-monoazido-6(I)-monodeoxy-beta-CD (two steps from beta-CD) and entailed: (i) protection of the remaining primary hydroxyls using tert-butyldimethylsilylchloride (TBDMSCl); (ii) exhaustive methylation of the secondary hydroxyls with methyl iodide and sodium hydride; (iii) removal of the TBDMS protecting groups with ammonium fluoride; (iv) reduction of the azido group as above. Route A was found to be less convenient than Route B due to the inherent difficulty of controlling the monotosylation of per-(2,3-O-methyl)-betaCD.     

Adaptation of the Nelson-Somogyi reducing-sugar assay to a microassay using microtiter plates

The Nelson-Somogyi assay for reducing sugars was adapted to microtiter plates. The primary advantages of this modified assay are (i) smaller sample and reagent volumes, (ii) elimination of boiling and filtration steps, (iii) automated measurement with a dual-wavelength scanning TLC densitometer, (iv) increased range and reproducibility, and (v) automated colorimetric readings by reflectance rather than absorbance.