Method For Analysis Of Carbohydrates Research Articles

The Determination of Carbohydrates by High-Performance Anion-Exchange Chromatography Coupled with Pulsed Amperometric Detection (HPAEC-PAD).

Chromatography techniques are widely used to separate, identify, and quantify molecules depending on their physicochemical properties. Standard methods range from simple size exclusion to separation based on affinity or ion exchange. Here, we present a method for the direct analysis of carbohydrates in Magnaporthe oryzae using high-performance anion-exchange chromatography (HPAEC) coupled with pulsed amperometric detection (PAD). The combination of HPAEC with PAD provides the highest selectivity and sensitivity with minimal sample preparation and cleanup time. Utilizing our HPAEC-PAD approach, we obtain reliable and highly reproducible determination of carbohydrates produced as osmotic stress response by M. oryzae. Thus, the method described provides a fast, precise, and comprehensive analysis of stress-dependent metabolic adjustments of carbohydrates not only relevant for M. oryzae but also applicable in other systems.

Analysis of Macro Nutrient on Rono Fish (Xenopoecilus oophorus) Freshwater

Fish are assumed to have high nutritional value with mineral content, vitamins, fats, and proteins containing essential amino acids that are needed for growth and human intelligence. One type of them is rono fish (Xenopoecilus oophorus) freshwater which is an endemic fish that comes from lake Poso, Central Sulawesi and still found in traditional markets of Tentena city, but the local community is less interested in this type of fish and the lack of literature that informs about the nutritional content of rono fish. The research aims to determine the macronutrient content in rono fish. The nutrient content analyzed in fish was on carbohydrates, proteins, and fats by using the anthrone sulfate method for carbohydrate analysis, Kjeldahl method for protein, and for fat analysis using the soxhlet method. The results of the analysis obtained that macronutrient in freshwater rono fish was followed carbohydrate was 9.26%, the protein was 37.79% and fat was 11.92%.

Raman Optical Activity and Raman spectroscopy of carbohydrates in solution.

This comprehensive study on selected 14 carbohydrates in water solution is an extension of previously published one focused only on solid state analysis. Here, Raman spectroscopy was used as a dedicated method for analysis of carbohydrates in solution, both using a normal effect (RS) and its chiral analogue: Raman Optical Activity spectroscopy (ROA). The compounds were selected as biologically important and representative of all groups: monosaccharides, disaccharides, trisaccharides, cyclodextrines and polysaccharides. RS and ROA spectra are presented together with an expanded discussion on various structures and conformations of studied carbohydrates in the solution taking into account particular regions, i.e. (1) low wavenumber region (250-600 cm-1), (2) anomeric region (600-950 cm-1), (3) fingerprint region (950-1200 cm-1) and (4) CH2and COH deformations region (1200-1500 cm-1). So, the following information can be obtained about: (1) the absolute configuration of the anomeric centre; (2) the configuration of the anomeric centre and the orientation of the anomeric hydroxyl group; (3) the ring structures and the relative orientation of substituents and (4) the conformation of the exocyclic CH2OH (4), respectively. Raman spectroscopy and Raman Optical Activity were shown as unique tools to study complex structures of carbohydrates.

Triple-Internal Standard Based Glycan Structural Assignment Method for Capillary Electrophoresis Analysis of Carbohydrates.

Despite the ever growing use of capillary electrophoresis in biomedical research and the biopharmaceutical industry, the development of data interpretation methods is lagging behind. In this paper we report the design and implementation of a coinjected triple-internal standard method to alleviate the need of an accompanying run of the maltooligosaccharide ladder for glucose unit (GU) calculation. Based on the migration times of the coinjected standards of maltose, maltotriose, and maltopentadecaose (bracketing the peaks of interest), a data processing approach was designed and developed to set up a virtual ladder that was used for GU calculation. The data processing was tested in terms of the calculated GU values of human IgG glycans, and the resulting relative standard deviation was ≤1.07%. This approach readily supports high-throughput capillary electrophoresis systems by significantly speeding up the processing time for glycan structural assignment.

Analysis of oligosaccharides in lignocellulosic biomass hydrolysates by high-performance anion-exchange chromatography coupled with mass spectrometry (HPAEC–MS)

The carbohydrate composition of lignocellulosic biomass hydrolysates is highly complex. High performance anion exchange chromatography coupled with pulsed amperometric detection (HPAEC–PAD), a widely used method for carbohydrate analysis, provides limited chemical information on the detected peaks. To improve the detection and increase the chemical information of the carbohydrates, HPAEC was coupled with mass spectrometry (MS). Using a pooled hydrolysate sample, it was shown that HPAEC–MS can separate and detect many oligosaccharides in one experimental run based on retention time and mass. The method was validated on its linearity, reproducibility and response factors. The analysis of a group of different biomass hydrolysates revealed that remaining disaccharides was the bottleneck of the hydrolysis process. As an analytical tool, HPAEC–MS provides information for the improvement of hydrolysate pretreatment method and enzyme cocktail quality. Besides, the consumption ability of microbial host strains for various mono- and oligosaccharides in hydrolysates can be assessed.

The usefulness of hydrazine derivatives for mass spectrometric analysis of carbohydrates

Over the last years, extensive studies have evaluated glycans from different biological samples and validated the importance of glycosylation as one of the most important post-translational modifications of proteins. Although a number of new methods for carbohydrate analysis have been published and there has been significant progress in their identification, the development of new approaches to study these biomolecules and understand their role in living systems are still vivid challenges that intrigue glycobiologists. In the last decade, the success in analyses of oligosaccharides has been driven mainly by the development of innovative, highly sensitive mass spectrometry techniques. For enhanced mass spectrometry detection, carbohydrate molecules are often derivatized. Besides, the type of labeling can influence the fragmentation pattern and make the structural analysis less complicated. In this regard, in 2003 we introduced the low scale, simple non-reductive tagging of glycans employing phenylhydrazine (PHN) as the derivatizing reagent. PHN-labeled glycans showed increased detection and as reported previously they can be analyzed by HPLC, ESI, or MALDI immediately after derivatization. Under tandem mass spectrometry conditions, PHN-derivatives produced useful data for the structural elucidation of oligosaccharides. This approach of analysis has helped to reveal new isomeric structures for glycans of known/unknown composition and has been successfully applied for the profiling of N-glycans obtained from serum samples and cancer cells. The efficacy of this labeling has also been evaluated for different substituted hydrazine reagents. This review summarizes all types of reducing-end labeling based on hydrazone-linkage that have been used for mass spectrometric analyses of oligosaccharides. This review is also aimed at correcting some past misconceptions or interpretations reported in the literature.

New Avenue for Mid-UV-Range Detection of Underivatized Carbohydrates and Amino Acids in Capillary Electrophoresis

Capillary electrophoresis (CE) appeared as an interesting alternative to chromatographic methods for carbohydrate analysis, but it can be difficult to implement, because of the lack of easily ionizable functions and chromophore groups. Recently, a promising method was proposed by Rovio et al. for the CE separation under extremely high alkaline conditions of neutral carbohydrates under their alcoholate form and their direct UV detection [Rovio et al. Electrophoresis 2007, 28, 3129-3135; and Rovio et al. J. Chromatogr. A 2008, 1185, 139-144], which is claimed to be due to the absorption of enediolate at 270 nm. Even so, most of the detected compounds in Rovio's paper (for example, sucrose) cannot give such enediolate, lacking a carbonyl group. In this work, a deeper insight was paid to the understanding of detection mechanism. In effect, unusual detection phenomena were observed in comparing reducing and nonreducing carbohydrate behaviors, which pointed to the existence of photochemical reactions in the detection window. A more systematic study of the influence of many parameters (carbohydrate nature, electrolyte pH, residence time in the detection window, and capillary diameter) was undertaken. In addition to this, most of this work was performed under cathodic (reversed) electro-osmotic flow conditions (using Polybrene-modified capillaries), to obtain much faster separations than under Rovio's conditions. This study also opens up new avenues for the detection in mid-UV range of non-UV-absorbing compounds bearing reducing moieties, such as amino acids.

The Beginning of Icodextrin

Correspondence to: C.D. Mistry, Peterborough District Hospital, Thorpe Road, Peterborough PE3 6DA United Kingdom. chandra.mistry@btopenworld.com Received 27 October 2009; accepted 9 September 2010. In the history of peritoneal dialysis (PD), 1976 marked a significant step forward when Popovich and Moncrief revolutionized the practice by introducing the concept of equilibration PD and extending the duration of dwell time to 4 – 10 hours (1). Despite this fundamental change in the practice of PD, the basic principle used to generate osmotic forces across the peritoneum remained unaltered. This principle relied on the traditional concept of osmotic flow across an “ideal” semipermeable membrane, necessitating making dialysis solution hypertonic to plasma with the addition of glucose as osmotic agent. Unfortunately, not being an “ideal” semipermeable membrane but being partially permeable to solutes, the peritoneum allows rapid absorption of glucose with progressive dissipation of the osmotic gradient and ultrafiltration of short duration. While this is of little significance during short dwell exchanges (30 – 60 minutes’ dwell in intermittent PD), it is not the case for long exchanges, such as in continuous ambulatory PD (CAPD) and automated PD, where reabsorption of initially ultrafiltered peritoneal fluid occurs. In addition, the continuous daily absorption of glucose aggravates longterm metabolic complications, including hyperlipidemia and obesity (2,3). Even as early as the 1980s there was clear recognition for an alternative osmotic agent that would minimize metabolic derangements and provide the ultrafiltration profile to suit long dwell exchanges. A range of different macromolecules was evaluated based on the simplistic concept that large molecular weight (MW) agents are less readily absorbed through the peritoneum and are likely to produce sustained ultrafiltration while reducing metabolic complications. Early investigations clearly identified the problems associated with use of nonphysiological hyperviscous macromolecules and defined the need for a neutral substance that is soluble, nonallergenic, and readily metabolized (3). Glucose polymer (GP), derived from hydrolyzed cornstarch, seemed a natural contender and several groups already held patents of diverse MW fractions. Among them, the Abbott group led the way by studying a narrow MW fraction (MW 1000 Da) in both animals (4) and humans (5). In Manchester, we were well placed to explore the potential of this novel agent as considerable experience had been developed while investigating GP (Caloreen) as an intravenous high-energy nutrient source in the management of patients with renal (6) and hepatic failure (7). We worked closely in collaboration with Jerry Milner, the holder of the patent for Caloreen, Fisons Pharmaceutical, who had established expertise in fractionating GP technology, and J. Fox, Department of Biochemistry, University of Birmingham, who had extensive experience in methods of carbohydrate analysis. For the initial clinical studies carried out in July 1983, we utilized a readily available dextrin formulation (Caloreen) with a bimodal MW distribution consisting of a 67% “low” MW fraction (chain length 12 glucose units); weight average MW (Mw) was 7000 Da and number average MW (Mn) was 960 Da. In contrast to glucose solution, predicting osmotic performance of a polydisperse GP sample with a relatively unknown peritoneal permselectivity proved difficult. Our preliminary studies suggested that 5% (52 mmol/L) and 10% (104 mmol/L) GP solutions were probably comparable to 1.36% (76 mmol/L) and 3.86% (214 mmol/L) glucose respectively (8). The first formal Phase 1 study, using solutions containing 5% (52 mmol/L) and 10% (104 mmol/L) of this GP formulation over a 6-hour dwell, was exciting and

Involvement of selectin-like carbohydrate binding specificity in human gamete interaction.

The recognition of carbohydrate epitopes by complimentary protein receptors has been shown to be a critical factor in gamete interaction in many different animal species. In this study it was hypothesized that, in the human, gamete binding requires an interaction between selectin ligands on the zona pellucida and putative egg binding proteins on the sperm surface. The hemizona assay (a unique internally controlled bioassay that evaluates tight binding of sperm to the zona) and advanced methods of carbohydrate analysis were used to test this hypothesis. From these tests it was shown that oligosaccharide recognition is also required for initial human gamete binding. This study suggests the existence of distinct egg binding proteins on human sperm that can bind to selectin ligands. Additionally, the results suggest a possible convergence in the types of carbohydrate sequences recognized during initial human gamete binding and immune/inflammatory cell interactions. Glycoconjugates that manifest selectin-ligand activity and that express specific carbohydrate epitopes have potent contraceptive and immunosuppressive effects. Such specific oligosaccharide sequences may provide an appropriate recognition signal for embryo development and protection.

A versatile method for stable carbon isotope analysis of carbohydrates by high‐performance liquid chromatography/isotope ratio mass spectrometry

We have developed a method to analyze stable carbon isotope ((13)C/(12)C) ratios in a variety of carbohydrates using high-performance liquid chromatography/isotope ratio mass spectrometry (HPLC/IRMS). The chromatography is based on strong anion-exchange columns with low strength NaOH eluents. An eluent concentration of 1 mM resulted in low background signals and good separation of most of the typical plant neutral carbohydrates. We also show that more strongly bound carbohydrates such as acidic carbohydrates can be separated by inclusion of NO(3) (-) as an inorganic pusher ion in the eluent. Analyses of neutral carbohydrate concentrations and their stable carbon isotope ratios are shown for plant materials and marine sediment samples both at natural abundance and for (13)C-enriched samples. The main advantage of HPLC/IRMS analysis over traditional gas chromatography based methods is that no derivatization is needed resulting in simple sample treatment and improved accuracy and reproducibility.

Evaluation of oligosaccharide methods for carbohydrate analysis in a fully human monoclonal antibody and comparison of the results to the monosaccharide composition determination by a novel calculation

Carbohydrates can change a drug's properties including solubility, affinity towards antigen, pharmacokinetics and pharmacodynamics. Due to this importance, carbohydrate composition is utilized as a parameter to evaluate a drug candidate's quality. In this study, the compositional monosaccharides of a drug candidate are measured by HPAEC-PAD, while the oligosaccharides are studied by HPAEC-PAD, CE-LIF and LC–MS. The advantages and limitations of these various approaches for oligosaccharide analysis are reviewed in this work. While the methods used for oligosaccharide analysis are well established we have devised a new and novel calculation for determining monosaccharide content using the relative percentages of the N-glycans. This calculation was used to evaluate the accuracy of the oligosaccharide determination methods by comparison of the N-glycan data to the experimental monosaccharide data. The results obtained from this novel calculation demonstrate that the relative abundance of carbohydrates as determined from these various approaches are consistent.

Ionic Liquid Matrixes Optimized for MALDI-MS of Sulfated/Sialylated/Neutral Oligosaccharides and Glycopeptides

1,1,3,3-tetramethylguanidium (TMG) salt of alpha-cyano-4-hydroxycinnamic acid (CHCA) (G(2)CHCA) was reported by Tatiana et al. as a useful ionic liquid matrix (ILM) for sulfated oligosaccharides to suppress the loss of sulfate groups. However, the report mainly referred to positive ion spectra only and amounts of 10 pmol or more of the analyte were used. Herein, we demonstrated highly sensitive detection of sulfated/sialylated/neutral oligosaccharides and preferential ionization of glycopeptides by optimizing a newly synthesized ILM: TMG salt of p-coumaric acid (G(3)CA) and the existing G(2)CHCA in both positive and negative ion extraction modes. Sulfated oligosaccharides were detected with high sensitivity (e.g., 1 fmol) in both ion extraction modes, and the dissociation of sulfate groups was suppressed especially using G(3)CA. Sialylated and neutral oligosaccharides were also detected with high sensitivity (e.g., 1 fmol) with positive ion extraction while the dissociation of sialic acids was suppressed especially using G(3)CA. Additionally, glycopeptide ions were detected preferentially using the ILMs among the digest of a glycoprotein, ribonuclease B, in both ion extraction modes but particularly in the negative ion mode. As a result, the use of optimized ILMs provides an effective method for carbohydrate analysis due to the highly sensitive soft-ionization achieved in both ion extraction modes as well as the homogeneity of analyte-matrix mixtures.

Carbohydrate analysis by a phenol–sulfuric acid method in microplate format

Among many colorimetric methods for carbohydrate analysis, the phenol–sulfuric acid method is the easiest and most reliable method. It has been used for measuring neutral sugars in oligosaccharides, proteoglycans, glycoproteins, and glycolipids. This method is used widely because of its sensitivity and simplicity. In its original form, it required 50–450 nmol of monosaccharides or equivalent for analysis and thus is inadequate for precious samples. A scaled-down version requiring only 10–80 nmol of sugars was reported previously. We have now modified and optimized this method to use 96-well microplates for high throughput, to gain greater sensitivity, and to economize the reagents. This modified and optimized method allows longer linear range (1–150 nmol for Man) and excellent sensitivity. Moreover, our method is more convenient, requiring neither shaking nor covering, and takes less than 15 min to complete. The speed and simplicity of this method would make it most suitable for analyses of large numbers of samples such as chromatographic fractions.

Isotope tag method for quantitative analysis of carbohydrates by liquid chromatography–mass spectrometry

We have previously demonstrated that liquid chromatography/mass spectrometry equipped with a graphitized carbon column (GCC-LC/MS) is useful for the structural analysis of carbohydrates in a glycoprotein. Here, we studied the monosaccharide composition analysis and quantitative oligosaccharide profiling by GCC-LC/MS. Monosaccharides were labeled with 2-aminopyridine and then separated and monitored by GCC-LC/MS in the selective ion mode. The use of tetradeuterium-labeled pyridylamino (d 4-PA) monosaccharides as internal standards, which were prepared by the tagging of standard monosaccharides with hexadeuterium-labeled 2-aminopyridine (d 6-AP), afforded a good linearity and reproducibility in ESIMS analysis. This method was successfully applied to the monosaccharide composition analysis of model glycoproteins, fetuin, and erythropoietin. For quantitative oligosaccharide profiling, oligosaccharides released from an analyte and a standard glycoprotein were tagged with d 0- and d 6-AP, respectively, and an equal amount of d 0- and d 4-PA oligosaccharides were coinjected into GCC-LC/MS. In this procedure, the oligosaccharides that existed in either analyte or a standard glycoprotein appeared as single ions, and the oligosaccharides that existed in both analyte and a standard glycoprotein were detected as paired ions. The relative amount of analyte oligosaccharides could be determined on the basis of the analyte/internal standard ion-pair intensity ratio. The quantitative oligosaccharide profiling enabled us to make a quantitative and qualitative comparison of glycosylation between the analyte and standard glycoproteins. The isotope tag method can be applicable for quality control and comparability assessment of glycoprotein products as well as the analysis of glycan alteration in some diseases.

Application of curve fitting in thin-layer chromatography-flame ionization detection analysis of the carbohydrate fraction in marine mucilage and marine snow samples from Italian seas.

This paper presents a thin-layer chromatographic-flame ionization detection (TLC-FID) procedure to characterize the carbohydrate fraction of marine mucilage and marine snow samples from the Italian Seas. The identification of the different carbohydrate subfractions is supported by the application of a deconvolution procedure based on a new mathematical function for describing chromatographic peaks and enhancing their resolution. The joint-approach TLC-FID analysis and deconvolution procedure allows for the characterization of the carbohydrate fraction of the marine samples in a single step without using the different derivatization procedures requested by the most common gas chromatography and high-performance liquid chromatographic methods for carbohydrate analysis. In fact, the results obtained by the TLC-FID procedure show that different neutral, uronic acid, and aminosugar subfractions can be present simultaneously in these samples. Moreover, the results support some hypotheses about the causes of the presence of mucilages in the Italian Seas.

Biochemical and functional characterization of the human zona pellucida

A successful interaction between spermatozoa and the zona pellucida is critical for fertilization. This biological step reflects multiple sperm functions, including the acquisition and completion of capacitation, recognition and binding to specific zona pellucida receptors, and induction of the physiological acrosome reaction. The recognition of carbohydrate sequences by complimentary receptors has been demonstrated in gamete interaction in different animal species. It has been proposed that, in the human, sperm binding to the zona pellucida requires a ‘selectin-like’ interaction. The hemizona assay (a unique internally controlled bioassay that evaluates tight binding of human spermatozoa to the homologous zona pellucida) and advanced methods of carbohydrate analysis have been used to test this hypothesis. Compelling evidence exists to demonstrate that oligosaccharide recognition is also required for specific, tight human gamete binding. The induction of the acrosome reaction using the physiological inducers, i.e. the zona pellucida and progesterone, was also examined. It has also been demonstrated that there is a priming effect of the steroid on the acrosome reaction inducing capacity of the zona pellucida. These studies may allow for a better understanding of human gamete interaction in physiological and pathological situations.

Analysis of N-linked oligosaccharides: progress towards the characterisation of glycoprotein-linked carbohydrates

The covalent attachment of carbohydrate to proteins is a very common co- or post-translational event in the biosynthesis of glycoproteins. The type and heterogeneity of these oligosaccharides can affect a range of physico-chemical and biological properties of a glycoprotein. Thus the development of sensitive, reliable and robust analytical methods for carbohydrate analysis is important in the pharmaceutical industry, especially in the recombinant production of experimental and therapeutic glycoproteins. In this report we have reviewed methodology for the in-gel enzymatic release of N-linked oligosaccharides from glycoproteins separated by electrophoresis. These oligosaccharides are derivatised by reductive amination using 3-acetamido-6-aminoacridine (AA-Ac), a novel, highly fluorescent probe. A major advantage of this technique is that glycan derivatives are amenable to analysis by an array of chromatographic and mass spectrometric methods, allowing the resolution and characterisation of a wide variety of glycan structures. It is hoped that in due course the methodology described will be applied to proteomics studies, especially in identifying the role of carbohydrate in protein function and disease.

High-Resolution Polyacrylamide Gel Electrophoresis of Carbohydrates Derivatized with a Visible Dye

A technique for carbohydrate analysis that is both inexpensive and easily performed is currently unavailable. In this communication we address the problem and have outlined a method for labeling saccharides with a visible dye, 4-amino-1,1′-azobenzene-3,4′-disulfonic acid, which has an absorption maximum of 489 nm and an extinction coefficient of 37,615, to facilitate visible detection at low levels. The visible dye was coupled by a reductive amination to different sugars. The sugar–dye adducts were then separated by electrophoresis on alkaline polyacrylamide gels. The gels were scanned with a densitometer, and visible sugar–dye adducts were qualitatively analyzed by identifying them according to their mobilities. The sugar–dye adducts were quantified by determining their densitometric volume. The kinetics of the reductive amination reactions, performed at 37°C, were different for each of three saccharides tested. The rate constants for glucose and fucose were 1.31 times greater and 1.8 times greater, respectively, than that of maltotriose. The reductive amination reactions were essentially complete after approximately 16 h under the given experimental conditions. A linear dose–response relationship was observed between the amount of sugar (monosaccharide, trisaccharide, or heptasaccharide) in the reductive amination reaction. The quantity of saccharide–dye adduct that could be visually detected for glucose, maltotriose, and maltoheptaose, was 25, 25, and 50 nmol, respectively. Sugar–dye adducts were separated from one another by varying the acrylamide concentration in the polyacrylamide gels. Sugar–dye adducts of monosaccharides, disaccharides, trisaccharides, and heptasaccharides were separated on alkaline 30% polyacrylamide gels with mobilities of 0.778, 0.667, 0.639, and 0.375. Adducts of glucose, fucose, galactose, and mannose were separated with mobilities of 0.844, 0.833, 0.820, and 0.810, respectively, on a 30 to 40% gradient polyacrylamide gel. Adducts of glucose and glucose derivatives were separated on a 35% polyacrylamide gel. This technique provides an inexpensive and easily performed method of carbohydrate analysis to laboratories that do not have the highly trained personnel nor the expensive equipment needed for other methods of carbohydrate analysis. The method is most applicable to research problems where sensitivity (20 pmol) is not a problem. The simplicity of the method also makes it easily incorporated into teaching laboratories.

Enzyme array-amperometric detection in carbohydrate analysis

The introduction of an enzyme array-electrochemical detection method for carbohydrate analysis is demonstrated by using two complex and one high mannose N-linked oligosaccharides. Instead of measuring the remaining uncleaved oligosaccharides in enzymatic digestion, released monosaccharides are directly quantified by pulsed amperometric detection at a gold electrode. The measured monosaccharide concentrations in combination with the enzyme array analysis provide structural characterization of oligosaccharides. The enzyme array-electrochemical detection method does not require any separation procedure or any prior labeling of oligosaccharides. However, this method is limited by the use of purified oligosaccharide samples and the nature of the enzyme array. The development of more sophisticated enzyme arrays relies upon the introduction of a bank of highly specific (bond, arm, aglycon) exoglycosidases.

Enzyme array‐amperometric detection in carbohydrate analysis

The introduction of an enzyme array–electrochemical detection method for carbohydrate analysis is demonstrated by using two complex and one high mannose N-linked oligosaccharides. Instead of measuring the remaining uncleaved oligosaccharides in enzymatic digestion, released monosaccharides are directly quantified by pulsed amperometric detection at a gold electrode. The measured monosaccharide concentrations in combination with the enzyme array analysis provide structural characterization of oligosaccharides. The enzyme array–electrochemical detection method does not require any separation procedure or any prior labeling of oligosaccharides. However, this method is limited by the use of purified oligosaccharide samples and the nature of the enzyme array. The development of more sophisticated enzyme arrays relies upon the introduction of a bank of highly specific (bond, arm, aglycon) exoglycosidases. ©1998 John Wiley & Sons, Inc. Biotechnol Bioeng 57: 545-551, 1998.