Determining carbohydrates in essential and non-essential                                            foodstuffs using ion c...
Figure 1                                                                                                                  ...
Figure 2Figure 2: Schematic diagram showing (a) inline dialysis cell (b) coupling to the sample changer and IC unit, and (...
Table 1                                                                                                           Figure 3...
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Determining carbohydrates in foodstuffs using IC


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Metrohm Solutions for determining carbohydrates in essential and non-essential
foodstuffs using ion chromatography

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Determining carbohydrates in foodstuffs using IC

  1. 1. Determining carbohydrates in essential and non-essential foodstuffs using ion chromatography Alfred Steinbach and Andrea Wille Herisau, Switzerland This article describes a straightforward ion chromatographic method that uses isocratic elution and pulsed amperometric detection (PAD) to sensitively determine water-soluble polyols and sugar alcohols as well as mono-, di-, and oligosaccharides in essential and non-essential foodstuffs. While carbohydrate determination of most foodstuffs requires only minimal sample preparation such as dilution and filtration, samples with strongly interfering matrices such as protein-containing dairy products have to be dialysed before injection. Carbohydrates or sacccharides, DNA. Because of the widespread use drawbacks, high-performance also called sugars, are the most and importance of carbohydrates, anion-exchange chromatography abundant organic molecules their determination is of considerable has increasingly been used for found in the biosphere. They interest in biological, environmental, carbohydrate determination. In include monosaccharides (e.g., clinical, and medical research. strongly alkaline mobile phases, glucose, fructose, xylose, or Additionally, quality control of sugar anions are separated on a mannose), disaccharides (e.g., foodstuffs assures consumers’ positively charged strong anion- sucrose, lactose, and maltose), top-quality carbohydrate supply. exchange resin. Subtle differences trisaccharides, oligosaccharides and The most commonly applied in pKa values of the hydroxyl groups polysaccharides. Carbohydrates analytical techniques for of the carbohydrates allow for an are synthesized in plants during determining carbohydrates are efficient separation of low-molecular photosynthesis using carbon 1 H-nuclear magnetic resonance saccharides. However, sensitive dioxide, water, and the energy from (NMR), fourier transform infrared and straightforward detection sunlight. They are one of the three spectroscopy (FT-IR), polyacrylamide of the separated carbohydrates macronutrients (proteins and fats gel electrophoresis (PAGE), mass has long been a challenge. As a being the other two) that provide the spectrometry (MS) together with result of missing chromophores human body with energy. Complex gas and liquid chromatography. and fluorophores, ultraviolet and polysaccharides are structural While spectroscopic methods fluorescence detectors cannot be elements in the cell walls of bacteria suffer from high instrumentation used and refractive index detection and plants and are stored for food costs and need highly skilled suffers from poor sensitivity and or structural support. In addition, operators, gas chromatographic the fact that gradient elution is not ribose and deoxyribose form part of methods require time-consuming applicable. Several post-column the structural framework of RNA and derivatizations. In view of these derivatization reactions have beenA Determining carbohydrates in foodstuffs using IC separation science — volume 3 issue 6
  2. 2. Figure 1 samples. Experimental Materials and Methods Sample preparation of malt extracts Instrumentation: is straightforward and only involves a Chromatographic equipment: 871 1:100 dilution. Afterwards the sample Advanced Bioscan, 838 Advanced IC can be directly injected. Sample Processor, 833 Advanced IC Liquid Handling Pump Unit, Results 833 Advanced IC Liquid Handling Glucose, fructose, and sucrose Dialysis Unit, 818 Advanced IC Pump. are primarily responsible for the Instrument control, data acquisition, foods` sweetness. As the latter two and processing were performed by sugars are not present at detectable Metrodata IC Net software (Metrohm concentrations [Figure 1], malt AG). extracts are only perceived half as Reagents and eluents: sweet as mainly sucrose-containing Carbohydrates were reagent grade products. Apart from glucose and purchased from Fluka (Sigma and maltose the investigated Aldrich, Buchs, Switzerland). All malt extract contains several standard solutions and eluents were maltooligosaccharides. prepared with deionized water with a The observed saccharide profile of specific resistance higher than the investigated sample corresponds 18 M Ω•cm. to the general malt composition.Figure 1: PAD carbohydrate chromatogram of a malt extract sample containing 593 mg/L glucose, 5642 mg/L maltose, Example 1 – Carbohydrate Example 2 – Carbohydrate content1029 mg/L maltotriose, 291 mg/L maltotetraose, 164 mg/L maltopentaose, 327 mg/L maltohexaose and 97 mg/Lmaltoheptaose. Before analysis, the malt extract sample was diluted 1:100. determination in malt extract in dairy products Viscous or dried malt extracts are In contrast to the straightforwardproposed for the spectrophotometric of any reaction products from the obtained from germinated barley sample preparation of solubledetection of carbohydrates. However, electrode surface. The third, negative and contain naturally present sugar constituents in malt extractsthe improved sensitivity of these potential (E3) reduces any surface enzymes, particularly amylase, which presented above, the analysis ofmethods is offset by the labour- oxide on the electrode surface. The convert starch to water-extractable protein-containing samples suchintensive and often error-prone entire three-stage process typically sugars. Malt extracts excel in their as dairy products can pose severederivatization procedure. lasts one second and is continuously high physiological and nutritional problems when directly injected. Because carbohydrates are repeated to prevent electrode fouling. values. Malt is added as a nutritional Precipitation of the proteins foulselectrochemically active, amperometric Apart from carbohydrates, PAD has supplement to the diets of infants the column und ultimately destroysdetection overcomes the mentioned proven to be effective for amino and elderly persons. Furthermore, it. This can be prevented by theshortcomings. A triple-step potential sugars, amino acids, biogenic amines, it is a very important intermediate integration of upstream precipitationwaveform referred to as pulsed sulfur-containing species, alcohols, and ingredient in infant and pet foods, procedures such as Carrezamperometric detection (PAD) is some antibiotics. variety and party breads, instant precipitation. However, apart fromapplied. First a positive potential (E1) is Several applications demonstrate coffee, beverages, ice creams, the labour-intensive preparation,used to determine the target analytes, the potential of ion chromatography pharmaceuticals, etc. Carrez precipitation suffers fromfollowed by a second, more positive followed by pulsed amperometric coprecipitation, inclusions, andpotential (E2) for oxidative removal detection in food and beverage enhanced sugar decomposition.A Determining carbohydrates in foodstuffs using IC separation science — volume 3 issue 6
  3. 3. Figure 2Figure 2: Schematic diagram showing (a) inline dialysis cell (b) coupling to the sample changer and IC unit, and (c) spiral-flow dialysis cell. In contrast, carbohydrate solution is the same as that in carbohydrates migrate from the 95 and 105% indicate quantitativeseparation from high-molecular the donor solution. This patented sample stream into the acceptor permeability of the membrane forproteins can be comfortably stopped-flow procedure takes about solution, the high-molecular protein the target carbohydrates and thusachieved via stopped-flow dialysis. 14 minutes and can be directly constituents stay on the sample the applicability of the dialysisThis technique is based on the coupled to an IC setup (Figure 2). As side of the membrane. Finally, the technique.selective diffusion of molecules the dialysis is performed during the acceptor solution is transferred to theor ions from one liquid (donor recording of the previous sample`s sample loop. Carbohydrate content in variousor sample solution) to another chromatogram, the overall analysis food and beverage products – An(acceptor solution) via a membrane. time is not prolonged. Results overviewThe driving force for the transfer is Figure 3 shows the chromatogram Carbohydrate analysis offers verythe concentration gradient across the Experimental of a fruit yoghurt dialysate with the far-reaching application possibilitiesmembrane. The molecular separation The carbohydrate content in fruit following peaks: the polyol inositol, for the analysis of beverages, foods,thresholds are generally determined yoghurt was determined according the sugar alcohol sorbitol and the and sweets (Table 1). Unlike theby the thickness and porosity of the to the following procedure: Before mono- and disaccharides glucose, above-mentioned protein matrix,membrane. dialysis, 10 g yoghurt is dissolved in galactose, fructose, lactose, and most foodstuff samples only require Unlike in dynamic dialysis, where 1 L ultrapure water and the resulting sucrose. Repetitive analyses showed sample preparation methods suchtwo solutions continuously pass solution diluted 1:10 (v/v). 10 mL no trending in peak areas, peak as comminution, dilution, extraction,through the dialysis module, in of this solution is passed along the heights or retention times, which sonication, and/or filtration.equilibrium dialysis, at least one dialysis membrane by the peristaltic suggests that sample proteins did Table 1 gives an overview of thesolution is temporarily stopped until pump, while the acceptor solution not pass the membrane. In contrast, determination of various polyols,the concentration in the acceptor remains at rest. Whereas dialysable carbohydrate recovery rates between sugar alcohols as well as mono-, di-,A Determining carbohydrates in foodstuffs using IC separation science — volume 3 issue 6
  4. 4. Table 1 Figure 3 Figure 3: PAD carbohydrate chromatogram of a fruit yoghurt dialyzate containing 0.2 mg/g inositol, 0.8 mg/g sorbitol, 21.0 mg/g glucose, 7.4 mg/g galactose, 16.1 mg/g fructose, 26.5 mg/g lactose, and 77.0 mg/g sucrose. Determination of mono-and disaccharides in milk products by high-performance anion- (search for 8.000.6018EN). exchange chromatography with pulsedTable 1: Selection of various sugar determinations in different food matrices amperometric detection, Anal. Chem. 2003; 485: 43–49.and oligosaccharides in different gold working electrode can be used 2. Meyer A., Raba C., and Fischer K., Ion-foodstuffs. to determine various carbohydrates pair RP-HPLC determination of sugars, The presented setup also facilitates in different food matrices. Whereas aminosugars, and uronic acids aftercarbohydrate analysis in plant most samples require no sample derivatization with p-aminobenzoic acid,extracts, blood, urine, pharmaceutical preparation other than extraction, Anal. Chem. 2001; 73: 2377–2382.products, explosives, or biofuels. comminution, dilution, or filtration, 3. P. Deepak, Carbohydrate analysis in foodHowever, these determinations are carbohydrate determination in products and beverages, Food&Pack, 40beyond the scope of this article. difficult matrices can be easily solved (August 2002). by using Metrohm’s proven dialysis 4. Takematsu M.M., Carreira W., Batista F.F.,Conclusion technique. Zanuni L., and Sertek P.A., Quantification ofHigh-performance anion-exchange carbohydrates and uronic acids in eucalyptuschromatography with pulsed References wood species by ion chromatography withamperometric detection using a 1. Cataldi T.R.I., Angelotti M., and Bianco G., PAD using a gold electrode, Pittcon 2008,A Determining carbohydrates in foodstuffs using IC separation science — volume 3 issue 6