• Oligosaccharides
    Analysis at Celignis


Carbohydrates can be classified into three groups:

1. Monosaccharides - a single sugar unit (e.g glucose, xylose, and fructose).

2. Oligosaccharides - up to ten monosaccharide residues joined together by glycosidic linkages.

3. Polysaccharides - generally considered to be polymers of more than ten monosaccharide residues. Examples include cellulose, hemicellulose, and starch.

Disaccharides are the smallest form of oligosaccharides and consist of two monosaccharide units. Sucrose ((iii) in the figure below), a disaccharide of glucose and fructose is the most important disaccharide in plants, occupying much of the mass balance in species such as sugarcane, sugar beets and sweet sorghum. Other important disaccharides include the dimers cellobiose and maltose ((i) and (ii) in the figure below, respectively). These are the repeating units of cellulose and starch, respectively, and can be obtained upon their partial hydrolysis.

There are numerous oligosaccharides that have more than two units. More than 500 oligosaccharides with betwen three and ten sugar units are known, many of which occur as free natural substances. Due to their low chain length, oligosaccharides are often soluble in water and can be removed from biomass during extractives analysis. Oligosaccharides commonly found in the vegetative parts of grasses and some barks include raffinose (a trisaccharide containing galactose, glucose, and fructose) and stachyose (a tetrasaccharide which consists of two galactose units, glucose, and fructose). These are illustrated in the figure below ((i) - raffinose, (ii) - stachyose).

Request a QuoteOligomers Analysis

Oligosaccharides in Pre-Treatment and Hydrolysis Liquids

Hydrolysis biorefining technologies aim to produce biofuels and/or platform chemicals from the polysaccharides of lignocellulosic biomass by breaking them apart into their constituent monosaccharide (single sugar) units. This can be done in various ways, including using enzymes and chemicals. Furthermore, these technologies often involve a pre-treatment stage to make the lignocellulosic matrix more amenable to subsequent hydrolysis and in many cases these pre-treatments can partially hydrolyse some of the polysaccharides.

This partial hydrolysis can result in a significant proportion of the hemicellulosic and/or cellulosic sugars being tied up in soluble oligosaccharides that are contained within the process liquor. It is important to know whether the soluble sugars are present in monosaccharide or oligomer forms as these will affect subsequent downstream processing steps and the effective valorisation of the liquid component.

Oligosaccharides in Bio-Oil

Similarly, the water extract of the bio-oil fraction produced in pyrolysis may contain oligosaccharides in addition to the monosaccharides and anhydrosugars.

Please click here to read more about bio-oil and the analysis methods that we use to characterise it.

Request a QuoteBio-Oil Oligomers

Analysis of Oligosaccharides at Celignis

Direct Analysis of Oligosaccharides

We can characterise some oligosaccharides directly using our ion chromatography system. These include sucrose, cellobiose, raffinose, and melibiose. These can be analysed in the water extracts of biomass samples (obtained from analysis packages P5 - Water Extractives, P6 - Full Extractives, and P10 - Sugars, Lignin, Extractives, Ash) using analysis package P12 - Sugars in Solvent Extract, or in general process liquids using analysis package P21 - Sugars in Solution. These packages are detailed below.

If you would like us to directly determine other oligosaccharides then this may be possible depending on whether a suitable chromatography method for separation and appropriate chemical standards for those oligosaccharides are available. Please get in touch with us to discuss this.

Determination of the Proportions of Each Sugar in the Oligomeric Form

Analysis package P13 - Sugars and Oligosaccharides in Solution can be used to determine the total oligomeric sugar content of a liquid and also the relative proportions that each monosaccharide contributes to this total amount. The package involves a number of stages of analysis.

Firstly, the liquid is analysed using our ion chromatography system for the free monosaccharides and disaccharides in solution, just as in package P21 - Sugars in Solution.

Then we subject the liquid to a mild form of acid hydrolysis involving 4% sulphuric acid and 1 hour in an autoclave at 121 degrees Celcius. These conditions will break apart any oligosaccharides into their constituent monosaccharide units.

The liquid is subsequently filtered and analysed again on our IC system, to determine the amounts of various monosaccharides, after correction for any losses in sugars associated with the hydrolysis process. For each monosaccharide there will now be two values, the concentration prior to hydrolysis and that after hydrolysis.

With the exception of fructose (which degrades to hydroxymethylfurfural during the autoclaving step) all monosaccharides should be present in greater concentration post-hydrolysis. Hence, the proportion of each sugar that is present in the original liquid in the oligomeric form can be calculated by subtracting the pre-hydrolysis concentration from the post-hydrolysis concentration. As we report the sucrose and cellobiose contents separately we correct the calculated oligomeric glucan content using those data.

The constituents determined in analysis package P13 - Sugars and Oligosaccharides in Solution are detailed below. In our reports for P13 we report the pre-hydrolysis, post-hydrolysis and oligomeric concentrations of each sugar.

We have a similiar analysis package to P13 that is used for determining the oligomeric composition of the water extract of pyrolysis bio-oils. Analysis package P62 - Sugars and Oligosaccharides in Bio-oil Water Extract differs in that, rather than quantifying the sugar alcohols it instead determines the amounts of various anhydrosugars (such as levoglucosan, cellobiosan, mannosan, and galactosan) that are present in the original liquid.

As with P13, analysis package P62 then involves the acid hydrolysis of the liquid to calculate the oligomeric composition. However, in this case the anhydrosugars will be hydrolysed to their hydrated aldose equivalents (i.e. levogluocosan and cellobiosan to glucose, mannosan to mannose, and galactosan to galactose) and we consider this in our calculations to ensure that the proportions of each sugar that are reported as being present in oligomers actually are. Analysis package P62 is detailed below:

Publications on Oligosaccharides By The Celignis Team

Swart, L. J., Bedzo, O. K. K., van Rensburg, E., Gorgens, J. F. (2022) Pilot-scale xylooligosaccharide production through steam explosion of screw press-dried brewers spent grains, Biomass Conversion and Biorefinery 12: 1295-1309


Brewers spent grains (BSGs) represent the largest quantity of solid waste from brewing, while xylooligosaccharides (XOS) produced from BSG show promising applications in food, beverage and health products. Production of XOS from a Weiss and malt BSG was scaled-up in steam explosion hydrothermal treatment using process conditions from bench-scale liquid hot water optimisations in stirred batch reactors. Three levels of moisture (15, 25 and 32% dry matter) achieved by screw press dewatering were evaluated by varying the treatment temperatures and times. Results show the highest XOS yields (73.1%) were obtained, for both BSGs, at process condition selected (180 C, 10 min) with 25% initial dry matter content. These yields were higher than reported bench-scale optimisations (61%), but obtained using 60% less water; hence, initial dry matter content was an important variable affecting XOS yield. The pilot-scale steam explosion results provide a departing point for a cost-effective commercial production of XOS from BSG.

Swart, L. J., Bedzo, O. K. K., van Rensburg, E., Gorgens, J. F. (2021) Intensification of Xylo-oligosaccharides Production by Hydrothermal Treatment of Brewers Spent Grains: The Use of Extremely Low Acid Catalyst for Reduction of Degradation Products Associated with High Solid Loading, Applied Biochemistry and Biotechnology 193: 1979-2003


Brewers' spent grains (BSG) make up to 85% of a brewery's solid waste, and is either sent to landfill or sold as cheap animal feed supplement. Xylo-oligosaccharides (XOS) obtained from BSG are antioxidants and prebiotics that can be used in food formulations as low-calorie sweeteners and texturisers. The effect of extremely low acid (ELA) catalysis in liquid hot water (LHW) hydrothermal treatment (HTT) was assessed using BSG with dry matter contents of 15% and 25%, achieved by dewatering using a screw press. Batch experiments at low acid loadings of 5, 12.5 and 20 mg/g dry mass and temperatures of 120, 150 and 170 C significantly affected XOS yield at both levels of dry mass considered. Maximum XOS yields of 76.4% (16.6 g/l) and 65.5% (31.7 g/l) were achieved from raw BSG and screw pressed BSG respectively, both at 170 C and using 5 mg acid/g dry mass, after 15 min and 5 min, respectively. These XOS yields were obtained with BSG containing up to 63% less water and temperatures more than 20 C lower than that reported previously. The finding confirms that ELA dosing in LHW HTT allows lowering of the required temperature that can result in a reduction of degradation products, which is especially relevant under high solid conditions. This substantial XOS production intensification through higher solid loadings in HTT not only achieved high product yield, but also provided benefits such as increased product concentrations and decreased process heat requirements.

Swart, L. J., Peterson, A. M., Bedzo, O. K. K., Gorgens, J. F. (2021) Techno-economic analysis of the valorization of brewers spent grains: production of xylitol and xylo-oligosaccharides, Journal of Chemical Technology & Biotechnology 96(6): 1632-1644


Brewers spent grains (BSG) represents around 85% of a brewery's solid waste and common disposal to landfill is increasingly more difficult. Yet BSG is a food-grade by-product with potential economic valorization that can significantly improve resource efficiency and reduction in carbon emissions. This study investigated valorization of BSG through the application of novel high solids hydrothermal processing technology in a small-scale biorefinery, annexed to a brewery. It focused on three scenarios for the production of: (A) the sugar replacement xylitol; (B) prebiotic xylo-oligosaccharide (XOS); and (C) co-production of xylitol and XOS. Economic assessment was conducted by comparing the capital and operating expenditure from process simulations created in Aspen Plus. The process models developed were supplemented with experimental data to improve accuracy.
Internal rate of return (IRR) values obtained were greater than the hurdle rate of 9.7% for all scenarios when considering a conservative market price for xylitol and XOS as US$4500 t-1, yet dedicated production of XOS was economically more favourable with a minimum required selling price (MRSP) of US$2509 t-1 compared to US$4153 t-1 for xylitol. Additionally, the scenario for co-production of xylitol and XOS achieved the lowest MRSP of US$2182 t-1. By-products significantly contributed to 32.7%, 14.2% and 27.5% of the revenue generated in scenarios A, B and C, respectively.
These results provide a good platform from which to develop the cost-effective commercial production of XOS and xylitol from BSG.

Bedzo, O. K. K., Mandegari, M. and Gorgens, J. F. (2020) Techno-economic analysis of inulooligosaccharides, protein, and biofuel co-production from Jerusalem artichoke tubers: A biorefinery approach, Biofuels Bioproducts & Biorefining-Biofpr 14(4): 776-793


Jerusalem artichoke (JA) is a crop with excellent potential for application in biorefineries. It can resist drought, pests, and diseases and can thrive well in marginal lands with little fertilizer application. The JA tubers contain considerable quantities of inulin, which is suitable for the production of inulooligosaccharides (IOS), as a high-value prebiotic, dietary fiber. In this study, five JA tuber biorefinery scenarios were simulated in Aspen Plus and further evaluated by techno-economic and sensitivity analyses. Production of IOS, proteins and animal feed was studied in scenarios A and C, applying various biorefinery configurations. Scenario B explored the option of producing only IOS and the sale of residues as animal feed. Scenarios D and E investigated the economic potential of biofuel generation from residues after IOS and protein production by generation of biogas and ethanol respectively, from residues. Based on the chosen economic indicators, scenario B resulted in the lowest minimum selling price (MSP) of 3.91 US$ kg-1 (market price 5.0 US$ kg-1) with correspondingly reduced total capital investment (TCI) and total operating cost (TOC) per mass unit produced of IOS of 18.91 and 2.59 US$ kg-1 respectively, compared with other studied scenarios. Considering the set production scale, it is more profitable when the residues are sold as animal feed instead of being converted into biofuel, due to the capital-intensive nature of the biofuel production processes. The coproduction of protein had a negative impact on the economics of the process as the associated capital and operating expenditure outweighed the associated revenue.

Bedzo, O. K. K., Trollope, K., Gottumukkala, L. D., Coetzee, G., Gorgens, J. F. (2019) Amberlite IRA 900 Versus Calcium Alginate in Immobilization of a Novel , Engineered B-fructofuranosidase for Short-Chain Fructooligosaccharide Synthesis from Sucrose, Biotechnology Progress 35(3): 1-9


The immobilization of B-fructofuranosidase for short-chain fructooligosaccharide (scFOS) synthesis holds the potential for a more efficient use of the biocatalyst. However, the choice of carrier and immobilization technique is a key to achieving that efficiency. In this study, calcium alginate (CA), Amberlite IRA 900 (AI900) and Dowex Marathon MSA (DMM) were tested as supports for immobilizing a novel engineered B-fructofuranosidase from Aspergillus japonicus for scFOS synthesis. Several immobilization parameters were estimated to ascertain the effectiveness of the carriers in immobilizing the enzyme. The performance of the immobilized biocatalysts are compared in terms of the yield of scFOS produced and reusability. The selection of carriers and reagents was motivated by the need to ensure safety of application in the production of food-grade products. The CA and AI900 both recorded impressive immobilization yields of 82 and 62%, respectively, while the DMM recorded 47%. Enzyme immobilizations on CA, AI900 and DMM showed activity recoveries of 23, 27, and 17%, respectively. The CA, AI900 immobilized and the free enzymes recorded their highest scFOS yields of 59, 53, and 61%, respectively. The AI900 immobilized enzyme produced a consistent scFOS yield and composition for 12 batch cycles but for the CA immobilized enzyme, only 6 batch cycles gave a consistent scFOS yield. In its first record of application in scFOS production, the AI900 anion exchange resin exhibited potential as an adequate carrier for industrial application with possible savings on cost of immobilization and reduced technical difficulty.