• Celignis Analysis Package
    P13 : Sugars and Oligosaccharides in Solution

This package is particularly suitable for analysing the liquid output streams from biomass pre-treatment, hydrolysis, and other conversion processes. It 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.

In our reports for this package we report the pre-hydrolysis, post-hydrolysis and oligomeric concentrations of each sugar.

If you are also interested in finding the amounts of various sugar-degradation products in your liquid then, in addition to this analysis package, we would also recommend analysis package P22 - Organic Acids and Furans which can quantify a number of products, including levulinic acid, furfural, and formic acid).

If you are considering this analysis package to evaluate the outcome of your pre-treatment process, we have outlined a recommended set of analysis packages that will allow you to get detailed information about process yields and efficiencies.


Constituents Determined

For every constituent determined via wet-chemical analysis each sample is analysed in duplicate and we provide the compositional data for each replicate along with the average value and the standard deviation between the duplicates. Our analytical protocols typically provide results with a high level of precision, as detailed here.

Examples of the data reports generated can be viewed on the Celignis Database. Please log on to the guest account using email "test@celignis.com" and password "celignis".

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Publications Involving P13 with Celignis Authors

Bedzo, O. K. K., Gottumukkala, L. D., Sasso, G. L., Kaminski, K., Schlage, W., Goffman, F., Ivanov, N., Hoeng, J., Hayes, D. J. (2024) Process development for efficient pectin extraction from tobacco residues and its characterisation, Biomass Conversion and Biorefinery 14: 29481-29501

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This study presents a method for effective pectin extraction from the laminae of three tobacco varieties as a means of biomass valorisation. Two pre-treatment methods (cold ethanol vs. accelerated solvent extraction [ASE] with ethanol) were compared for their capacities to produce a high pectin yield. Enzymatic extraction of pectin was also tested as a green extraction procedure and compared to the acid extraction approach. The optimisation experiments revealed that cold ethanol extraction followed by acid hydrolysis is the most convenient method for pectin extraction; the optimal set of conditions for hydrolysis were identified as 90°C, pH 1.5, and 4 h of extraction. Applying these optimised conditions to the three Nicotiana rustica tobacco varieties yielded pectin recoveries of 66.2%, 57.8%, and 56.7% from the NRT63, Bakoum Miena, and NRT61 samples, respectively. Tobacco pectins were found to have a medium molecular weight and low methoxy content. These results highlight the potential of tobacco residues as feedstock for to produce pectin with dietary applications.

Outside Publications Including Celignis P13 Data

van der Waals, J., O’Callahan, D., Campion, S., Widsten, P., Thumm, A., Murton, K., West, M., Suckling, I., Fahmy, M. and Vaidya, A.A. (2025) Boosting softwood hemicellulose Hydrolysis: Enzymes from a new fungi Penicillium rotoruae remarkably Improve CTec-2 hydrolysis efficiency and reduce sugar production costs, Bioresource Technology133486

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Economic production of fermentable sugars from lignocellulosic biomass is critical for the biorefinery applications in the bioeconomy industry. This study demonstrates effective enzymatic hydrolysis of recalcitrant softwood using newly identified fungus Penicillium rotoruae. Initially, nineteen fungal isolates were screened on softwood galactoglucomannan (GGM), with nine showing strong responses in the liquid culture. Trichoderma viride, Penicillium rotoruae, and Amorphotheca resinae showed highest B-mannanase, B-mannosidase, and a-galactosidase activities. P. rotoruae demonstrated superior main chain cleaving enzyme activities, while A. resinae excelled in the side chain cleaving activity. The crude enzyme of P. rotoruae was evaluated on two Pinus radiata substrates. Using soluble GGM, P. rotoruae released 34.3% monomeric sugars (32.1 g/L reducing sugars), outperforming commercial CTec-2 (22.9% and 23.2 g/L respectively). Co-application of CTec-2 with P. rotoruae enzymes increased monomeric sugar yield to 56.3%, with galactose, mannose, and glucose increasing 20-, 3.6-, and 2.2-fold respectively. Using insoluble pulp, co-application yielded 88% of monomeric sugars (20.2 g/L reducing sugars) representing an increase of 20% soluble sugars relative to CTec-2 used alone. Techno-economic analysis indicated an increase in annual EBITDA, a positive ROCE and sugar cost savings of NZD 125/t demonstrating significant economic potential for softwood biorefineries.

Widsten, P., Murton, K., Thumm, A. and Bridson, J.H. (2023) Isolation and purification of high-molecular weight hemicelluloses from radiata pine wood chips prior to thermo-mechanical pulp (TMP) production, Holzforschung 77(5): 338-347

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A novel thermo-mechanical pulping (TMP) process has been developed to produce a by-product rich in high-molecular weight (MW) hemicelluloses, a potential raw material for barrier coatings and films. This process uses prehydrolysis to solubilise the hemicelluloses followed by chip compression to separate the soluble material from the wood matrix. The pressate from the chip compression stage was dark-coloured and had a high content of high-MW hemicelluloses and lignin. However, isolating the high-MW material from the pressate directly by ultrafiltration was not feasible because of membrane fouling by dissolved lignin and wood extractives, while coloured impurities are undesirable for many potential applications. To solve these problems the pressate was purified using XAD adsorbent resin to remove low-MW lignin, extractives and colour. Ultrafiltration of the purified pressate yielded lightly coloured high-MW hemicellulose with a low content of lignin and well suited for barrier films and coatings.

Smit, A.T., Verges, M., Schulze, P., van Zomeren, A. and Lorenz, H. (2022) Laboratory-to pilot-scale fractionation of lignocellulosic biomass using an acetone organosolv process, ACS Sustainable Chemistry & Engineering 10(32): 10503-10513

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Acetone organosolv fractionation of beech and birch wood at the lab-scale results in high sugar yields from the (hemi)cellulose and the isolation of a high-purity lignin. In this study, the process is scaled up to validate the technology at the pilot scale using industrial-size beech and birch wood chips and low liquid-to-solid ratios as a next step toward commercialization. Translation of the fractionation process to the pilot-scale showed a similar performance as compared to the lab-scale processing with a good conversion of the wood polymeric pentoses to mostly monomeric sugars and a high delignification. Continuous lignin precipitation by solvent evaporation using the LigniSep process resulted in the formation of nonsticky lignin aggregates with a good filterability. The improved lignin yields and advanced process design as compared to the traditional dilutive lignin precipitation approaches are likely to translate to a better process economy. The pulp washing efficiency and the recovery of (nonprecipitable) lignin from the aqueous hemicellulose stream still need to be improved for an efficient process design. However, the fractionation performance and high product concentrations in the spent liquor provide an excellent start position for improved process design at the commercial scale.

Torr, K.M., de Miguel Mercader, F., Murton, K.D., Harbers, T.J., Cooke-Willis, M.H., van de Pas, D.J. and Suckling, I.D. (2020) Fast pyrolysis of pine wood pretreated by large pilot-scale thermomechanical refining for biochemical production, Industrial & Engineering Chemistry Research 59(49): 21294-21304

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Pretreatments of lignocellulosic biomass prior to fast pyrolysis need to give a better yield and quality of pyrolysis oil and be practical to implement at a commercial scale. This study demonstrated, at a large pilot scale, that thermomechanical refining can reduce the particle size, hemicellulose content, and mineral content of pine wood chips, leading to improved yields of carbohydrate-derived chemicals on fast pyrolysis. Additionally, a hemicellulose-rich byproduct can be generated during the pretreatment for separate valorization. The impact of different presteaming times, citric acid infusion, and disc refining on the yield and composition of the pyrolysis oil was studied. Presteaming of citric acid-infused wood chips at 173 C for 3 min and refining to wood fiber proved most effective in improving the yield of pyrolysis oil organics, including the valuable biochemicals levoglucosan and hydroxyacetaldehyde, while reducing the yields of low-value char and noncondensable gases.



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