• Biomass Hydrolysis
    Bioprocess Development
    At Celignis Biomass Lab


Lignocellulosic Biomass

Lignocellulosic biomass is defined as a plant, or plant-derived, material that is mostly composed of cellulose, hemicellulose, and lignin. Lignocellulosic feedstocks are highly abundant, covering many biomass types including grasses, wood, energy crops (e.g. Miscanthus and coppices), agricultural residues (e.g. straws and corn stover), and municipal wastes.

Lignocellulosic feedstocks are highly abundant and can often be sourced sustainably, at low cost, without leading to land-use conflicts. As a result, there is currently great interest in obtaining chemicals, fuels, and biomaterials from such biomass.

Hydrolysis of Lignocellulose

A major pathway by which many lignocellulosic feedstocks are processed is known as hydrolysis, where monomeric sugars are released from the lignocellulosic polysaccharides (i.e. cellulose and hemicellulose). Typically, these polysaccharides are hydrolysed by acid or, more commonly, by enzymes. The hydrolysis of cellulose will yield monomeric glucose (as cellulose is a hompolysaccharide, i.e only containing one type of sugar), whilst the hydrolysis of hemicellulose will yield a variety of different sugars covering the hexoses (6-carbon sugars) glucose, galactose, and mannose, and the pentoses (5-carbon sugars) xylose and arabinose, depending on the type of hemicellulose. Hydrolysis of hemicellulose can also yield uronic acids and acetyl groups.

However, the hydrolysis of lignocellulosic polysaccharides is not easy and is influenced by the complex inter-associations between hemicellulose and cellulose and between these polysaccharides and lignin in the lignocellulosic matrix. In particular, the crystalline nature of much cellulose and the existence of a physical barrier of lignin surrounding the cellulose fibres are said to be major contributors to the recalcitrance of cellulose.

The mechanism of hydrolysis is further complicated by the fact that different process intensities are required for the hydrolysis of cellulose versus hemicellulose. The more intense conditions required for cellulose hydrolysis may degrade the sugars hydrolysed from hemicellulose (to products such as furfural and formic acid).

For this reason, most hydrolysis technologies employ pre-treatment processes that aim to break apart the matrix (and in particular the associations between lignin and cellulose), reduce cellulose crystallinty, and (in some cases) hydrolyse hemicelluloses, hence separating the hydrolysate from cellulose which can then undergo more severe/targeted treatment.

Click below to read more about bioprocess development for biomass pretreatment.

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History of Hydrolysis of Lignocellulosic Biomass

Early Work (Acid Hydrolysis)

The potential of lignocellulosic biomass as a source of fermentable sugars was recognized as early as the late 19th century, with notable work by researchers such as Charles Tanret. However, the robust structure of lignocellulose—comprising cellulose, hemicellulose, and lignin—posed significant challenges. Acid hydrolysis emerged as an early method for biomass conversion, making use of sulphuric acid to cleave glycosidic bonds in cellulose and hemicellulose.

The acid hydrolysis of lignocellulose materials was commercialised in the late 19th century and several dilute-acid facilities existed in the USA, Germany, Japan, and Russia by World War 1 while concentrated acid hydrolysis facilities were being built between 1937 and the late 1960s. However, these were uneconomic where fossil fuels were available and very few facilities were operational at the end of the 20th century.

Early Enzymatic Hydrolysis

In the mid-20th century, a paradigm shift occurred with the introduction of enzymatic hydrolysis. Enzymes produced by microorganisms, such as Trichoderma reesei, were found to efficiently convert cellulose into glucose. This biological hydrolysis was more environmentally friendly and yielded higher sugar concentrations. Despite its promise, the cost of producing cellulase enzymes was prohibitive, which led to a research emphasis on reducing enzyme costs and improving their efficiency.

Research Advances

It was recognised that the recalcitrance of lignin remained a major challenge in developing efficient biological hydrolysis processes. As a result, pre-treatment processes were introduced to enhance the accessibility of cellulose and hemicellulose to enzymes. These processes include dilute acid pretreatment, steam explosion, and organosolv methods. Each of these methods disrupts the lignocellulosic structure in various ways, increasing the efficiency of subsequent enzymatic hydrolysis.

There were also significant advances, in the late 20th century and early 21st century, on the developement of enzymes that were more efficient in hydrolysing biomass and less sensitive to inhibition. Additionally, improvements were also made in the engineering of enzyme production systems and enzyme recycling technologies. These all helped to reduce the costs associated with the use of enzymes for hydrolysing biomass. As a result, there are a handful of commercial-scale biorefineries using enzymes for the hydrolysis of lignocellulosic feedstocks.

Ongoing Research

Research continues for developing improved pretreatments and more robust enzymes. Additionally, the early 21st century has seen a particular focus on the development of consolidated bioprocessing (CBP). This approach consolidates enzyme production, saccharification, and fermentation into a single step. CBP employs engineered microbes, such as Clostridium thermocellum, that can both produce cellulolytic enzymes and ferment sugars into ethanol. This process further simplifies the biomass-to-product conversion process and reduces costs.

Acid Hydrolysis of Lignocellulosic Biomass

Dilute-Acid Hydrolysis

This involves the use of acids, in relatively low concentrations and at elevated temperatures, to hydrolyse the biomass polysaccharides. The dynamics and outputs of the process are highly dependent on its severity, based on factors such as the type and concentration of acid and the temperatures used. Since cellulose requires more severe hydrolysis conditions than hemicellulose, the dilute-acid approach usually involves two stages. The fist stage focuses on the hydrolysis of hemicellulose and the second stage then involves more severe conditions that allow for cellulose to be hydrolysed.

Click below to read more about dilute-acid hydrolysis and bioprocess development to optimise this approach.

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Concentrated Acid Hydrolysis


Click below to read more about concentrated acid hydrolysis.

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Enzymatic Hydrolysis of Lignocellulosic Biomass

Enzymatic hydrolysis is an alternative, biological, approach for obtaining monomeric sugars from lignocellulose. It involves the action of cellulases, for the hydrolysis of cellulose, and hemicellulases for the hydrolysis of hemicellulose. These enzymes can be produced from baceria and fungi, with a number of different types of enzymes required for effective hydrolysis of these polysaccharides. For example, cellulose hydrolysis involves the activities of endoglucanases, exoglucanases, and betaglucosidases.

Some of the different enzymatic hydrolysis technologies are listed and described below.

Separate Hydrolysis and Fermentation (SHF)

This process involves two distinct stages: hydrolysis, in which the cellulose and hemicellulose (if not already removed in the pretreatment) are broken down into simple sugars; and fermentation, where these sugars are transformed into biofuels.

Click below to read more about SHF and bioprocess development to optimise this approach.

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Simultaneous Saccharification and Fermentation (SSF)

SSF involves the concurrent breakdown (hydrolysis) of cellulose (and hemicellulose, if present) into monomeric sugars (saccharification), and the conversion of these sugars into products via fermentation. Unlike in SHF, in SSF both stages take place in the same reactor.

Click below to read more about SSF and bioprocess development to optimise this approach.

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Simultaneous Saccharification and Co-Fermentation (SSCF)

SSCF is a modification of the Simultaneous Saccharification and Fermentation (SSF) method, incorporating a step to ferment pentose sugars alongside the conventional hexose sugars.

Click below to read more about SSCF and bioprocess development to optimise this approach.

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Consolidated Bioprocessing (CBP)

Consolidated Bioprocessing (CBP) is an even more integrated approach in which the enzyme production, hydrolysis, and fermentation steps all occur in one step and one reactor.

Click below to read more about CBP and bioprocess development to optimise this approach.

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Other Celignis Services for Bioprocess Development

Global Recognition as Bioprocess Experts

Celignis provides valued services to over 1000 clients. We understand how the focus of bioprocess projects can differ between countries and have advised a global network of clients. We also have customs-exemptions for samples sent to us allowing us to quickly get to work no matter where our clients are based.

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Biomass can be rich in bioactive compounds of high value for food, feed, cosmetic, and pharmaceutical applications. We develop bespoke extraction methods suitable for your needs with high selectivity, efficiency and low environmental impact.

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The choice of pretreatment method varies with the type of biomass and the end-product requirements. At Celignis we can determine the most suitable pretreatment for your feedstock and determine the optimum conditions in lab-scale trials followed by higher TRL scale-ups.

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Enzymes are biological catalysts that have a wide variety of applicaitons in the bioeconomy, ranging from the liberation of sugars from lignocellulosic biomass to the functionalisation of biomass-derived chemicals and materials for higher-value applications. We are experts in the design and use of enzymatic approaches.

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Development of fermentation processes requires knowledge of an array of important factors including: biomass, the microbes used, nutrient media, and fermentation conditions. We're experienced in many fermentations and can help you determine and optimise yields of an array of different fermentation products.

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Downstream Processing

How the various outputs (solid and liquid) of a bioprocess are dealt with is often overlooked until later in bioprocess development, leading to excessive costs and complications. We consider and tackle these issues, and others such as product recovery, early-on as being integral to the bioprocess.

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Lab-Scale Optimisations

We consider that optimising a bioprocess at the lab-scale is the most cost-effective approach to explore a range of different scenarios in search of optimal process conditions. Based on the outputs of these experiments we can then test the chosen set of conditions at higher TRL levels.

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TRL Scale-Up

At our dedicated Celignis Bioprocess laboratories we have all the necessary upstream and downstream apparatus to undertake bioprocess projects up to a tehcnology readiness level (TRL) of 6, with reactor and processing capacities of up to 100 litres.

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Technoeconomic Analyses

Our technoeconomic experts can evaluate your bioprocess, considering various scale, technology, and feedstock options. We apply accurate costing models to determine CAPEX/OPEX of simulated and pilot-scale processes which are then used to determine key economic indicators (e.g. IRR, NPV).

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Biobased Chemicals

A large array of chemicals and materials are possible from biomass and wastes. These can involve chemical or biological approaches, or a combination of the two. Based on your desired end-product we can design and test the most appropriate bioprocess.

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From Process Refinements to an Entire New Process

We work closely with you to understand your objectives and timelines. We then propose a project, usually covering a series of deliverables and stage-gates. Often our projects involve optimising conditions at the lab-scale before replicating the conditions at higher TRL levels.

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Research Collaborations

Celignis is active in several bioprocess research projects. These include projects funded by the EU's CBE-JU, with Celignis being a Full Industry Member of the BIC. We're open to participating in future collaborative research projects where our extensive infrastructure and expertise in bioprocesses can be leveraged.

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