Malic Acid Bioproduction Engineering Service

Malic Acid is a dicarboxylic acid widely used as a food acidulant, flavoring agent, and precursor for chemical synthesis. Traditional chemical synthesis involves high pollution due to the use of hazardous chemicals (e.g., maleic anhydride) and generates racemic mixtures that require costly separation. While biosynthesis is environmentally friendly, a major challenge is low yield in biosynthesis , as Malic Acid is a central metabolic intermediate quickly consumed by the host organism for growth and energy production, leading to low final titers.

CD Biosynsis offers a synthetic biology service focused on establishing an efficient and sustainable bioproduction route. Our core strategy involves modification of the Aspergillus oryzae metabolic pathway (a robust industrial fungal host) to enhance the flux through the glyoxylate cycle or CO2 fixation pathways, which are critical for Malic Acid synthesis. This is coupled with the overexpression of malate dehydrogenase (or other key pathway enzymes like fumarase or pyruvate carboxylase) to boost the conversion rate to Malic Acid and prevent its degradation. This integrated approach aims to deliver a high-titer, high-yield, and food-grade L-Malic Acid product from renewable sugar feedstock.

Pain Points

Developing a sustainable and economically competitive Malic Acid bioproduction route faces these key challenges:

• High Pollution in Chemical Synthesis: The use of toxic intermediates and high-energy processes leads to significant environmental waste and costly remediation .
• Low Yield in Biosynthesis: Malic Acid is a branch point in the TCA cycle, meaning it is quickly metabolized to other products (e.g., succinate, fumarate) or used for biomass production, limiting accumulation.
• Product Toxicity: High concentrations of Malic Acid, especially in its acidic form, can inhibit the host cell's growth and viability, capping the maximum achievable titer.
• Stereoisomer Purity: Chemical synthesis yields a mix of L- and D-Malic Acid. Bioprocesses must produce the single, preferred L-isomer (food-grade) with high purity.

A cost-effective solution must successfully redirect the carbon flux toward Malic Acid synthesis and block its degradation.

Solutions

CD Biosynsis utilizes advanced metabolic engineering to optimize Malic Acid production in Aspergillus oryzae :

Modification of Aspergillus oryzae Metabolic Pathway

           

We employ genome editing to inactivate enzymes that consume Malic Acid (e.g., Malate enzyme) and upregulate CO2 fixation pathways (Pyruvate carboxylase) to increase Malate supply.

Overexpression of Malate Dehydrogenase

We specifically overexpress Malate dehydrogenase (or Fumarase) to boost the conversion rate from oxaloacetate/fumarate to Malic Acid, creating an efficient sink.

NADH and Cofactor Optimization

We engineer the host metabolism to ensure sufficient NADH supply (required for the MDH reaction) and maintain the redox balance crucial for high-yield production.

Product Tolerance and Transport Enhancement

We introduce or upregulate Malate transporters to export Malic Acid out of the cell , reducing product inhibition and increasing the final secreted titer.

This systematic approach is focused on overcoming the native host's tendency to consume Malic Acid, maximizing its extracellular secretion.

Advantages

Our Malic Acid engineering service is dedicated to pursuing the following production goals:

High L-Malic Acid Purity

Biosynthesis naturally yields the single, enantiomerically pure L-isomer , meeting food and pharmaceutical standards and avoiding expensive separation.

Significantly Increased Titer

Blockage of consumption pathways and optimized transport ensures maximum Malic Acid accumulation in the culture broth.

Environmentally Sustainable Process

Microbial production from sugar feedstock eliminates the need for hazardous chemical inputs and high energy usage. [Image of Cost Reduction Icon]

Food-Grade Host Safety

Aspergillus oryzae is a generally recognized as safe (GRAS) organism, simplifying regulatory hurdles for food and feed applications.

Efficient Substrate Conversion

Metabolic balancing maximizes the carbon yield (g Malic Acid}/\text{g substrate), improving raw material economics.

We provide a specialized platform aimed at maximizing the quality and cost-effectiveness of Malic Acid biomanufacturing.

Process

Our Malic Acid strain engineering service follows a rigorous, multi-stage research workflow:

• TCA Cycle and Malate Consumption Blockage: Knock out competing enzymes (Malate enzyme, Fumarase) to prevent Malic Acid degradation or conversion to other products.
• Pathway Upregulation and CO2 Fixation: Overexpress key anaplerotic enzymes (Pyruvate carboxylase or PEP carboxylase) to increase the flow of carbon into the C}_4$ synthesis route.
• Enzyme Overexpression and NADH Balancing: Introduce multiple copies of the Malate dehydrogenase gene and co-express genes that enhance NADH regeneration to ensure high conversion efficiency.
• Transport Engineering and Tolerance: Screen or engineer novel Malate transporters to enhance export and select strains with inherent pH tolerance to reduce the need for excessive neutralization.
• Fermentation Performance Validation: Test the final engineered strain in fed-batch fermentation to assess Malic Acid titer, yield, and stereochemical purity .
• Result Report Output: Compile a detailed Experimental Report including gene modification data, flux analysis, and fermentation metrics (yield, titer, and purity) , supporting process transfer.

Technical communication is maintained throughout the process, focusing on timely feedback regarding yield and product purity.

Explore the potential for a high-yield, sustainable L-Malic Acid supply. CD Biosynsis provides customized strain and enzyme engineering solutions:

• Detailed Metabolic Flux and Titer Analysis Report , demonstrating success in carbon redirection and titer increase.
• Consultation on fermentation control strategies optimized for high-density growth and acid neutralization.
• Experimental reports include complete raw data on carbon yield (g Malic Acid}/\text{g sugar) and stereochemical purity , essential for regulatory and commercial use.