Enzymatic Diol Stereo-Selective Production via Epoxide Hydrolase

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Solution Overview

Problem

Current chemical synthesis methods for producing (8R,9S)-cis-cedrene diol are inefficient, resulting in low yields and requiring extensive purification, while existing biotechnological methods lack a suitable method for producing this specific diol.

Innovation Solution

A method involving the conversion of trans-diol to hydroxyketone and subsequent stereo-selective reduction using a 3-alpha-hydroxysteroid dehydrogenase enzyme, which can catalyze both oxidation and reduction steps, allowing for high-purity and high-yield production of (8R,9S)-cis-cedrene diol.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Manufacturing precision

If chemical epoxidation followed by acid-catalyzed hydrolysis is used to produce cis-cedrene diol, then the desired product can be obtained, but the yield is low and extensive purification is required

Engineering Contradiction:

Improvepurity of cis-cedrene diolVSAvoidyield of cis-cedrene diol

Core Design Contradiction:

Manufacturing precisionVSProductivity

Solution Approach 1:

The patent replaces chemical synthesis methods with enzymatic biocatalysis. Specifically, epoxide hydrolase enzymes are used to catalyze the ring-opening of cedrene epoxide to produce cis-cedrene diol, substituting the mechanical/chemical acid-catalyzed hydrolysis process. This enzymatic approach provides high stereoselectivity for the cis-isomer, eliminating the need for extensive purification and significantly improving both yield and purity simultaneously.

Inventive Principle:

Principle #28Mechanics substitution (Replace mechanical system)

2Manufacturing precision

If chemical cis-dihydroxylation using osmium tetroxide is used to synthesize cis-cedrene diol, then the desired product can be obtained, but toxic substances must be used

Engineering Contradiction:

Improvestereoselectivity for cis-diolVSAvoidtoxicity of osmium tetroxide

Core Design Contradiction:

Manufacturing precisionVSObject-affected harmful factors

Solution Approach 1:

The patent substitutes the toxic chemical osmium tetroxide system with biologically derived epoxide hydrolase enzymes. These enzymes naturally catalyze the formation of cis-diols from epoxides with high stereoselectivity, providing the same manufacturing precision without the harmful toxicity associated with osmium-based reagents.

Inventive Principle:

Principle #28Mechanics substitution (Replace mechanical system)

Solution Approach 2:

The patent employs readily available commercial substrates (α-cedrene, cedrene epoxide, or cedrol) that can be processed by the enzymatic system. The enzymatic catalysts are used in catalytic amounts and can be regenerated or replaced, avoiding the need for expensive and toxic stoichiometric reagents like osmium tetroxide.

Inventive Principle:

Principle #27Cheap short-living objects (Disposable)

3Productivity

If microbial biotransformation of (8R)-cedrol is used to produce cedrene diol, then biotechnological production can be achieved, but the wrong epimer or trans-diol is produced

Engineering Contradiction:

Improvebiotechnological production efficiencyVSAvoidstereoisomer configuration

Core Design Contradiction:

ProductivityVSManufacturing precision

Solution Approach 1:

The patent changes the substrate parameter from cedrol to cedrene epoxide or α-cedrene as the starting material. This parameter change enables the epoxide hydrolase to produce the desired cis-cedrene diol with correct stereochemistry, whereas previous attempts using cedrol as substrate led to wrong epimers or trans-diols through different reaction mechanisms.

Inventive Principle:

Principle #35Parameter changes

Applied Scientific Principles

This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.

Function Achieved in This Case

This method achieves efficient conversion of (8R,9R)-trans-cedrene diol to (8R,9S)-cis-cedrene diol with high stereo-selectivity and yield, eliminating the need for expensive regeneration systems and reducing the complexity of purification processes.

Implementation Method 1

the conversion(s) is/are catalyzed by an enzyme, which is encoded by a nucleic acid sequence of SEQ ID NO: 1 or wherein the enzyme comprises an amino acid sequence of SEQ ID NO: 2

Methodology Applied

Scientific EffectEnzyme catalysis: Enzyme

Implementation Method 2

A method involving the conversion of trans-diol to hydroxyketone and subsequent stereo-selective reduction using a 3-alpha-hydroxysteroid dehydrogenase enzyme

Methodology Applied

Scientific EffectStereo-selective reduction:

Implementation Method 3

conversion of trans-diol to hydroxyketone and subsequent stereo-selective reduction

Methodology Applied

Scientific EffectRedox reactions: Redox Reactions

Data Source

PatentUS12281343B2Biotechnological production of diols

Publication Date: 2025.04.22 SYMRISE GMBH & CO KG

AI summary

The present invention relates to a method for the stereo selective production of a trans-diol or a cis-diol or a hydroxyketone comprising the step(s) (i) conversion of a trans-diol or a cis-diol to a hydroxyketone and/or (ii) conversion of a hydroxyketoneto a cis-diol or a trans-diol, catalyzed by an enzyme, which is encoded by a nucleic acid sequence of SEQ ID NO: or wherein the enzyme comprises an amino acid sequence of SEQ ID NO: 2. The present invention also relates to the use of an enzyme encoded by a nucleic acid sequence of SEQ ID NO: 1 or wherein the enzyme comprises an amino acid sequence of SEQ ID NO: 2 for the conversion of a trans-diol to a cis-diol or for the conversion of a trans-diol or a cis-diol to a hydroxy ketone and/or the conversion of a hydroxyketone to a trans-diol or a cis-diol.