Sandalwood-like scents are expressed not only through natural sandalwood essential oil but also through a variety of synthetic fragrance ingredients. If their manufacture could selectively react only the necessary parts of a molecule and produce the three-dimensional forms suited to the scent, it might change how raw materials and processing steps are used.
A study published in 2024 by Maria C. Cancellieri and colleagues investigated combined-enzyme routes to four sandalwood fragrance ingredients. The central questions were whether only the desired reactions could proceed while preserving molecular forms important to odor, and whether the approach would offer advantages over conventional chemical processing.
Some bonds need to remain; others need to change
The four ingredients were Brahmanol, Firsantol, Sandalore, and Ebanol. These are individual synthetic ingredients with sandalwood-like odors, not mixtures reproducing sandalwood essential oil itself. In the study, the researchers worked stepwise from α-pinene, a plant-derived starting material, toward the target structures.
The challenge is deciding which of several bonds in an intermediate compound should change. When a molecule is reduced, for example by adding hydrogen, altering the double bond in the ring that is important to odor as well as the intended site in the side chain produces a different substance from the target.
A second challenge is stereochemistry. Molecules with the same atomic connectivity but different spatial orientations at particular positions are called stereoisomers. They may look similar in a flat structural drawing without necessarily smelling the same. Producing forms without selecting the desired one can expend materials and effort on forms that contribute little to the intended odor.
Two enzymes working sequentially in one vessel
The researchers combined two enzymes with different roles: an ene reductase and an alcohol dehydrogenase. The former reduces a particular carbon–carbon double bond, while the latter converts an oxygen-containing group into an alcohol. Enzymes can be understood here as biologically derived catalysts that promote targeted reactions.
They selected the yeast enzyme OYE2 for the first reaction. They then used one of two alcohol dehydrogenases with different properties according to the desired stereochemistry. Combining the form of the starting intermediate with the enzyme added later changed the stereochemistry of the resulting molecule.
The two reactions proceeded sequentially in the same vessel. They operated in a mainly aqueous medium at about 30°C, with a glucose-based system to regenerate the cofactors supporting enzyme activity. Extraction, purification, distillation, and other final steps removed impurities that might interfere with odor evaluation.
This does not mean that enzymes alone made the products all the way from a natural raw material. Earlier steps included chemical reactions, and conventional reducing agents were used to prepare some intermediates. The study focused on replacing reduction steps that particularly require selectivity with enzyme-based reactions.
Obtaining the intended form without changing too much
In a comparison using an intermediate leading to Sandalore, the OYE2 reaction converted about 96% of the starting material, with the desired product accounting for more than 99% of the products. Suppressing by-products in which the ring double bond was also reduced was an important result. The isolated yield of the purified target product was 87%.
These percentages describe different things. The 96% describes how much starting material reacted; more than 99% describes how selectively the desired product was formed among the products; and 87% describes how much was ultimately isolated. None is a percentage of people correctly distinguishing an odor.
The chemical reduction methods tested either reacted poorly or produced mixtures that included compounds with a reduced ring. For this substrate, the enzyme was better at selecting the reaction site.
The researchers also prepared different stereochemical forms of all four fragrance ingredients. Selectivity was not identical for every intermediate, however. Even small changes in molecular bulk or rigidity changed which form OYE2 preferentially produced.
Stereochemistry was examined using methods including nuclear magnetic resonance, which probes the environments around atoms. Some of the stereochemical bias generated by OYE2 was estimated indirectly, assuming that the stereochemical purity of the starting material was maintained through the intervening steps. Not every result was therefore measured directly with the same precision.
Sandalore’s odor also depended on stereochemistry
The prepared Sandalore forms were evaluated by fragrance experts. Of the four stereochemical forms, the one described as most pleasant had creamy, sandalwood-like characteristics, with an impact score of 8 out of 10 and a tenacity score of 9 out of 10.
Other forms tended toward amber or dry wood, or had an unpleasant initial impression followed by a marked decline in intensity. The results show that, even within the same Sandalore framework, stereochemistry affects odor quality and persistence.
This was a descriptive expert evaluation, not a large consumer-preference study. Nor was the tenacity score a measurement of how many hours the material lasted on skin. A score of nine cannot be read as nine hours of wear.
The authors explain these differences through a model in which the orientation of particular parts of the molecule alters its fit with odorant receptors. However, receptor binding or activation was not directly measured in this study; this is an interpretation connecting the odor evaluations with existing ideas.
Environmental advantages and challenges were not the same
The researchers compared conventional chemical and enzymatic methods for part of the production of Brahmanol. They examined the reduction steps from the same intermediate to the target ingredient, not an assessment covering raw-material production through product use and disposal.
The enzyme route avoided the high-pressure hydrogen and high temperatures needed in the chemical comparator and combined two steps in one vessel. On EcoScale, which deducts points for factors including hazards, reaction conditions, yield, and workup, the enzyme route scored 82 out of 100, while the two chemical steps scored 69 and 46.
On the other hand, the enzyme route performed worse on the metric of waste generated relative to the amount of product. Using materials such as glucose and working in relatively dilute solutions increases the amounts of substances and liquids used per unit of product.
The authors emphasize the advantages of the enzyme route, arguing that waste mass alone does not adequately capture differences in selectivity, hazards, equipment, and workup. What they demonstrated was promise across several criteria, not that using enzymes reduces every form of environmental burden.
The study received Italian and European public support, and companies supplied commercial fragrance ingredients and α-pinene. Assessors at CreaSens assisted with the expert odor evaluation. The authors declared no conflicts of interest.
Summary
This study demonstrated how enzymes can select reaction sites and stereochemical forms while preserving desired structures in the synthesis of sandalwood fragrance ingredients. The odor differences among Sandalore forms also illustrate why making the same molecular framework is not enough: the particular three-dimensional form matters.
The significance lies in the possibility of designing both the choice of odor-relevant molecules and the way those molecules are made. By exploiting mild conditions and the potential to combine steps while addressing challenges such as waste generation, this approach could offer options that consider both fragrance quality and manufacturing methods.
References
Cancellieri, M. C., Maggioni, D., Di Maio, L., Fiorito, D., Brenna, E., Parmeggiani, F., & Gatti, F. G.(2024). Biocatalytic approaches for a more sustainable synthesis of sandalwood fragrances. Green Chemistry, 26, 6150–6159. DOI: 10.1039/d4gc00746h. Full text.
The original article is published under CC BY-NC 3.0; journal rights notice: © The Royal Society of Chemistry 2024.
