produced (Belin et al. 1992). These methods lead to the formation of one major
product involving either one (biotransformation) or several (bioconversion) biochemical steps (Cheetham 1997). The biotransformation processes involve different
biochemical reactions such as oxidation, reduction, hydrolytic reactions, dehydration and formation of new C–C bonds. In fact, the flavour production in
microorganisms is carried out by enzymes which comprise hydrolytic enzymes,
transferases, oxidoreductases and lyases (Schreier 1997). The yield of flavoured
product in bioconversion and biotransformation processes is also higher as compared to de novo synthesis. Therefore, approaches for production of flavour
compounds are also economical (Welsh et al. 1989; Amalraj et al. 2017). In the
beginning of the era of aroma production, both de novo and biotransformation
methods were used for the production of flavour compounds. The microorganisms
were isolated, screened and selected for their unique aroma production properties.
But, in order to produce completely new set of flavours and to improve the fragrance
yield and notes, the microorganisms have been modified by the techniques of
metabolic and genetic engineering, which can also be termed as synthetic biology.
The genetically engineering strains of bacteria for flavour production were first
used in 2010, when there was an acute shortage of fragrance oil, extracted from
Pogostemon cablin (Patchouli), an essential oil used as a fragrance in incense sticks,
personal and health care products. Due to heavy rains in Indonesia, the medicinal
shrub was destroyed which led to a poor harvest of fragrant oil (Gupta et al. 2015;
Mahajan and Phatak 2019). Biotechnological interventions solved the crisis when
the firms like Allylix, Isobionics and Evolva used genetically engineering bacteria
(e.g. Pseudomonas sp.) and yeast (Saccharomyces sp.) that could produce plant oils
by fermenting sugars (Mahajan and Phatak 2019).The gene coding for a particular
aroma character was identified and isolated from its known source. Following the
cloning approaches, the target gene was cloned and expressed either in E. coli or
Saccharomyces cerevisiae, thus fragrance producing recombinants were produced.
Microbial synthesis of aroma compounds which is classified into different groups is
described in the next section.
7.3.2.1 Fruity and Floral Terpenes
Terpenes are the flavoured components of essential oils, consisting of five carbon
isoprene units which are assembled to each other, while terpenoids are modified
class of terpenes with different functional groups and oxidized methyl group at
various C positions (Perveen 2018). Only a few terpenes have been reported to be
produced by de novo methods using microorganisms, usually fungi belonging to
group Ascomycetes and Basidiomycetes (Gupta et al. 2015). Fungi such as
Kluyveromyces lactis and Ceratocystis moniliformis produce de novo fruity and
floral flavoured terpenes such as citronellol, linalool and geraniol (Drawert and
Barton 1978; Bluemke and Schrader 2001). However, the methods involving biotransformation of terpenes are milder and produce less toxic wastes and, hence, are
better alternative for the production of natural aroma compounds (Dionísio et al.
2012; Bier et al. 2011).The pathway for biotransformation of geraniol into methyl
heptenone has been elucidated for Penicillium digitatum and citral lyase enzyme
7 Biotechnological Interventions for Production of Flavour and Fragrance. . .
137
product involving either one (biotransformation) or several (bioconversion) biochemical steps (Cheetham 1997). The biotransformation processes involve different
biochemical reactions such as oxidation, reduction, hydrolytic reactions, dehydration and formation of new C–C bonds. In fact, the flavour production in
microorganisms is carried out by enzymes which comprise hydrolytic enzymes,
transferases, oxidoreductases and lyases (Schreier 1997). The yield of flavoured
product in bioconversion and biotransformation processes is also higher as compared to de novo synthesis. Therefore, approaches for production of flavour
compounds are also economical (Welsh et al. 1989; Amalraj et al. 2017). In the
beginning of the era of aroma production, both de novo and biotransformation
methods were used for the production of flavour compounds. The microorganisms
were isolated, screened and selected for their unique aroma production properties.
But, in order to produce completely new set of flavours and to improve the fragrance
yield and notes, the microorganisms have been modified by the techniques of
metabolic and genetic engineering, which can also be termed as synthetic biology.
The genetically engineering strains of bacteria for flavour production were first
used in 2010, when there was an acute shortage of fragrance oil, extracted from
Pogostemon cablin (Patchouli), an essential oil used as a fragrance in incense sticks,
personal and health care products. Due to heavy rains in Indonesia, the medicinal
shrub was destroyed which led to a poor harvest of fragrant oil (Gupta et al. 2015;
Mahajan and Phatak 2019). Biotechnological interventions solved the crisis when
the firms like Allylix, Isobionics and Evolva used genetically engineering bacteria
(e.g. Pseudomonas sp.) and yeast (Saccharomyces sp.) that could produce plant oils
by fermenting sugars (Mahajan and Phatak 2019).The gene coding for a particular
aroma character was identified and isolated from its known source. Following the
cloning approaches, the target gene was cloned and expressed either in E. coli or
Saccharomyces cerevisiae, thus fragrance producing recombinants were produced.
Microbial synthesis of aroma compounds which is classified into different groups is
described in the next section.
7.3.2.1 Fruity and Floral Terpenes
Terpenes are the flavoured components of essential oils, consisting of five carbon
isoprene units which are assembled to each other, while terpenoids are modified
class of terpenes with different functional groups and oxidized methyl group at
various C positions (Perveen 2018). Only a few terpenes have been reported to be
produced by de novo methods using microorganisms, usually fungi belonging to
group Ascomycetes and Basidiomycetes (Gupta et al. 2015). Fungi such as
Kluyveromyces lactis and Ceratocystis moniliformis produce de novo fruity and
floral flavoured terpenes such as citronellol, linalool and geraniol (Drawert and
Barton 1978; Bluemke and Schrader 2001). However, the methods involving biotransformation of terpenes are milder and produce less toxic wastes and, hence, are
better alternative for the production of natural aroma compounds (Dionísio et al.
2012; Bier et al. 2011).The pathway for biotransformation of geraniol into methyl
heptenone has been elucidated for Penicillium digitatum and citral lyase enzyme
7 Biotechnological Interventions for Production of Flavour and Fragrance. . .
137