Biotransform
677
Part E | 28
28. Biotransformation of Nitriles by Marine Fungi
Julieta Rangel de Oliveira, Mirna H. Reagli Seleghim, André L. Meleiro Porto
In this study, a screening of 12 marine fungi
(Penicillium miczynskii CBMAI 930, Penicillium
raistrickii CBMAI 931, Aspergillus sydowii CBMAI
933, Aspergillus sydowii CBMAI 934, Aspergillus
sydowii CBMAI 935, Bionectria sp. CBMAI 936,
Penicillium oxalicum CBMAI 1185, Penicillium
citrinum CBMAI 1186, Penicillium decaturense
CBMAI 1234, Penicillium raistrickii CBMAI 1235,
Cladosporium sp. CBMAI 1237, and Aspergillus
sydowii CBMAI 1241) was conducted in order
to evaluate the enzymatic potential of these
microorganisms for biotransformation of phenylacetonitrile 1. These microorganisms were isolated
from sponges and algae collected on the northern part of the coast of São Paulo State, Brazil.
The screening was carried out on a solid mineral medium supplemented with glucose and
phenylacetonitrile 1 as the only source of nitrogen. Afterwards, the microorganisms adapted
were tested in a liquid medium containing
20, 40, and 60 L of phenylacetonitrile 1. The
phenylacetonitrile 1 was biotransformed into
the 2-hydroxyphenylacetic 1a acid by eight
of the marine fungi selected. The enzymes
which hydrolyzed nitriles in these catalytic systems were inducible. Since the mycelium of
A. sydowii CBMAI 934 grew strongly in solid
and liquid mineral media in the presence of
phenylacetonitrile 1, this fungus was selected for
the enzymatic hydrolysis reactions using other
nitriles, such as 4-fluorophenylacetonitrile 2,4chlorophenylacetonitrile 3, 4-methoxyphenylacetonitrile 4, cyclohexenylacetonitrile 5, and
2-cyanopyridine 14, yielding their corresponding carboxylic acids: 4-fluorophenylacetic 2a
(51%), 4-chlorophenylacetic 3a (55%), 4methoxyphenylacetic 4a (43%), cyclohexenylacetic acid 5a (28%), and the amide, 2pyridinecarboxamide 14a, respectively. This chapter reports on the study on the biotransformation
of nitriles by marine microorganisms which is summarized in the context of directions for future
research.
28.1 Overview ............................................. 678
28.1.1 Biocatalysis and
Biotransformation ..................... 678
28.1.2 Marine Microorganisms .............. 678
28.1.3 Nitriles ..................................... 679
28.1.4 Nitrilases, Nitrile Hydratases,
and Amidases ........................... 682
28.1.5 Application of Nitrilases,
Nitrile Hydratase, and Amidases
to Biocatalysis........................... 684
28.1.6 Biodegradation of Nitriles .......... 689
28.2 Experimental Methods ......................... 691
28.2.1 Isolation, Identification,
and Preparation of Stock Cultures
of Marine Fungi......................... 691
28.2.2 Cultivation of Marine Fungi
on a Solid Medium
in the Presence
of Phenylacetonitrile 1 ............... 691
28.2.3 Biotransformation
of Phenylacetonitrile 1
by Marine Fungi
in a Liquid Medium ................... 691
28.2.4 Isolation
of 2-Hydroxyphenylacetic Acid 1a
from Culture Media Produced
by A. sydowii CBMAI 934 ............ 691
28.2.5 Biotransformation and Isolation
of Phenylacetonitrile
Derivatives 2–4 by A. sydowii
CBMAI 934................................. 692
28.2.6 Biotransformation
of Cyclohexenylacetonitrile 5
by A. sydowii CBMAI 934 ............ 693
28.2.7 Biotransformation
of Aromatic Nitriles.................... 693
28.2.8 Growth of Marine Fungus
A. sydowii CBMAI 934 in the
Presence of 2-Cyanopyridine 14
in Solid and Liquid Media .......... 694
677
Part E | 28
28. Biotransformation of Nitriles by Marine Fungi
Julieta Rangel de Oliveira, Mirna H. Reagli Seleghim, André L. Meleiro Porto
In this study, a screening of 12 marine fungi
(Penicillium miczynskii CBMAI 930, Penicillium
raistrickii CBMAI 931, Aspergillus sydowii CBMAI
933, Aspergillus sydowii CBMAI 934, Aspergillus
sydowii CBMAI 935, Bionectria sp. CBMAI 936,
Penicillium oxalicum CBMAI 1185, Penicillium
citrinum CBMAI 1186, Penicillium decaturense
CBMAI 1234, Penicillium raistrickii CBMAI 1235,
Cladosporium sp. CBMAI 1237, and Aspergillus
sydowii CBMAI 1241) was conducted in order
to evaluate the enzymatic potential of these
microorganisms for biotransformation of phenylacetonitrile 1. These microorganisms were isolated
from sponges and algae collected on the northern part of the coast of São Paulo State, Brazil.
The screening was carried out on a solid mineral medium supplemented with glucose and
phenylacetonitrile 1 as the only source of nitrogen. Afterwards, the microorganisms adapted
were tested in a liquid medium containing
20, 40, and 60 L of phenylacetonitrile 1. The
phenylacetonitrile 1 was biotransformed into
the 2-hydroxyphenylacetic 1a acid by eight
of the marine fungi selected. The enzymes
which hydrolyzed nitriles in these catalytic systems were inducible. Since the mycelium of
A. sydowii CBMAI 934 grew strongly in solid
and liquid mineral media in the presence of
phenylacetonitrile 1, this fungus was selected for
the enzymatic hydrolysis reactions using other
nitriles, such as 4-fluorophenylacetonitrile 2,4chlorophenylacetonitrile 3, 4-methoxyphenylacetonitrile 4, cyclohexenylacetonitrile 5, and
2-cyanopyridine 14, yielding their corresponding carboxylic acids: 4-fluorophenylacetic 2a
(51%), 4-chlorophenylacetic 3a (55%), 4methoxyphenylacetic 4a (43%), cyclohexenylacetic acid 5a (28%), and the amide, 2pyridinecarboxamide 14a, respectively. This chapter reports on the study on the biotransformation
of nitriles by marine microorganisms which is summarized in the context of directions for future
research.
28.1 Overview ............................................. 678
28.1.1 Biocatalysis and
Biotransformation ..................... 678
28.1.2 Marine Microorganisms .............. 678
28.1.3 Nitriles ..................................... 679
28.1.4 Nitrilases, Nitrile Hydratases,
and Amidases ........................... 682
28.1.5 Application of Nitrilases,
Nitrile Hydratase, and Amidases
to Biocatalysis........................... 684
28.1.6 Biodegradation of Nitriles .......... 689
28.2 Experimental Methods ......................... 691
28.2.1 Isolation, Identification,
and Preparation of Stock Cultures
of Marine Fungi......................... 691
28.2.2 Cultivation of Marine Fungi
on a Solid Medium
in the Presence
of Phenylacetonitrile 1 ............... 691
28.2.3 Biotransformation
of Phenylacetonitrile 1
by Marine Fungi
in a Liquid Medium ................... 691
28.2.4 Isolation
of 2-Hydroxyphenylacetic Acid 1a
from Culture Media Produced
by A. sydowii CBMAI 934 ............ 691
28.2.5 Biotransformation and Isolation
of Phenylacetonitrile
Derivatives 2–4 by A. sydowii
CBMAI 934................................. 692
28.2.6 Biotransformation
of Cyclohexenylacetonitrile 5
by A. sydowii CBMAI 934 ............ 693
28.2.7 Biotransformation
of Aromatic Nitriles.................... 693
28.2.8 Growth of Marine Fungus
A. sydowii CBMAI 934 in the
Presence of 2-Cyanopyridine 14
in Solid and Liquid Media .......... 694