Bacterial Enzymes and Multi-enzymatic Systems for Cleaning-up Dyes from the Environment - Microbial Degradation of Synthetic Dyes in Wastewaters - page 45

catalytic incompetent 6-coordinated low spin state in the Bs enzyme, while the
major population in PpDyP is the 5-co-ordinated quantum mechanically mixed spin
state, as observed by resonance Raman (Sezer et al. 2013 ).

4.2 The Catalytic Pathway for Biotransformation
of Anthraquinonic Dyes by DyPs

The transformation of the anthraquinonic dye reactive blue 5 was carried out using
DyP from Thanatephorus cucumeris Dec 1 (Sugano et al. 2009 ). Changes in the
visible spectrum of RB5 treated with DyP resulted in a decrease in the intensity of
the dye absorption band, at

λ max = 600 nm, along with an increase in absorption at

400 500 nm as the color of the solution became red-brown.

Analysis of the

-

nal enzymatic reaction mixtures by NMR and MS techniques
showed that the anthraquinone dye reactive blue 5 was transformed by DyP from T.
cucumeris Dec 1 to three reaction products detected by their distinct molecular ion
signals. The

ed as phthalic acid by comparison with an
authentic sample. The second one (2), with a molecular mass of 472 g mol − 1 , can be
attributed to a reactive blue 5 molecule without the anthraquinone frame (see
Fig. 16 ). Finally, the third product (3) can be obtained from compound (2) which
loss a 2,5-diaminobenzene sulfonic acid (ABS) molecule.
Based on these results, a reasonable degradation pathway of reactive blue 5 by
DyP was proposed as shown in Fig. 16 . The

rst product (1) was identi

nal red-brown color of the reaction
mixture of reactive blue 5 biotransformation and the absence of o-ABS and m-or
p-ABS as

nal products, suggest the presence of other products resulting from the
dimerisation and polymerization reactions of ABS type substrates by DyP action. In
fact, this was con rmed with the o-ABS reaction with DyP, leading to the formation

of high weight colored products, from which compound 4, containing an azo group,
was identi

ed.

4.3 Combined Sequential Enzymatic Treatment for Dye
Degradation and Detoxi

cation

In order to set-up enzymatic processes for maximal decolorization as well as
detoxi cation, a sequential enzymatic procedure was performed combining the

PpAzoR reduction of azo dyes to the oxidation of aromatic amines by CotA-
laccase. It is know that laccases catalyze the oxidation of ortho- or para-substituted
phenolic or aromatic amine substrates by one electron abstraction to form free
radicals that undergo further coupling, polymerization, demethylation, or quinone
formation (Abadulla et al. 2000 ; Kandelbauer and Guebitz 2005 ). In particular, we
have shown recently that CotA-laccase catalyzed the homocoupling of primary