184
M. R. Nikolic et al.
Beer and Eshel (1985) proposed equations for calculation of concentration
(mg/mL), which are not affected by the concentration of interfering components,
and which are presently mainly used for determination of PBPs from red algae:
[PE] = ((A 564 − A 592 ) − (A 455 − A 592 ) 0.20) 0.12;
[PC] = ((A 618 − A 645 ) − (A 592 − A 645 ) 0.15) 0.15.
Finally, Sampath-Wiley and Neefus (2007) described equations for estimation
of PBPs content in aqueous extracts (mg/mL), which are, according to the authors,
more accurate than previously published methods:
[R − PC] = 0.154(A 618 − A 730 );
[R − PE] = 0.1247((A 564 − A 730 ) − 0.4583(A 618 − A 730 )).
For quantification of PCB obtained by methanolysis of purified PC, the extinction
coefficient of 37900 M
−1 cm
−1 in MeOH/HCl solution (Cole et al. 1967) is frequently
used.
While chromophore is a light-sensitive part of PBPs, apoprotein part confers the
stability with respect to pH and temperature. Typically, the thermostability, the effect
of pH itself, or the effect of pH on thermal stability of the PBPs are measured by incubating protein samples at chosen temperatures/pH values, followed by measurements
of the characteristic absorbance maximum at regular time intervals, and calculation of
the remaining concentration of PBP (C R , %) relative to the initial concentration (e.g.,
Rahman et al. 2017; Wu et al. 2016; González-Ramírez et al. 2014; Liu et al. 2009).
Differences in denaturation midpoint, defined as the temperature (Tm) at which 50%
of the protein still remains in solution (C R = 50% value), the purity ratio (e.g.,
A 620 /A 280 for PC), and the half-life value (t 1/2 , the time taken for the initial protein
concentration to be reduced by half) are also used to compare the stability of PBPs
from various algal species. The examination of conformational state and functional
dynamics of PBPs by measurements of optical properties of chromophores have been
recently applied to compare the stability of the full length and truncated α-subunit of
cyanobacterial PE. Urea-induced denaturation α-subunit transitions were monitored
and the role of the truncated region in PE stability was also investigated by molecular
dynamics simulations (MDS) (Anwer et al. 2015).
Improving the stability of PBPs is an important goal for their practical application as natural colors in the food and cosmetic industry and as a fluorescent probe
and analytical reagent (Stanic-Vucinic et al. 2018). PBPs spectroscopic properties,
including degradation kinetics with determination of degradation rate constant (k d ),
are wildly used to examine improvement of PBPs storage stability by encapsulation
(Suzery et al. 2015; Purnamayati et al. 2018), nanofiber encapsulation, and/or in the
presence of preservatives (Braga et al. 2016; Bhattacharya et al. 2018).
M. R. Nikolic et al.
Beer and Eshel (1985) proposed equations for calculation of concentration
(mg/mL), which are not affected by the concentration of interfering components,
and which are presently mainly used for determination of PBPs from red algae:
[PE] = ((A 564 − A 592 ) − (A 455 − A 592 ) 0.20) 0.12;
[PC] = ((A 618 − A 645 ) − (A 592 − A 645 ) 0.15) 0.15.
Finally, Sampath-Wiley and Neefus (2007) described equations for estimation
of PBPs content in aqueous extracts (mg/mL), which are, according to the authors,
more accurate than previously published methods:
[R − PC] = 0.154(A 618 − A 730 );
[R − PE] = 0.1247((A 564 − A 730 ) − 0.4583(A 618 − A 730 )).
For quantification of PCB obtained by methanolysis of purified PC, the extinction
coefficient of 37900 M
−1 cm
−1 in MeOH/HCl solution (Cole et al. 1967) is frequently
used.
While chromophore is a light-sensitive part of PBPs, apoprotein part confers the
stability with respect to pH and temperature. Typically, the thermostability, the effect
of pH itself, or the effect of pH on thermal stability of the PBPs are measured by incubating protein samples at chosen temperatures/pH values, followed by measurements
of the characteristic absorbance maximum at regular time intervals, and calculation of
the remaining concentration of PBP (C R , %) relative to the initial concentration (e.g.,
Rahman et al. 2017; Wu et al. 2016; González-Ramírez et al. 2014; Liu et al. 2009).
Differences in denaturation midpoint, defined as the temperature (Tm) at which 50%
of the protein still remains in solution (C R = 50% value), the purity ratio (e.g.,
A 620 /A 280 for PC), and the half-life value (t 1/2 , the time taken for the initial protein
concentration to be reduced by half) are also used to compare the stability of PBPs
from various algal species. The examination of conformational state and functional
dynamics of PBPs by measurements of optical properties of chromophores have been
recently applied to compare the stability of the full length and truncated α-subunit of
cyanobacterial PE. Urea-induced denaturation α-subunit transitions were monitored
and the role of the truncated region in PE stability was also investigated by molecular
dynamics simulations (MDS) (Anwer et al. 2015).
Improving the stability of PBPs is an important goal for their practical application as natural colors in the food and cosmetic industry and as a fluorescent probe
and analytical reagent (Stanic-Vucinic et al. 2018). PBPs spectroscopic properties,
including degradation kinetics with determination of degradation rate constant (k d ),
are wildly used to examine improvement of PBPs storage stability by encapsulation
(Suzery et al. 2015; Purnamayati et al. 2018), nanofiber encapsulation, and/or in the
presence of preservatives (Braga et al. 2016; Bhattacharya et al. 2018).