8. Construct a plot in which the sample temperature (y, in
C) is
plotted versus the time point in the temperature run (x, in s);
and fit using a linear regression line (y ¼ ax+b, where a represents the actual cooling/warming rate in
C/s and b the offset
or start temperature in
C) (see Note 15).
9. List the time points (x, in s) at which spectra were recorded
during the scan (each labeled with its own #), and calculate the
actual temperature at which they were collected (y, in
C), using
the linear regression equation determined above. In a spreadsheet program, enter column listing: (1) spectra numbers,
(2) time points of collection, and (3) calculated sample temperature during collection. Results from spectral analysis can be
added later, after exporting the spectra using the spectra acquisition software.
10. Extract and save the individual spectra (each labeled with its
own #) that are acquired during the temperature run, in a
separate folder; label with date (and experiment run number).
3.5 Spectral
Analysis: Diffusion
Coefficients
of Protective
Molecules Permeating
into Tissues
Permeation of molecules into tissues can be evaluated by analyzing
characteristic spectral bands or regions in the FTIR spectra, which
are specific for a particular compound. These regions are typically
selected in the 1500–900 cm
À1 wave number range. Also components in mixtures can be simultaneously monitored by simultaneous monitoring of specific bands that have no overlap (see
Fig. 5a). Permeation kinetics can be evaluated by plotting the area
of characteristic bands as a function of the time (see Fig. 5b). The
data in such plots can be fitted using established mathematical models to derive specific diffusion coefficients:
1. Use tissue saturated with the component of interest (overnight
equilibration) to assign specific bands and spectral regions that
can be used to monitor permeation as a function of time. In
order to study compounds in mixtures, select spectral regions
for each compound that do not overlap with each other.
2. Use the following procedure to calculate baseline-corrected
band areas of selected regions for each spectrum (each labeled
with its own #):
(a) Open spectrum and select the characteristic region for the
protective compound of interest.
(b) Determine the baseline-corrected band area using the
beginning and end point of the selected region, and
save it.
(c) Steps (a) through (b) need to be applied to all the spectra
that have been recorded during the run. For mixtures, this
has to be done for all of the components.
340
Willem F. Wolkers and Harrie ¨ tte Oldenhof
C) is
plotted versus the time point in the temperature run (x, in s);
and fit using a linear regression line (y ¼ ax+b, where a represents the actual cooling/warming rate in
C/s and b the offset
or start temperature in
C) (see Note 15).
9. List the time points (x, in s) at which spectra were recorded
during the scan (each labeled with its own #), and calculate the
actual temperature at which they were collected (y, in
C), using
the linear regression equation determined above. In a spreadsheet program, enter column listing: (1) spectra numbers,
(2) time points of collection, and (3) calculated sample temperature during collection. Results from spectral analysis can be
added later, after exporting the spectra using the spectra acquisition software.
10. Extract and save the individual spectra (each labeled with its
own #) that are acquired during the temperature run, in a
separate folder; label with date (and experiment run number).
3.5 Spectral
Analysis: Diffusion
Coefficients
of Protective
Molecules Permeating
into Tissues
Permeation of molecules into tissues can be evaluated by analyzing
characteristic spectral bands or regions in the FTIR spectra, which
are specific for a particular compound. These regions are typically
selected in the 1500–900 cm
À1 wave number range. Also components in mixtures can be simultaneously monitored by simultaneous monitoring of specific bands that have no overlap (see
Fig. 5a). Permeation kinetics can be evaluated by plotting the area
of characteristic bands as a function of the time (see Fig. 5b). The
data in such plots can be fitted using established mathematical models to derive specific diffusion coefficients:
1. Use tissue saturated with the component of interest (overnight
equilibration) to assign specific bands and spectral regions that
can be used to monitor permeation as a function of time. In
order to study compounds in mixtures, select spectral regions
for each compound that do not overlap with each other.
2. Use the following procedure to calculate baseline-corrected
band areas of selected regions for each spectrum (each labeled
with its own #):
(a) Open spectrum and select the characteristic region for the
protective compound of interest.
(b) Determine the baseline-corrected band area using the
beginning and end point of the selected region, and
save it.
(c) Steps (a) through (b) need to be applied to all the spectra
that have been recorded during the run. For mixtures, this
has to be done for all of the components.
340
Willem F. Wolkers and Harrie ¨ tte Oldenhof