Chemistry Reference
In-Depth Information
12.2.2
Solid phase emission (fluorescence) procedures
In solid phase spectrofluorimetric procedures, a similar relationship between the analytical signal and the
initial concentration of analyte in solution can be established. The fluorescence signal of the analyte sorbed
on the solid support,
I
, is given by the following expression:
I
=
F
F
I
0
e
S
l
S
Cs
(12.8)
where
f
F
is the fluorescence efficiency in the solid phase,
I
o is the intensity of the excitation light beam and
e
S,
l
S,
and
Cs
are as above described. From Equations (12.4) and (12.8), it follows:
I
=
F
F
I
0
e
S
l
S
C
O
V1000
/(
m
+
V
/
D
)
(12.9)
and if the fraction V/D is neglected, as it usually occurs:
I
=
F
F
I
0
e
S
l
S
C
O
V1000/m
(12.10)
indicating a dependence of the fluorescence signal on
m
and
V
similar to that found for solid phase molecular
spectrophotometry.
In flow mode, similar relations to previous described equations can be obtained. However, there is an
additional advantage: when the system is flowing for a few seconds, the packing keeps constant and the
baseline shows a constant signal, so the spectrophotometric solid phase based measurements can be performed
at only one wavelength [11].
12.3
Batch mode procedures
SPMS performed in the UV region is an easy approach used when the analyte shows intrinsic absorption in
the UV region and can be selectively retained on the solid microbeads. No reagent is needed for the procedure
except a buffer solution which is often required to set the optimum sample solution pH value to allow the
optimized analyte retention. Therefore, the waste generated does not contain any potential hazardous reagent.
Moreover, the solid support can be regenerated and reutilized, although this advantage usually is not exploited
by the authors.
Interestingly, a key feature in UV-SPMS is the compatibility of the solid support with the nature of the
detector. While Sephadex exchanger gels or Silica C
18
are, in practical terms, transparent enough in the UV
region to allow measurements compatibility [9], other support such as aromatic polymers (type Dowex or
Amberlite) cannot be used for UV-SPMS [11]. In batch mode, UV-SPMS has been applied for the
determination of both cationic and anionic species in a wide variety of matrices. It must be noted that the
main applications reported are limited to active pharmaceutical formulations [12, 13]. The developed
procedures represent a simple GAC alternative to other procedures for quality control which include the use
of more sophisticated and/or less environmental friendly methods, even official ones. In these cases, relatively
low sample volume is used (typically 10 ml) and, consequently, low equilibration time is needed.
An illustrative example of UV-SPMS application is the microdetermination of thiamine (vitamin B
1
) in the
presence of other B-group vitamins [9]. Its cationic nature allows the selective retention on Sephadex CMC-
25 cation-exchange gel microbeads. The effect of solid phase amount is shown in Figure 12.1. Sensitivity
(expressed as apparent molar absorptivity) increases from 5.63 × 10
5
to 2.19 × 10
7
l mol
-1
cm
-1
for a 100-fold
sample volume increase.
