Handbook of Size Exclusion Chromatography and Related ...

thisfactisprovidedbytheelutionprofilesoflow-anglescattering(Fig.18),which
do not approach baseline for several void volumes.
Foraqueousdissolvedglucans(dn/dc) 630isintherange0.150 +0.03mL/g
and no major error is introduced working with this value for starch glucans. Any
possible small error is partially compensated by computation of absolute
molecular weights from mass and laser light-scattering experiments if
concentration (DRI-profile) and optical constant (LS-profile) are also processed
with this value.
Ifsampleconcentration (conc) isknown andrecoveryfrom theSECsystem
can be assumed to be 100%, specific refractive index increment may be
determined from area of the SEC elution profiles according to Eq. (5):
dn
¼DRI cal const DRI sens
dc l
area DRI
conc loop vol
For DMSO-dissolved native starch glucans atypical recovery of 30–60% will be
obtained with aqueous SEC eluents. Recovery increases with increasing
dissolution process; however, no preferential dissolution of individual glucans is
observed: normalized DRI-eluograms (mass ev), which are computed from raw
data DRI elution profiles (raw mass) according to Eqs (6) and (7) for several
dissolutionperiodsformorethan7daysmatchwithinexperimentalerror(Fig.13).
raw mass
mass ev ¼ Ð (6)
int stop
int startraw mass
ð int stop
int start
(5)
mass ev ¼1:0 (7)
where mass ev ¼normalized elution profile of mass fractions, raw mass ¼
elution profile of mass fractions (raw data), and int start, int stop ¼integration
range ¼limits of selective separation range.
Molecular weightsfor starchglucansmay beobtainedfrom SECseparation
combinedwith mass detection bymeans ofmolecular weight calibration obtained
either from peak position calibration or broad standard calibration, for instance
with dextrans (Fig. 14). Results, however, are relative results in terms of
calibration material (e.g., dextran) equivalent molecular weights. Typically,
molecular weight distributions are visualized as normalized (area ¼1.0)
differential distribution of mass fractions (m MWD d) according to Eq. (8) and
molar (number) fractions (n MWD d) according Eq. (9) (Fig. 15).
m(M) i ¼ dm(M)
dM
with
ð 1
m(M)
n(M) i ¼ Ð i=M
1
0 [m(M) i=M] dM with
© 2004 by Marcel Dekker, Inc.
0
m(M) dM ¼ m MWD d ¼ 1:0 (8)
ð 1
0
n(M) dM ¼ n MWD d ¼ 1:0 (9)