To investigate diffusion processes in agarose gel, nanoparticles with sizes in the range between 1 and 140 nm have been tested by means of fluorescence correlation spectroscopy. Understanding the diffusion properties in agarose gels is interesting, because such gels are good models for microbial biofilms and cells cytoplasm. The fluorescence correlation spectroscopy technique is very useful for such investigations due to its high sensitivity and selectivity, its excellent spatial resolution compared to the pore size of the gel, and its ability to probe a wide range of sizes of diffusing nanoparticles. The largest hydrodynamic radius (R(c)) of trapped particles that displayed local mobility was estimated to be 70 nm for a 1.5% agarose gel. The results showed that diffusion of particles in agarose gel is anomalous, with a diverging fractal dimension of diffusion when the large particles become entrapped in the pores of the gel. The latter situation occurs when the reduced size (R(A)/R(c)) of the diffusing particle, A, is >0.4. Variations of the fractal exponent of diffusion (d(w)) with the reduced particle size were in agreement with three-dimensional Monte Carlo simulations in porous media. Nonetheless, a systematic offset of d(w) was observed in real systems and was attributed to weak nonelastic interactions between the diffusing particles and polymer fibers, which was not considered in the Monte Carlo simulations.
Fluorescence correlation spectroscopy (FCS) was used to
determine diffusion coefficients (D) of the Suwannee
River fulvic and humic acids (SRFA and SRHA) and to
measure the effects of pH, ionic strength, Ca, and humic
substance (HS) concentration on the value of D. For these
HS, average diffusion coefficients were in the range of (2−3) × 10-10 m2 s-1 (corresponding to hydrodynamic
diameters of approximately 1.5−2.1 nm). Small, but
significant, decreases in the diffusion coefficients were
observed with decreasing pH, most likely indicating that a
small degree of aggregation (formation of dimers and
trimers) was occurring. The effect of ionic strength (up to
100 mM) was either small or insignificant, but, where a
change occurred, there was a tendency toward a reduction
in the diffusion coefficient with increasing salt concentration.
No effect of HS concentration (1−50 mg L-1) or hydration
time (1−14 days) was observed. There were no observable
effects of Ca in comparison to Na, at an ionic strength
of 5 mM. For all conditions studied, SRFA had a higher
diffusion coefficient than SRHA which is in agreement with
literature data on their respective molar masses. Since
this is the first application of FCS to HS, the technique was
systematically tested for artifacts, in particular photo
bleaching and the effect of fluorescence excitation
wavelength. Despite a small amount of photobleaching
due to the intense laser excitation, FCS was found to be
suitable for use with fulvic or humic acids.
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