Characterization of Humic Substances by Environmental Scanning Electron Microscopy

Abstract

Environmental scanning electron microscopy (ESEM) is a new technique capable of imaging micron and submicron particles. Here, we have applied it to image and quantify natural aquatic organic matter (standard Suwannee River humic acid, SRHA). Uniquely, we have observed the humic aggregate structures as a function of humidity and pH. Large aggregates of tens of micrometers were observed as the dominant material under all conditions, although much smaller material was also observed. Fractal dimensions (D) were calculated between 1.48 and 1.70, although these values were not statistically different under conditions of low humidity. However, D values calculated at high humidities (85%) during the rehydration phase were significantly lower (1.48+/-0.01) than in the initial dehydration phase (1.69+/-0.01). This hysteresis indicated that full rehydration of the HS was either kinetically slow or irreversible after dehydration. Fractal analysis of ESEM images was also performed to probe the change in aggregate structure as a function of pH. Minimum values were calculated at neutral pHs, rising by 0.1-0.2 at both high and low pHs because of a combination of the physical chemistry of HS and the impacts of the drying regime within the ESEM. Thus, ESEM was an important complementary technique to other analytical methods. At present, ESEM cannot be used to image nonperturbed natural samples. However, the method is an ideal method for probing the changes in colloid structure as function of hydration state and has the potential to perform fully quantitative and nonperturbing analysis of colloidal structure.

Visualisation of natural aquatic colloids and particles ? a comparison of conventional high vacuum and environmental scanning electron microscopy

“…However, care must be taken not to drastically change the nature of the samples. The present work showed no evidence that changes in the sample structure or aggregation had taken place, but Redwood et al 29 demonstrated that material such as humic substances, the major organic constituents of soils and aquatic environments, 36,37 tend to aggregate when subjected to dehydration. Fig.…”

“…Another study that quantified the structures of standard Suwannee River humic acids using fractal dimensions 29 also observed the formation of large crystals of NaCl during dehydration of their NaCl-based sample matrix in ESEM, although those appeared differently shaped than in this study.…”

“…32 It is capable of a resolution limit of ca. 30 nm at a relative humidity as high as 65%, 29 which is slightly higher than with SEM (ca. 1 nm).…”

“…30 nm at a relative humidity as high as 65% although imaging must be performed very carefully. 29 However, many environmental particles are relatively amorphous and gel-like and our experience in this work suggests that a resolution limit of about 50 nm is realistic here (Fig. 5D, which is a highly magnified version of Fig.…”

Size fractionation and characterization of nanocolloidal particles in soils

“…Moreover, the low-density particles in the fractions would have had a relatively large size distribution. Electron microscope imaging of colloidal fractions Both TEM and SEM have often been used to visualise the size and shape of aquatic colloids and other particles (Wilkinson et al 1999;Redwood et al 2005). Here, the purified nanocolloid fractions were analyzed by TEM to validate laser granulometry measurements and assess the capability of CFF to retain nanocolloidal particles.…”