Enhanced visualization of polysaccharides from aqueous suspensions

Abstract

Abstract. Aqueous suspensions of polysaccharides such as those prepared for domestic and industrial applications or present in natural waters, although difficult to visualize by conventional transmission electron microscopy (TEM) because of their poor electron density, can be characterized at the ultrastructural level by using mild en bloc staining and contrast enhancement by energy-filtered TEM (EF-TEM). The advantages and drawbacks of the proposed method are discussed in relation to the different parameters controlling the quality of final images. It is shown, with synthetic polysaccharides, purified algal fibrils and lacustrine exocellular polymers as key examples, that optimizing specimen preparation and visualization parameters allows unbiased identification of organic substructures never revealed or strongly degraded by classical microscopic procedures.Key words: fibrillar polysaccharides, contrast enhancement, energyfiltered transmission electron microscopy (EF-TEM), densitometry, stain, uranyl acetate.Carbohydrates in nature show high diversity and complexity in their composition, structure and function [1,2]. They are produced as neutral or acidic compounds by all living species, from micro-organisms to higher animal cells, and it is not surprising to detect them in our environment (soils, waters), as a consequence of exocellular release (extracellular polysaccharides, EPS) or degradation process in the natural reservoirs [3].Because of their thickening properties, native or derivatized polysaccharide biopolymers from botanical and microbial sources have found widespread * To whom correspondence should be addressed applications for industrial, domestic and environmental uses where rheology, gelling or pseudoplasticity is of concern [4,5].The behaviour of polysaccharides in synthetic formulations and natural suspensions will primarily depend on their macromolecular structure, varying from totally linear (cellulose) to strongly branched (spherocolloids like amylopectin or gum arabic) compounds; due to minimal steric hindrances, the former have a strong tendency to pack into water-insoluble thick fibres, while the latter are usually highly hydrated (hydrocolloids) and form gels which can contain up to 99.5% water [6].Polysaccharides have a high complexing capacity for di-and trivalent ions [7,8]. Furthermore, the hydroxyl groups of polysaccharides tend to interact with the hydrated surfaces of dispersed mineral particles (oxyhydroxides) and with hydrophilic organic material by hydrogen binding, and to form biofilms on submerged surfaces [9,10]. These properties are demonstrated in soils, where fungal polysaccharides act as binding/aggregating agents between clay particles and as water stabilizers [11][12][13]. Polysaccharides are used in water treatment plant for the removal of undesirable substances by cross-linking, micro-gel formation and flocculation. However, the role of EPS on the stability of suspended mineral particles and on the transport processes of micropollutants in freshwaters and marine envi...

Internucleobase-Interaction-Directed Self-Assembly of Nanofibers from Homo- and Heteroditopic 1,ω-Nucleobase Bolaamphiphiles

“…To precisely characterize the resultant morphologies and their size dimensions, we observed the individual self-assembled structures using energy-filtering transmission electron microscopy (EF-TEM) . Unlike conventional TEM, EF-TEM provides a useful tool for observing low-contrast organic samples without the need for prior staining . The EF-TEM for the helical ropes of T-10-T also showed clear images similar to those observed using light microscopy (Figure b and c).…”

Identification of stoichiometric iron-phosphorus colloids produced in a eutrophic lake

“…In order to determine with a high accuracy the molar ratio [PO 4 ] part : [Fe] part in individual iron-rich particles, hydrous iron oxide colloids containing phosphates were synthetised under chemical conditions similar to the lake and used as EDS calibration standards: Fe-P-rich colloids were obtained by slow (ca. 12 h) oxidation of a deoxygenated ([O 2 ] t0 < 0.5 mg/l) solution containing [Fe 2+ ] 10 mM in [Ca(HCO 3 ) 2 ] 1.5 mM, [H 2 PO 4 -] 1.25-5 mM and polysaccharide fibrils (xanthan 1 mg/l produced by Xanthomonas campestris; Lienemann et al, 1997) • Direct in situ collection of particulate phases: Machined Teflon plates (f = 8 cm) holding TEM gold grids (collodion coated, carbon covered) were vertically inserted in the 60-120 m layer of the water column for occasional periods varying between one and two months; this procedure allowed for the direct in situ recovery of freshly formed and reactive non-sedimenting particles for microscopic (TEM grids) and bulk chemical (Teflon plates) analyses. The plates were retrieved and the particulate material adhering onto their surface was mineralized (HCl 10 -1 M for colorimetry, HNO 3 3% + H 2 NOH • HCl 10 -1 M for ICP-AES; 24 h stirring at room temperature) prior to analysis by colorimetry (PO 4 ) and ICP-AES (Fe, Mn).…”

Texture analysis of TEM micrographs of alginate gels for cell microencapsulation

“…It is many years since TEM analysis of polysaccharides in solution and gel has been performed (Lamberti and Sefton, 1983;Lienemann et al, 1997), although images of polysaccharide mixtures are not so common (Willenberg et al, 2006;Yongsheng et al, 2008), and the quality is quite often not outstanding. At the best of our knowledge, quantitative analysis of the TEM image of alginate gels was previously reported only by one group Rayment et al, 2009).…”