The Induction of Birefringence in Pellicles of Bacterial Cellulose From Acetobacter Xylinum by Lipids

Abstract

Soine lipids, particularly diglycerides, cholesterol, or cholesterol derivatives, are able to induce visible birefringence in pellicles of estracellular bacterial cellulose produced by Acetobacter xylinzrnz when they are added in snlall amounts to the surface of cc~ltures. This birefringence is due to orientation of a fraction of the cellulose microfibrils, not the cells, in the pellicle. Orientation may begin with small microscopic domains which are present in pellicles of untreated cultures and some of which inay overgrow others under the influence of a lipid. How the lipids induce alignment of the microfibrils in the bacterial cult~ire is unknown but the process appears to involve an interaction between ilemly formed extracellular microfibrils and the ~nicrocrpstalline surface of the lipid. IntroductionDiffuse and weak areas of birefringence in pellicles of bacterial cellulose have been recognized for a long time but have not been investigated systernatically, probably because they were attributed to slight stretching or compression of films and were therefore only of trivial interest. Recently, two additional forms of birefringence in pellicles of cellulose from cultures of Acetobacter xylinz~m have beell investigated in more detail (Colvin 1965(Colvin , 1966 because they reflect definite orientation of extracellular cellulose microfibrils in a systenl where the factors causing alignment cannot be mechanical or hydrodynan~ic. The purpose of the present paper is to report that several classes of lipids may induce birefringence in pellicles produced by cultures of il. xylinum, t o show t h a t this birefringence is due t o partial orientation of the cellulose microfibrils, and to describe some of the properties of the cultures which are associated with this orientation. Materials and MethodsPellicles of bacterial cellulose were grown and their birefringence observed and recorded by methods previously described (Colvin 1965). Compounds were applied to the surfaces of static cultures containing 100 1111 of seeded medium per 15-cm Pctri plate either as an ethanol solution (1 ~ng/ml or a saturated solutioil if the solubility of the c o~~l p o u l~d in ethanol was below this limit) or as small crystals. When ethanol solutions were used, results were coinpared with cultures to which an equal volume of ethanol (10 drops) had been added a t the same time (12-24 11 after pouring the medium, when only a slight cloudilless had appeared without any forlllatio~l of a coherent pellicle). When crystals were used, 1-2 mg of the compound u7as dropped on surfaces of cultures im~nediately after the medium uras poured. Results were compared to plates to which nothing had been added. 'N.R.C. Contribution KO. 9472. Canadian Jourllal of Microbiology. Volume 13 (196;) Can. J. Microbiol. Downloaded from www.nrcresearchpress.com by McMaster University on 12/22/14 For personal use only.Sections of pellicles froin cultures were obtained as follows. Areas of interest in the pellicle were carefully dissected between crossed Polaroid filin...

Induction of orientation of bacterial cellulose microfibrils by a novel terpenoid from Acetobacter xylinum

“…At all stages of lipid fractionation, each portion was tested for its ability to induce birefringence in the freshly grown pellicle of A. xylinum cultures. As shown in previous studies (Colvin, 1965(Colvin, ,1966Carson et al, 1967) and confirmed in this investigation for the particular lipids involved (see the Results section) birefringence in the pellicle or a part of it implies partial orientation of the cellulose microfibrils within that part. Therefore whenever a compound or fraction is called active or biological activity is mentioned in the succeeding text, this refers to the capacity of the fraction to induce birefringence and mutual orientation of microfibrils during the assay procedure.…”

“…The white waxy mass, which was crystallized from fraction 8, had an activity that was qualitatively different from that shown by the alcohol from fraction 2 or by 22-hydroxyhopane, both of which induced the development of a wide-spread, diffuse birefringence. The substance in fraction 8 induced a much more localized birefringence, confined to narrow, branched lines and similar to that produced by cholesteryl oleate (Carson et al, 1967, Fig. 1).…”

“…All plates were observed between crossed Polaroid sheets periodically up to 48h and scored for birefringence on a scale from 0 to 5. This method is qualitative rather than quantitative and the type of birefringence observed differs with different compounds (Carson et al, 1967). The majority of compounds, however, induced the same type of birefringence (Carson et al, 1967, Fig.…”

“…Previous studies by optical microscopy and transmission electron microscopy have shown that birefringence in the pellicles of A. xylinum is associated with partial orientation of the component cellulose microfibrils (Colvin, 1965(Colvin, , 1966Carson et al, 1967). It was considered worthwhile to verify the association, taking advantage of a recently available scanning electron microscope.…”

“…Some of the cellulose microfibrils produced in the external 14S medium of Acetobacter are aligned as bands (Ben-Hayyim & Ohad, 1965), as parts of spherulites (Colvin, 1965), or as thread-like structures (Colvin, 1966). Enhanced orientation can be induced by the addition of small amounts of certain classes of lipids to the medium (Carson et al, 1967). The most-active compounds are sterols.…”