An investigation of agitation speed as a factor affecting the quantity and monomer distribution of alginate from Azotobacter vinelandii ATCC® 9046

Abstract

Alginate is a copolymer of β-D: -mannuronic and α-L: -guluronic acids. Distribution of these monomers in the alginate structure is one of the important characteristics that affect the commercial value of the polymer. In the present work, the effect of agitation speed in the range of 200-700 rpm on alginate production by Azotobacter vinelandii ATCC(®) 9046 was investigated at a dissolved oxygen tension of 5% of air saturation. Experiments were conducted in a fermentor operated in batch mode for 72 h while the production of biomass and alginate, the consumption of substrate and the change in culture broth viscosity and monomer distribution of the polymer were monitored. Results showed that the growth rate of the bacteria increased from 0.165 to 0.239 h(-1) by the increase of mixing speed from 200 to 400 rpm. On the other hand, alginate production was found to be the most efficient at 400 rpm with the highest value of 4.51 g/l achieved at the end of fermentation. The viscosity of culture broth showed similar trends to alginate production. Viscosity was recorded as 24.61 cP at 400 rpm while it was only 4.26 cP at 700 rpm. The MM- and GG-block contents were almost equal in most of the culture times at 400 rpm. On the other hand, GG-blocks dominated at both low and high mixing speeds. Knowing that GG-blocks make rigid and protective gels with divalent cations, due to the higher GG-block content, the gel formation potential is higher at 200 rpm as well at 700 rpm, which might originate from the unfavorable environmental conditions that the bacteria were exposed to.

Guluronic acid content as a factor affecting turbidity removal potential of alginate

“…All bacterial alginate samples were produced by Azotobacter vinelandii ATCC ® 9046 in modified Burk's medium (Moral and Sanin 2012), deacetylated by alkaline treatment, and purified by acid precipitation followed by ethanol precipitation as described earlier (Ertesvåg and Skjåk-Braek 1999). Samples coded as AS#1 -4 produced in a laboratory fermenter (BIOSTAT ® A plus, Sartorious BBI Systems, Germany) at constant dissolved oxygen tension, 5 %, and 30 °C during 72 hours of fermentation.…”

“…It is known that if M-blocks formed by consecutive mannuronic acid residues are dominant, soft and more flexible gels are formed; whereas if stretches of consecutive guluronic acid residues (G-blocks) are dominant, strong gels that are mostly resembled by an "egg-box structure" are formed (Grant et al 1973). Production of alginate from Azotobacter vinelandii has been widely studied by using shake flasks and fermenters (Clementi et al 1999;Diaz-Barrera et al 2010;Mejia et al 2010;Peña et al 1997;Savalgi and Savalgi 1992), and by differing the growth conditions, it is possible to reproducibly produce alginates with different content of G-blocks (Moral and Sanin 2012;Sabra et al 2000;Sabra et al 1999). A. vinelandii encode seven different secreted mannuronan C-5-epimerases (AlgE1-7) which introduce G residues in the polymer, and these may be used to increase the amount of G and G-blocks in vitro (Ertesvåg 2009).…”

Shaken flasks by resonant acoustic mixing in the biosynthesis of alginate by Azotobacter vinelandii with non‐Newtonian rheological characteristics

“…It is known that increasing the oxygen supply by increasing agitation intensity enhances alginate production in shake‐flask cultures,, and in stirred‐tank bioreactors;, , , this relation is conserved to a limit, after which enhancement of the agitation‐aeration conditions causes a decrease in alginate production , , , , …”

“…(e)). Oxygen‐limited conditions when growing A. vinelandii in shaken flasks has been widely documented,, , , , and this is related to the high respiration activity of this bacteria , , . In this sense, the DOT profile observed at 20 g, with no oxygen limitation, has not been reported for shake‐flask cultures of A. vinelandii .…”

“…The highest apparent viscosity of 15.47 ± 6.93 mPa.s was achieved by the cultures grown at 20 g , whereas at 5, 10, and 15 g , values of 3.03 ± 0.18, 5.22 ± 0.42, and 1.26 ± 0.03 mPa.s were obtained, respectively. The high viscosity attained at 20 g can be attributed to the accumulation of high concentrations of alginate, as viscosity is a function of alginate concentration, among other polysaccharide characteristics such as M/G ratio, molecular weight and acetylation , , , . .…”