ПРОДУКТИВНІСТЬ СИНТЕЗУ ЕКЗОПОЛІСАХАРИДУ BACILLUS MUCILAGINOSUS ЗАЛЕЖНО ВІД ДЖЕРЕЛА АЗОТУ І ПОХОДЖЕННЯ ПОСІВНОГО МАТЕРІАЛУ
It was established that Bacillus mucilaginosus C-3 requires introduction of nitrogen sourceinto the culture medium in form of nitrate as themost optimal for maximal accumulation of bacterial biomass and sporulation process. Introduction of the ammonium form was less efficient. It was shown that the...
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| Datum: | 2015 |
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Institute of Agrocultural Microbiology and Agro-industrial Manufacture of NAAS of Ukraine
2015
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| author | Малиновська, І. М. |
| author_facet | Малиновська, І. М. |
| author_institution_txt_mv | [
{
"author": "І. М. Малиновська",
"institution": "Національний науковий центр «Інститут землеробства НААН»"
}
] |
| author_sort | Малиновська, І. М. |
| baseUrl_str | https://smic.in.ua/index.php/journal/oai |
| collection | OJS |
| datestamp_date | 2026-07-22T10:10:42Z |
| description | It was established that Bacillus mucilaginosus C-3 requires introduction of nitrogen sourceinto the culture medium in form of nitrate as themost optimal for maximal accumulation of bacterial biomass and sporulation process. Introduction of the ammonium form was less efficient. It was shown that the intensity of nitrogensource use by B. mucilaginosus is highly depended on the physiological state of the inoculum cells. At sowing of spore inoculum theoptimal concentration of potassium nitrate was2.5 g/l resulting in 1010 cells/ml, while under theuse of vegetative cells the optimal concentrationof potassium nitrate was 1.0 g/l leading to theaccumulation of 109 cells/ml.Use of spore inoculum had ensured themaximum productivity of extracellular polysaccharide synthesis. With the higher number ofB. mucilaginosus subcultures in a vegetativestate the efficiency of the exopolysaccharidesynthesis has decreased: after the second passage — by 20.0 %, after the fourth — by 56.5 %,after the sixth — at 127.8 %. After eighth passage the culture loses its ability to synthesizepolysaccharide, especially on the medium withthe N : C ratio 1 : 3 and resume this ability onlyafter sporulation and the significant increase ofthe N : C ratio. |
| doi_str_mv | 10.35868/1997-3004.21.12-17 |
| first_indexed | 2025-07-17T12:24:03Z |
| format | Article |
| fulltext |
EXOPOLYSACCHARIDE SYNTHESIS PRODUCTIVITY
BY BACILLUS MUCILAGINOSUS DEPENDING
ON THE NITROGEN SOURCE AND ORIGIN OF INOCULUM
I. M. Malinovska
The extracellular polysaccharide Bacillus
mucilaginosus has broad prospects for the use
in practice: in medicine (immune-stimulating,
immunomodulatory, antiviral, antitumor
activity) and animal husbandry (food additive
in the diet of farm animals, preserving forage),
in oil and mining industry, in the production of
ceramics and lubricants, substitutes of known
technical oils, antiwear additive in brake and
cooling solutions [1-3]. But the works, devoted
to the study of principles of growth and
synthesis of extracellular polysaccharide B.
mucilaginosus, are controversial [4-7]. Thus,
according to V. H. Aleksandrov and
V. S. Podhorskyi et al. [5; 6], the potential
growth of culture on nitrogen-free
environments is possible. Most scientists
advocate the need for nitrogen administration
to the culture medium composition [4; 7]. The
most thorough study was conducted by
K. I. Surman [7]. She showed that B.
mucilaginosus belongs to oligonitrophils and
needs small amounts of nitrogen in the
ammonium form (0.03% NH4NO3) for its
development. The contradiction between the
researchers is not only on the need of nitrogen
administration to the culture medium, but also
on its form. Thus, V. H. Aleksandrov and
H. A. Zak [6] believe that organic nitrogen
inhibits the growth of bacteria. Other
researchers argue that peptone (0.05%) and
yeast water (2.0%) stimulate the growth and
sporogenesis of B. mucilaginosus [1; 2].
The divisiveness of obtained data is,
possibly, associated with conducting
experiments with different strains of B.
mucilaginosus, or a mixture of options that are
different in physiological and biochemical
properties. There is a need for research
extension in the direction of physiological and
biochemical characteristics of individual
strains of B. mucilaginosus.
Materials and methods. The subject of the
research was C-3 strain of B. mucilaginosus
obtained from the collection of the Institute of
Mineral Resources of the Ministry of Geology
of Ukraine, where it was given to in 1963 by
V. H. Alexandrov. For the cultivation of
bacteria A-27 medium was used, g/l: sucrose -
5.0; MgSO4x7H2O – 0.5; Na2HPO4x2N2O -
2.0; FeCl3x6H2O – 0.05; potassium aluminium
silicate - 1.0; chalk, powdered with 0.1 g of
silicate - 1.0; pH 7.2 [6]. For the cultivation of
bacteria mineral medium A of such
composition was also used (g/l): glucose -15.4;
MgSO4x7H2O - 0,7; Na2HPO4x2N2O - 2.0;
KNO3 - 1.0; FeCl3x6H2O - 0,05. The
composition varied depending on
environmental conditions and research
objective. Different N:C ratio in the
environment were created by increasing
glucose concentrations. For obtaining B.
mucilaginosus culture, which does not
synthesize exopolysaccharide (EPS), it was 3-5
times passaged in liquid medium with N:C ratio
= 1:3.
Growing B. mucilaginosus was conducted
in a periodical process which was carried out in
750 ml flasks with 100 ml of medium on a
shaker at 240 rev./min. The culture medium
temperature was 30°C, cultivation time – 24
hours. The volume of seed material made up
5% of the medium volume. To study the impact
of seed material origin on polysaccharide
synthesis, the concentration of cells and spores
in the medium was determined by Goriaiev
chamber, while the concentration of spores
after seed material addition to the medium was
6.5x107 units/ml, the concentration of
vegetative cells – 7.0-7.4x107 cells/ml.
Spore-based seed material was pre-heated at
the temperature of 80°C for 10 min. to kill
vegetative cells.
The content of cells that synthesize
exopolysaccharide was determined by
weighing technique. The cells were
precipitated at 40000 g for 40 min. Weighing
cups with biomass were dried to constant
weight at the temperature of 105°C. To take
into account the number of cells in the culture
liquid, the direct count method in Goriaiev
chamber and limiting dilutions method
followed by bacterial suspension seeding on
agar medium were used.
The amount of extracellular
polysaccharide was determined by weighing
technique following the three-fold ethanol
volume precipitation. For this, the culture
liquid (18-24 hrs.) was 2-3 times diluted with
physiological solution, centrifuged to
precipitate the cells at 40 000 g for 40 min. The
supernatant was poured off and polysaccharide
was precipitated with three volumes of ethanol
from it. Precipitated EPS was separated from
alcohol by centrifugation at 6000 rpm for 10
min. Ethanol precipitate was dissolved in
distilled water and dialyzed against drinking
water one day, against distilled – two days. The
polysaccharide was lyophilized to get dry.
Thus, it was found that the optimal
nitrogen form for C-3 strain of B.
mucilaginosus is nitrate form of nitrogen, with
which the largest amount of biomass is
accumulated and the process of sporulation
takes place, which is not provided by the
ammonium form of nitrogen.
The content of 2.5 g/l of potassium nitrate
in medium A provides maximum accumulation
of both biomass and extracellular
polysaccharide B. mucilaginosus.
The Intensity of nitrogen use by B.
mucilaginosus culture depends on the
physiological condition seed material cells: if
seed material is used in the form of spores,
maximum amount of biomass and extracellular
polysaccharide is produced.
As the number of B. mucilaginosus culture
re-seeding increased in a vegetative state the
efficiency of nitrogen source use and the ability
to synthesize exopolysaccharide significantly
decreases: after the second re-seeding – by
20.0%, after the fourth – by 56.5%, after the
sixth – by 127.8%.
|
| id | oai:ojs2.smic.in.ua:article-140 |
| institution | Agriciltural microbiology |
| keywords_txt_mv | keywords |
| language | English Ukrainian |
| last_indexed | 2026-07-23T01:01:53Z |
| publishDate | 2015 |
| publisher | Institute of Agrocultural Microbiology and Agro-industrial Manufacture of NAAS of Ukraine |
| record_format | ojs |
| resource_txt_mv | smicinua/27/aa3e4e232a65ba5b9a5051ff99712427.pdf |
| spelling | oai:ojs2.smic.in.ua:article-1402026-07-22T10:10:42Z EXOPOLYSACCHARIDE SYNTHESIS PRODUCTIVITY BY BACILLUS MUCILAGINOSUS DEPENDING ON THE NITROGEN SOURCE AND ORIGIN OF INOCULUM ПРОДУКТИВНІСТЬ СИНТЕЗУ ЕКЗОПОЛІСАХАРИДУ BACILLUS MUCILAGINOSUS ЗАЛЕЖНО ВІД ДЖЕРЕЛА АЗОТУ І ПОХОДЖЕННЯ ПОСІВНОГО МАТЕРІАЛУ Малиновська, І. М. Bacillus mucilaginosus, exopolysaccharide, nitrogen source, inoculum, spores Bacillus mucilaginosus, позаклітинний полісахарид, джерело азоту, посівний матеріал, спори It was established that Bacillus mucilaginosus C-3 requires introduction of nitrogen sourceinto the culture medium in form of nitrate as themost optimal for maximal accumulation of bacterial biomass and sporulation process. Introduction of the ammonium form was less efficient. It was shown that the intensity of nitrogensource use by B. mucilaginosus is highly depended on the physiological state of the inoculum cells. At sowing of spore inoculum theoptimal concentration of potassium nitrate was2.5 g/l resulting in 1010 cells/ml, while under theuse of vegetative cells the optimal concentrationof potassium nitrate was 1.0 g/l leading to theaccumulation of 109 cells/ml.Use of spore inoculum had ensured themaximum productivity of extracellular polysaccharide synthesis. With the higher number ofB. mucilaginosus subcultures in a vegetativestate the efficiency of the exopolysaccharidesynthesis has decreased: after the second passage — by 20.0 %, after the fourth — by 56.5 %,after the sixth — at 127.8 %. After eighth passage the culture loses its ability to synthesizepolysaccharide, especially on the medium withthe N : C ratio 1 : 3 and resume this ability onlyafter sporulation and the significant increase ofthe N : C ratio. Встановлено, що Bacillus mucilaginosus C-3 потребує внесення у середовище культивування джерела азоту в нітратній формі, за якої накопичується найбільша кількість біомаси і проходить процес споруляції, чого не забезпечує амонійна форма азоту. Інтенсивність використання азоту B. mucilaginosus залежить від фізіологічного стану клітин посівного матеріалу: при засіванні спорового посівного матеріалу оптимальна концентрація нітрату калію складає 2,5 г/л і накопичується 1010 кл./мл, при засіванні середовища вегетативними клітинами оптимальна концентрація нітрату калію складає 1,0 г/л і накопичується 109 кл./мл. Концентрація нітрату калію 2,5 г/л забезпечує максимальне накопичення як біомаси, так і позаклітинного полісахариду B. mucilaginosus. За використання спорового посівного матеріалу спостерігається найвища продуктивність синтезу позаклітинного полісахариду. Зі збільшенням кількості пересівів B. mucilaginosus у вегетативному стані здатність до синтезу екзополісахариду істотно зменшується: за другого пересіву — на 20,0 %, за четвертого — на 56,5 %, за шостого — на 127,8 %. Після 8–9 пересівів культура втрачає здатність синтезувати полісахарид, особливо на середовищі зі співвідношенням N : C = 1 : 3, і поновлює цю здатність лише після проходження процесу споруляції або значного підвищення співвідношення N : C. Institute of Agrocultural Microbiology and Agro-industrial Manufacture of NAAS of Ukraine 2015-08-06 Article Article Рецензована Стаття application/pdf application/pdf https://smic.in.ua/index.php/journal/article/view/140 10.35868/1997-3004.21.12-17 Agricultural microbiology; Vol. 21 (2015): Agriciltural microbiology; 12-17 Сільськогосподарська мікробіологія; Том 21 (2015): Сільськогосподарська мікробіологія; 12-17 1997-3004 10.35868/1997-3004.21 en uk https://smic.in.ua/index.php/journal/article/view/140/166 https://smic.in.ua/index.php/journal/article/view/140/167 Авторське право (c) 2015 I. M. Malinovska https://creativecommons.org/licenses/by/4.0 |
| spellingShingle | Bacillus mucilaginosus позаклітинний полісахарид джерело азоту посівний матеріал спори Малиновська, І. М. ПРОДУКТИВНІСТЬ СИНТЕЗУ ЕКЗОПОЛІСАХАРИДУ BACILLUS MUCILAGINOSUS ЗАЛЕЖНО ВІД ДЖЕРЕЛА АЗОТУ І ПОХОДЖЕННЯ ПОСІВНОГО МАТЕРІАЛУ |
| title | ПРОДУКТИВНІСТЬ СИНТЕЗУ ЕКЗОПОЛІСАХАРИДУ BACILLUS MUCILAGINOSUS ЗАЛЕЖНО ВІД ДЖЕРЕЛА АЗОТУ І ПОХОДЖЕННЯ ПОСІВНОГО МАТЕРІАЛУ |
| title_alt | EXOPOLYSACCHARIDE SYNTHESIS PRODUCTIVITY BY BACILLUS MUCILAGINOSUS DEPENDING ON THE NITROGEN SOURCE AND ORIGIN OF INOCULUM |
| title_full | ПРОДУКТИВНІСТЬ СИНТЕЗУ ЕКЗОПОЛІСАХАРИДУ BACILLUS MUCILAGINOSUS ЗАЛЕЖНО ВІД ДЖЕРЕЛА АЗОТУ І ПОХОДЖЕННЯ ПОСІВНОГО МАТЕРІАЛУ |
| title_fullStr | ПРОДУКТИВНІСТЬ СИНТЕЗУ ЕКЗОПОЛІСАХАРИДУ BACILLUS MUCILAGINOSUS ЗАЛЕЖНО ВІД ДЖЕРЕЛА АЗОТУ І ПОХОДЖЕННЯ ПОСІВНОГО МАТЕРІАЛУ |
| title_full_unstemmed | ПРОДУКТИВНІСТЬ СИНТЕЗУ ЕКЗОПОЛІСАХАРИДУ BACILLUS MUCILAGINOSUS ЗАЛЕЖНО ВІД ДЖЕРЕЛА АЗОТУ І ПОХОДЖЕННЯ ПОСІВНОГО МАТЕРІАЛУ |
| title_short | ПРОДУКТИВНІСТЬ СИНТЕЗУ ЕКЗОПОЛІСАХАРИДУ BACILLUS MUCILAGINOSUS ЗАЛЕЖНО ВІД ДЖЕРЕЛА АЗОТУ І ПОХОДЖЕННЯ ПОСІВНОГО МАТЕРІАЛУ |
| title_sort | продуктивність синтезу екзополісахариду bacillus mucilaginosus залежно від джерела азоту і походження посівного матеріалу |
| topic | Bacillus mucilaginosus позаклітинний полісахарид джерело азоту посівний матеріал спори |
| topic_facet | Bacillus mucilaginosus exopolysaccharide nitrogen source inoculum spores Bacillus mucilaginosus позаклітинний полісахарид джерело азоту посівний матеріал спори |
| url | https://smic.in.ua/index.php/journal/article/view/140 |
| work_keys_str_mv | AT malinovsʹkaím exopolysaccharidesynthesisproductivitybybacillusmucilaginosusdependingonthenitrogensourceandoriginofinoculum AT malinovsʹkaím produktivnístʹsintezuekzopolísaharidubacillusmucilaginosuszaležnovíddžerelaazotuípohodžennâposívnogomateríalu |