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Aquatic model for engine oil degradation by rhamnolipid producing Nocardiopsis VITSISB

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ABSTRACT

The present study was focused on isolation, screening, characterization and application of biosurfactant producing marine actinobacteria. Twenty actinobacteria were isolated from marine water sample and were primarily screened for biosurfactant production using hemolytic activity method. Among the 20 isolates, six showed positive result for hemolytic activity and those were taken for further secondary screening tests such as oil collapse method, oil spreading method and emulsification method. From the results of secondary screening analysis, two isolates (SIS-3 and SIS-20) were selected and further used to carry out biosurfactant characterization test such as pH, density, surface tension and viscosity determination. Comparing biosurfactant characterization results, SIS-3 was chosen for further analysis and application. FT-IR and GC–MS were carried out for analysis of biosurfactant from isolate SIS-3 and the compound detected was rhamnolipid. The isolate (SIS-3) was identified as Nocardiopsis using 16S rRNA gene sequencing and named as ‘Nocardiopsis VITSISB’ (KC958579) which was further applied for immobilizing whole cells for engine oil degradation by constructing an aquatic model and using natural products such as soybean meal, sugarcane juice as nutrient source. The oil was efficiently degraded by rhamnolipid producing Nocardiopsis VITSISB (KC958579) within 25 days which indicated that the strain can act as a natural candidate for the bioremediation of oil spill in ocean.

No MeSH data available.


Emulsification index (E24) of the isolates in two media. Data are given as mean ± SD (n = 3). Results were considered significant at p < 0.05
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Fig1: Emulsification index (E24) of the isolates in two media. Data are given as mean ± SD (n = 3). Results were considered significant at p < 0.05

Mentions: Total 20 marine actinobacteria were isolated from marine water sample. Out of 20 isolates, six isolates showed positive result in hemolytic activity on blood agar plates. The results are presented in Table 1. Two media (SS, SS + OIL) were used for the production of biosurfactant by marine actinobacteria. The emulsification index for the isolates in two media is mentioned in Fig. 1. The result of secondary screening of biosurfactant by six isolates is provided in Table 2. Biosurfactant characterization test were conducted and the results are summarized in Table 3. The surface tension was observed as 62.808 dyne/cm for SIS-3, 62.808 dyne/cm for SIS-20, and 68.6 dyne/cm for water. With reference to biosurfactant viscosity measurement result, SIS-3 showed 10.7 × 10−3 poise, SIS-20 showed 12.6 × 10−3 poise and sodium lauryl sulphate showed 10.1 × 10−3 poise, so lower the viscosity more would be the positive effect on emulsion stability of biosurfactant. Density measurement result showed SIS-3 as 1.16 g/cm3, SIS-20 as 1.18 g/cm3 and water as 68.6 g/cm3. The orcinol assay showed positive result by formation of blue-green color. The constancy of biosurfactant produced by SIS-3 was examined over an extensive variation of temperatures.The surface tension reduction (dyne/cm) and percentage of emulsification activities were highest at the temperature of 50 °C as shown in Fig. 2. The biosurfactant showed relatively high efficiency at pH 12 where the emulsification activity showed almost 66 % emulsion and the surface tension reduced to 56.8 dyne/cm as shown in Fig. 3. The effect of sodium chloride addition on biosurfactant produced from SIS-3 was studied as shown in Fig. 4. Optimum constancy of biosurfactant was observed at 10 % NaCl concentration. At higher concentration of sodium chloride, the biosurfactant showed 76 % of the emulsification activity and more reduction in surface tension than in lower concentration of NaCl. Molecular compositions of biosurfactant produced by SIS-3 were evaluated by FT-IR as shown in Fig. 5. The broad band observed in biosurfactant was at 3,444 cm−1 which corresponded to the hydroxy group, H-bonded OH stretch in glycolipid. The peak at 1,689 cm−1 corresponded to C=O stretching of carbonyl groups which is characteristic for ester compounds. The band 1,396 cm−1 confirmed the presence of alkyl groups. The C–O–C stretching in the rhamnose was found at 1,083 cm−1 and the stretching of aromatic C–H in-plane bend occurred at 991 cm−1. The GC–MS analysis of chloroform:methanol extracts of SIS-3 is shown in Fig. 6. The results showed that the major chemical constituent of chloroform:methanol extracts of SIS-3 was phenol, 3,5-bis(1,1-dimethylethyl)- by comparison of mass spectral data and retention times. The other major constituents present in the extract were benzene, 1,3-bis(1,1-dimethylethyl),dibutyl phthalate,2-tert-butyl-4,6-bis(3,5-di-tert-butyl-4 hydroxybenzyl) phenol and dodecane, 1-fluoro. The major chemical constituent phenol,3,5-bis(1,1-dimethylethyl) was responsible for surfactant activity. The molecular weight of phenol, 3,5-bis(1,1-dimethylethyl) (C14H22O) was 206. The taxonomic position of Nocardiopsis VITSISB (KC958579) was determined based on 16S rRNA partial gene sequencing. A phylogenetic tree was constructed based on the neighbor-joining method using Treeview software as shown in Fig. 7. The oil layer in the aquatic model was reduced after duration of 25 days as shown in Fig. 8. HPLC Chromatogram of degraded oil showed more number of peaks as compared to control engine oil as shown in Fig. 9. Table 1


Aquatic model for engine oil degradation by rhamnolipid producing Nocardiopsis VITSISB
Emulsification index (E24) of the isolates in two media. Data are given as mean ± SD (n = 3). Results were considered significant at p < 0.05
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

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Fig1: Emulsification index (E24) of the isolates in two media. Data are given as mean ± SD (n = 3). Results were considered significant at p < 0.05
Mentions: Total 20 marine actinobacteria were isolated from marine water sample. Out of 20 isolates, six isolates showed positive result in hemolytic activity on blood agar plates. The results are presented in Table 1. Two media (SS, SS + OIL) were used for the production of biosurfactant by marine actinobacteria. The emulsification index for the isolates in two media is mentioned in Fig. 1. The result of secondary screening of biosurfactant by six isolates is provided in Table 2. Biosurfactant characterization test were conducted and the results are summarized in Table 3. The surface tension was observed as 62.808 dyne/cm for SIS-3, 62.808 dyne/cm for SIS-20, and 68.6 dyne/cm for water. With reference to biosurfactant viscosity measurement result, SIS-3 showed 10.7 × 10−3 poise, SIS-20 showed 12.6 × 10−3 poise and sodium lauryl sulphate showed 10.1 × 10−3 poise, so lower the viscosity more would be the positive effect on emulsion stability of biosurfactant. Density measurement result showed SIS-3 as 1.16 g/cm3, SIS-20 as 1.18 g/cm3 and water as 68.6 g/cm3. The orcinol assay showed positive result by formation of blue-green color. The constancy of biosurfactant produced by SIS-3 was examined over an extensive variation of temperatures.The surface tension reduction (dyne/cm) and percentage of emulsification activities were highest at the temperature of 50 °C as shown in Fig. 2. The biosurfactant showed relatively high efficiency at pH 12 where the emulsification activity showed almost 66 % emulsion and the surface tension reduced to 56.8 dyne/cm as shown in Fig. 3. The effect of sodium chloride addition on biosurfactant produced from SIS-3 was studied as shown in Fig. 4. Optimum constancy of biosurfactant was observed at 10 % NaCl concentration. At higher concentration of sodium chloride, the biosurfactant showed 76 % of the emulsification activity and more reduction in surface tension than in lower concentration of NaCl. Molecular compositions of biosurfactant produced by SIS-3 were evaluated by FT-IR as shown in Fig. 5. The broad band observed in biosurfactant was at 3,444 cm−1 which corresponded to the hydroxy group, H-bonded OH stretch in glycolipid. The peak at 1,689 cm−1 corresponded to C=O stretching of carbonyl groups which is characteristic for ester compounds. The band 1,396 cm−1 confirmed the presence of alkyl groups. The C–O–C stretching in the rhamnose was found at 1,083 cm−1 and the stretching of aromatic C–H in-plane bend occurred at 991 cm−1. The GC–MS analysis of chloroform:methanol extracts of SIS-3 is shown in Fig. 6. The results showed that the major chemical constituent of chloroform:methanol extracts of SIS-3 was phenol, 3,5-bis(1,1-dimethylethyl)- by comparison of mass spectral data and retention times. The other major constituents present in the extract were benzene, 1,3-bis(1,1-dimethylethyl),dibutyl phthalate,2-tert-butyl-4,6-bis(3,5-di-tert-butyl-4 hydroxybenzyl) phenol and dodecane, 1-fluoro. The major chemical constituent phenol,3,5-bis(1,1-dimethylethyl) was responsible for surfactant activity. The molecular weight of phenol, 3,5-bis(1,1-dimethylethyl) (C14H22O) was 206. The taxonomic position of Nocardiopsis VITSISB (KC958579) was determined based on 16S rRNA partial gene sequencing. A phylogenetic tree was constructed based on the neighbor-joining method using Treeview software as shown in Fig. 7. The oil layer in the aquatic model was reduced after duration of 25 days as shown in Fig. 8. HPLC Chromatogram of degraded oil showed more number of peaks as compared to control engine oil as shown in Fig. 9. Table 1

View Article: PubMed Central - PubMed

ABSTRACT

The present study was focused on isolation, screening, characterization and application of biosurfactant producing marine actinobacteria. Twenty actinobacteria were isolated from marine water sample and were primarily screened for biosurfactant production using hemolytic activity method. Among the 20 isolates, six showed positive result for hemolytic activity and those were taken for further secondary screening tests such as oil collapse method, oil spreading method and emulsification method. From the results of secondary screening analysis, two isolates (SIS-3 and SIS-20) were selected and further used to carry out biosurfactant characterization test such as pH, density, surface tension and viscosity determination. Comparing biosurfactant characterization results, SIS-3 was chosen for further analysis and application. FT-IR and GC&ndash;MS were carried out for analysis of biosurfactant from isolate SIS-3 and the compound detected was rhamnolipid. The isolate (SIS-3) was identified as Nocardiopsis using 16S rRNA gene sequencing and named as &lsquo;Nocardiopsis VITSISB&rsquo; (KC958579) which was further applied for immobilizing whole cells for engine oil degradation by constructing an aquatic model and using natural products such as soybean meal, sugarcane juice as nutrient source. The oil was efficiently degraded by rhamnolipid producing Nocardiopsis VITSISB (KC958579) within 25&nbsp;days which indicated that the strain can act as a natural candidate for the bioremediation of oil spill in ocean.

No MeSH data available.