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Isolation and characterization of novel thermophilic lipase-secreting bacteria.

Rabbani M, Bagherinejad MR, Sadeghi HM, Shariat ZS, Etemadifar Z, Moazen F, Rahbari M, Mafakher L, Zaghian S - Braz. J. Microbiol. (2014)

Bottom Line: These strains were identified by PCR amplification of 16s rRNA genes using universal primers.Fermentation increased the activity up to 50%.The growth medium, designed for lipase production, increased the activity up to 4.55 folds.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmaceutical Biotechnology, School of Pharmacy and Pharmaceutical Sciences, Isfahan University of Medical Sciences, Isfahan, Iran.

ABSTRACT
The purpose of the present study was to screen and identify the lipase-producing microorganisms from various regions of Iran. Samples collected from hot spring, Persian Gulf, desert area and oil-contaminated soil, were analyzed for thermophilic extracellular-lipase producing organisms. Six strains with high activity on rhodamine B plates were selected for chemical identification and further study. Among these isolated bacteria, four strains show higher activity in pH-Stat method at 55 °C. These strains were identified by PCR amplification of 16s rRNA genes using universal primers. Fermentation increased the activity up to 50%. The growth medium, designed for lipase production, increased the activity up to 4.55 folds. The crude supernatant of ZR-5 after fermentation and separation the cells, was lyophilized and the activity was measured. Total activity of this strain was 12 kU/g that shows its potential for industrial uses. Further study is required for purification of enzyme and calculation its specific activity. Immobilization is another approach should be considered.

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Related in: MedlinePlus

Time course of lipase activity in shake flask method (37 °C, 220 rpm in LB with 2% v/v olive oil at pH 7.2). The lipase activity was measured using pH-Stat method (Titrando 902, Metrohm, Switzerland) and 12.5% v/v olive oil in gum Arabic (5% w/v) as substrate. The reaction was monitored for 10 min at 58 °C, pH 8 and 15 rpm. The points are means of three repeats.
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f3-bmj-44-4-1113: Time course of lipase activity in shake flask method (37 °C, 220 rpm in LB with 2% v/v olive oil at pH 7.2). The lipase activity was measured using pH-Stat method (Titrando 902, Metrohm, Switzerland) and 12.5% v/v olive oil in gum Arabic (5% w/v) as substrate. The reaction was monitored for 10 min at 58 °C, pH 8 and 15 rpm. The points are means of three repeats.

Mentions: Six strains of bacteria that showed positive results on Rhodamine test (ZR-1, ZR-5, PG-9, PG-31, PG-1 and WW) were further analyzed for lipase activity using pH-Stat method at 55 °C. Plate assay is normally used for primary screening of lipase activity (Samad et al., 1989). In this study Rhodamine B assay was used. Rhodamine B assay is a non-specific method for screening esterase activity and hence it cannot be used by itself to measure the lipase activity (Shu et al., 2010). Plate assay for primary screening of lipase activity are Thermophilic lipase activity of the selected strains grown on shake flask was measured by pH-Stat method using an auto-titrator at 55 °C. The activity measured after different incubation times (8–18 h with 2 h intervals) showed that ZR-1, ZR-5 and WW have the highest activity after 14 h of incubation, being respectively, 3.01, 2.33 and 1.87 U/mL (Figure 3). The optimum activity for PG-1, however, was 2.95 U/mL at 18 h after incubation (Figure 3). Other two strains, PG-9 and PG-31 did not show lipase activity using olive oil as substrate in pH-stat method at 55 °C.


Isolation and characterization of novel thermophilic lipase-secreting bacteria.

Rabbani M, Bagherinejad MR, Sadeghi HM, Shariat ZS, Etemadifar Z, Moazen F, Rahbari M, Mafakher L, Zaghian S - Braz. J. Microbiol. (2014)

Time course of lipase activity in shake flask method (37 °C, 220 rpm in LB with 2% v/v olive oil at pH 7.2). The lipase activity was measured using pH-Stat method (Titrando 902, Metrohm, Switzerland) and 12.5% v/v olive oil in gum Arabic (5% w/v) as substrate. The reaction was monitored for 10 min at 58 °C, pH 8 and 15 rpm. The points are means of three repeats.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3958176&req=5

f3-bmj-44-4-1113: Time course of lipase activity in shake flask method (37 °C, 220 rpm in LB with 2% v/v olive oil at pH 7.2). The lipase activity was measured using pH-Stat method (Titrando 902, Metrohm, Switzerland) and 12.5% v/v olive oil in gum Arabic (5% w/v) as substrate. The reaction was monitored for 10 min at 58 °C, pH 8 and 15 rpm. The points are means of three repeats.
Mentions: Six strains of bacteria that showed positive results on Rhodamine test (ZR-1, ZR-5, PG-9, PG-31, PG-1 and WW) were further analyzed for lipase activity using pH-Stat method at 55 °C. Plate assay is normally used for primary screening of lipase activity (Samad et al., 1989). In this study Rhodamine B assay was used. Rhodamine B assay is a non-specific method for screening esterase activity and hence it cannot be used by itself to measure the lipase activity (Shu et al., 2010). Plate assay for primary screening of lipase activity are Thermophilic lipase activity of the selected strains grown on shake flask was measured by pH-Stat method using an auto-titrator at 55 °C. The activity measured after different incubation times (8–18 h with 2 h intervals) showed that ZR-1, ZR-5 and WW have the highest activity after 14 h of incubation, being respectively, 3.01, 2.33 and 1.87 U/mL (Figure 3). The optimum activity for PG-1, however, was 2.95 U/mL at 18 h after incubation (Figure 3). Other two strains, PG-9 and PG-31 did not show lipase activity using olive oil as substrate in pH-stat method at 55 °C.

Bottom Line: These strains were identified by PCR amplification of 16s rRNA genes using universal primers.Fermentation increased the activity up to 50%.The growth medium, designed for lipase production, increased the activity up to 4.55 folds.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmaceutical Biotechnology, School of Pharmacy and Pharmaceutical Sciences, Isfahan University of Medical Sciences, Isfahan, Iran.

ABSTRACT
The purpose of the present study was to screen and identify the lipase-producing microorganisms from various regions of Iran. Samples collected from hot spring, Persian Gulf, desert area and oil-contaminated soil, were analyzed for thermophilic extracellular-lipase producing organisms. Six strains with high activity on rhodamine B plates were selected for chemical identification and further study. Among these isolated bacteria, four strains show higher activity in pH-Stat method at 55 °C. These strains were identified by PCR amplification of 16s rRNA genes using universal primers. Fermentation increased the activity up to 50%. The growth medium, designed for lipase production, increased the activity up to 4.55 folds. The crude supernatant of ZR-5 after fermentation and separation the cells, was lyophilized and the activity was measured. Total activity of this strain was 12 kU/g that shows its potential for industrial uses. Further study is required for purification of enzyme and calculation its specific activity. Immobilization is another approach should be considered.

Show MeSH
Related in: MedlinePlus