Production and Properties of Alkaline Dextranase from a Newly Isolated Brevibacterium

About 500 strains of dextranase producing microorganisms were examined in detail for pH- activity and enzyme stability. A gram positive bacterium identified as belonging to the genus Brevibacterium was found to produce alkaline dextranase. Maximal dextranase synthesis was obtained when grown aerobically at 26°C for 3 days in a medium containing 1 % dextran, 2% ethanol, 1 % polypeptone and 0.05 % yeast extract together with trace amounts of inorganic salts.

Brevibacterium dextranase had an optimum pH of 8.0 for activity at 37°C and an optimal temperature at 53°C at pH 7.5. The enzyme was quite stable over the range of pH 5.0 to 10.5 on 24 hr incubation at 37°C, especially on alkaline pH. The enzyme was also heat stable at 60°C for 10 min.     

Themoanaerobacterium calidifontis sp. nov., a novel anaerobic, thermophilic, ethanol-producing bacterium from hot springs in China

A novel thermophilic Gram staining positive strain Rx1 was isolated from hot springs in Baoshan of Yunnan Province, China. The strain was characterized as a hemicellulose-decomposing obligate anaerobe bacterium that is rod-shaped (diameter: 0.5–0.7 μm; length: 2.0–6.7 μm), spore-forming, and motile. Its growth temperature range is 38–68 °C (optimum 50–55 °C) and pH range is 4.5–8.0 (optimum 7.0). The maximum tolerance concentration of NaCl was 3 %. Rx1 converted thiosulfate to elemental sulfur and reduced sulfite to hydrogen sulfide. The bacterium grew by utilizing xylan and starch, as well as a wide range of monosaccharide and polysaccharides, including glucose and xylose. The main products of fermentation were ethanol, lactate, acetate, CO₂, and H₂. The maximum xylanase activity in the culture supernatant after 30 h of incubation at 55 °C was 16.2 U/ml. Rx1 DNA G + C content was 36 mol %. 16S rRNA gene sequence analysis indicated that strain Rx1 belonged to the genus Thermoanaerobacterium of the family ‘Thermoanaerobacteriaceae’ (Firmicutes), with Thermoanaerobacterium aciditolerans 761–119 (99.2 % 16S rRNA gene sequence similarity) being its closest relative. DNA–DNA hybridization between Rx1 and T. aciditolerans 761–119 showed 36 % relatedness. Based on its physiological and biochemical tests and DNA–DNA hybridization analyses, the isolate is considered to represent a novel species in the genus Thermoanaerobacterium, for which the name Thermoanaerobacterium calidifontis sp. nov. is proposed, with the type strain is Rx1 (=JCM 18270 = CCTCC M 2011109).     

Extracellular solvent stable cold-active lipase from psychrotrophic Bacillus sphaericus MTCC 7526: partial purification and characterization

Psychrotropic Bacillus sphaericus producing solvent stable cold-active lipase upon growth at low temperature was isolated from Gangotri glacier. Optimal parameters for lipase production were investigated and the strain was able to produce lipase even at 15 °C. An incubation period of 48 h and pH 8 was found to be conducive for cold-active lipase production. The addition of trybutyrin as substrate and lactose as additional carbon source increased lipase production. The enzyme was purified up to 17.74-fold by ammonium sulphate precipitation followed by DEAE cellulose column chromatography. The optimum temperature and pH for lipase activity were found to be 15 °C and 8.0, respectively. The lipase was found to be stable in the temperature range 20–30 °C and the pH range 6.0–9.0. The protein retained more than 83 % of its initial activity after exposure to organic solvents. The lipase exhibited significant stability in presence of acetone and DMSO retaining >90 % activity. The enzyme activity was inhibited by 10 mM CuSO₄ and EDTA but showed no loss in activity after incubation with other metals or inhibitors examined in this study.     

Paenibacillus brassicae sp. nov., isolated from cabbage rhizosphere in Beijing, China

A novel Gram-positive, rod-shaped, motile, spore-forming, nitrogen-fixing bacterium, designated strain 112^T, was isolated from cabbage rhizosphere in Beijing, China. The strain was found to grow at 10–40 °C and pH 4–11, with an optimum of 30 °C and pH 7.0, respectively. Phylogenetic analysis based on a fragment of the full-length 16S rRNA gene sequence revealed that strain 112^T is a member of the genus Paenibacillus. High levels of 16S rRNA gene similarities were found between strain 112^T, Paenibacillus sabinae DSM 17841^T (97.82 %) and Paenibacillus forsythiae DSM 17842^T (97.22 %). However, the DNA–DNA hybridization values between strain 112^T and the type strains of these two species were 10.36 and 6.28 %, respectively. The predominant menaquinone was found to be menaquinone 7 (MK-7). The major fatty acids were determined to be anteiso-C_15:0 and C_16:0. The major polar lipids were found to be diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol and unknown aminophospholipids. The cell wall peptidoglycan was found to contain meso-diaminopimelic acid. The DNA G+C content was determined to be 55.4 mol%. On the basis of its phenotypic characteristics, 16S rRNA gene sequence analysis and the value of DNA–DNA hybridization, strain 112^T is considered to represent a novel species of the genus Paenibacillus, for which the name Paenibacillus brassicae sp. nov. is proposed. The type strain is 112^T (= ACCC 01125^T = DSM 24983^T).     

Bacillus alkalicola sp. nov., An Alkaliphilic,Gram-Positive Bacterium Isolated from Zhabuye Lake in Tibet,China

A Gram-positive, alkaliphilic bacterium, designated strain Zby6^T, was isolated from Zhabuye Lake in Tibet, China. The strain was able to grow at pH 8.0–11.0 (optimum at pH 10.0), in 0–8 % (w/v) NaCl (optimum at 3 %, w/v) and at 10–45 °C (optimum at 37 °C). Cells of the isolate were facultatively anaerobic and spore-forming rods with polar flagellum. The predominant isoprenoid quinone was MK-7, and its cell wall peptidoglycan contained meso-diaminopimelic acid. The major cellular fatty acids were iso-C_15:0, C_16:0 and anteiso-C_15:0. The major polar lipids consisted of phosphatidylglycerol, diphosphatidylglycerol, and phosphatidylethanolamine. The genomic DNA G+C content of the isolate was 38.9 mol%. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain Zby6^T was a member of the genus Bacillus and most closely related to Bacillus cellulosilyticus DSM 2522^T (97.7 % similarity). The DNA–DNA relatedness value between strain Zby6^T and B. cellulosilyticus DSM 2522^T was 59.2 ± 1.8 %. Comparative analysis of genotypic and phenotypic features indicated that strain Zby6^T represents a novel species of the genus Bacillus, for which the name Bacillus alkalicola sp. nov. is proposed; the type strain is Zby6^T (=CGMCC 1.10368^T = JCM 17098^T = NBRC 107743^T).     

Thermus tengchongensis sp. nov., isolated from a geothermally heated soil sample in Tengchong, Yunnan, south-west China

A Gram-stain negative aerobic bacterium, designated YIM 77924^T, was isolated from a geothermally heated soil sample collected at Rehai National Park, Tengchong, Yunnan province, south-west China. Growth was found to occur from 55 to 75 °C (optimum 65 °C), pH 6.0–8.0 (optimum pH 7.0) and 0–1 % NaCl (w/v). Cells were observed to be rod-shaped and the colonies convex, circular, smooth, yellow and non-transparent. Phylogenetic analysis based on the 16S rRNA gene sequence indicated that strain YIM 77924^T belongs to the genus Thermus. The 16S rRNA gene sequence similarity values between strain YIM 77924^T and other species of the genus Thermus were all below 97 %. The polar lipids of strain YIM 77924^T were determined to be aminophospholipid, phospholipid and glycolipid. The predominant respiratory quinone was determined to be MK-8 and the G+C content was 66.64 mol%. The major fatty acids identified were iso-C_16:0, iso-C_15:0, iso-C_17:0 and C_16:0. On the basis of the morphological and chemotaxonomic characteristics as well as genotypic data, strain YIM 77924^T is proposed to represent a novel species, Thermus tengchongensis sp. nov., in the genus Thermus. The type strain is YIM 77924^T (=KCTC 32025^T = CCTCC AB2012063^T).     

Studies on dextranase from Penicillium aculeatum

A strain of Penicillium aculeatum has been found to synthesize large quantities of dextranase (1,6-α-d-glucan 6-glucanohydrolase, EC 3.2.1.11) in culture filtrate. Some of the conditions governing the enzyme production have been standardized. The enzyme in crude state was found to be highly stable, its activity being maximum at 50 to 60°C and at pH 5 to 6. About 90% of the substrate dextran was converted to isomaltose in a 4 h period at 40°C. The enzyme when purified by salt and solvent fractionation gave 1500 units per mg protein and retained its activity over a long period when stored at 4°C.     

Optimization, purification and characterization of novel thermostable, haloalkaline, solvent stable protease from Bacillus halodurans CAS6 using marine shellfish wastes: a potential additive for detergent and antioxidant synthesis

A protease producing marine bacterium, Bacillus halodurans CAS6 isolated from marine sediments, was found to produce higher enzyme by utilizing shrimp shell powder. Optimum culture conditions for protease production were 50 °C, pH 9.0, 30 % NaCl and 1 % shrimp shell powder (SSP) and the protease purified with a specific activity of 509.84 U/mg. The enzyme retained 100 % of its original activity even at 70 °C, pH 10.0 and 30 % NaCl for 1 h. The purified protease exhibited higher stability when treated with ionic, non-ionic (72–94 %) and commercial detergents (76–88 %), and organic solvents (88–126 %). Significant blood stain removal activity was found with the enzyme in washing experiments. The culture supernatant supplemented with 1 % SSP showed 93.67 ± 2.52 % scavenging activity and FT-IR analysis of the reaction mixture confirmed the presence of antioxidants such as cyclohexane and cyclic depsipeptide with aliphatic amino groups. These remarkable qualities found with this enzyme produced by Bacillus halodurans CAS6 could make this as an ideal candidate to develop the industrial process for bioconversion of marine wastes and antioxidant synthesis.     

Lysobacter thermophilus sp. nov., isolated from a geothermal soil sample in Tengchong, south-west China

A Gram-negative and aerobic bacterium, designated YIM 77875^T, was isolated from a geothermal soil sample collected at Rehai National Park, Tengchong, Yunnan Province, south-west China. Bacterial growth occurred from 37 to 65 °C (optimum 50 °C), pH 6.0–8.0 (optimum pH 7.0) and 0–1 % NaCl (w/v). Cells were rod-shaped and colonies were convex, circular, smooth, yellow and non-transparent. Phylogenetic analysis based on the 16S rRNA gene sequence indicated that strain YIM 77875^T belongs to the genus Lysobacter. The 16S rRNA gene sequence similarity values between strain YIM 77875^T and other species of the genus Lysobacter were all below 94.7 %. The polar lipids of strain YIM 77875^T were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine and five unknown phospholipids. The predominant respiratory quinone was Q-8 and the G+C content was 68.8 mol%. Major fatty acids were iso-C_16:0, iso-C_15:0 and iso-C_11:0. On the basis of the morphological and chemotaxonomic characteristics, as well as genotypic data, strain YIM 77875^T represents a novel species, Lysobacter thermophilus sp. nov., in the genus Lysobacter. The type strain is YIM 77875^T (CCTCC AB 2012064^T = KCTC 32020^T).     

Isolation and characterization of marine diesel oil-degrading Acinetobacter sp. strain Y2

The marine diesel oil-degrading bacterium Acinetobacter sp. strain Y2 was isolated from oil-polluted seawater sampled from Dinghai port, Zhoushan City, Zhejiang Province, China. The isolated bacterium was identified as Acinetobacter sp. based on its 16S rDNA gene sequence as well as various morphological and physiological characteristics. The degradation characteristics of strain Y2 were studied and its parameters for oil degradation optimized. These optimal conditions were determined to be an initial pH of 7.5, an incubation temperature of 30 °C, an initial diesel oil concentration of 2 % (v/v), and an initial inoculating bacteria concentration of 3?×?10⁷ cells/mL. The results from the gas chromatography–mass spectrometry analysis showed that strain Y2 could almost completely degrade all components of diesel oil, with a degradation ratio of up to 80 % after 10 days of incubation at the optimal conditions.     

Characterization of a marine-derived dextranase and its application to the prevention of dental caries

The dextranase added in current commercial dextranase-containing mouthwashes is largely from fungi. However, fungal dextranase has shown much higher optimum temperature than bacterial dextranase and relatively low activity when used in human oral cavities. Bacterial dextranase has been considered to be more effective and suitable for dental caries prevention. In this study, a dextranase (Dex410) from marine Arthrobacter sp. was purified and characterized. Dex410 is a 64-kDa endoglycosidase. The specific activity of Dex410 was 11.9 U/mg at optimum pH 5.5 and 45 °C. The main end-product of Dex410 was isomaltotriose, isomaltoteraose, and isomaltopentaose by hydrolyzing dextran T2000. In vitro studies showed that Dex410 effectively inhibited the Streptococcus mutans biofilm growth in coverage, biomass, and water-soluble glucan (WSG) by more than 80, 90, and 95 %, respectively. The animal experiment revealed that for short-term use (1.5 months), both Dex410 and the commercial mouthwash Biotene (Laclede Professional Products, Gardena, CA, USA) had a significant inhibitory effect on caries (p = 0.0008 and 0.0001, respectively), while for long-term use (3 months), only Dex410 showed significant inhibitory effect on dental caries (p = 0.005). The dextranase Dex410 from a marine-derived Arthrobacter sp. strain possessed the enzyme properties suitable to human oral environment and applicable to oral hygiene products.     

Roseovarius azorensis sp. nov., isolated from seawater at Espalamaca,Azores

A Gram-negative, motile, non-spore forming, rod shaped aerobic bacterium, designated strain SSW084^T, was isolated from a surface seawater sample collected at Espalamaca (38°33′N; 28°39′W), Azores. Growth was found to occur from 15 to 40 °C (optimum 30 °C), at pH 7.0–9.0 (optimum pH 7.0) and with 25–100 % seawater or 0.5–7.0 % NaCl in the presence of Mg²⁺ and Ca²⁺; no growth was found with NaCl alone. Colonies on seawater nutrient agar were observed to be punctiform, white, convex, circular, smooth, and translucent. Strain SSW084^T did not grow on Zobell marine agar and tryptic soy agar even when seawater supplemented. The major respiratory quinone was found to be Q-10 and the G + C content was determined to be 61.9 mol%. Phylogenetic analysis based on the 16S rRNA gene sequence indicated that strain SSW084^T belongs to the genus Roseovarius and that its closest neighbours are Roseovarius tolerans EL-172^T, Roseovarius mucosus DFL-24^T and Roseovarius lutimaris 112^T with 95.7, 95.4 and 95.3 % sequence similarity respectively. The remaining species of Roseovarius showed <95 % similarity. The polar lipids of strain SSW084^T were determined to be phosphatidylethanolamine, diphosphatidylglycerol, phosphatidylglycerol, phosphatidylcholine, an unidentified lipid and one unidentified aminolipid. The major fatty acids identified were identified as C_18:1 ω7c (52.5 %) and C_16:0 (13.8 %). On the basis of phenotypic, molecular and chemotaxonomic characteristics, strain SSW084^T is considered to represent a novel species of the genus Roseovarius, for which Roseovarius azorensis sp. nov is proposed. The type strain is SSW084^T (=KCTC 32421^T = MTCC 11812^T).     

一株产右旋糖苷酶海洋细菌的筛选鉴定

【目的】筛选鉴定产右旋糖苷酶的海洋细菌,并对其所产右旋糖苷酶的酶学性质及在变异链球菌牙菌斑生物膜中的应用进行初步研究。【方法】利用平板透明圈法从海洋环境中筛选产右旋糖苷酶的细菌,根据菌株形态特征、生理特征及16S rDNA序列确定其分类学地位,采用体外生物膜模型研究该酶对变异链球菌牙菌斑生物膜形成的抑制作用。【结果】从海泥中筛选出一株产右旋糖苷酶的细菌KQ11,初步鉴定为节杆菌(Arthrobacter sp.)。该菌株的最适生长温度为30°C,最适生长pH 7.5,最适生长NaCl浓度为0.4%。右旋糖苷酶的最适作用温度为45°C,最适作用pH为5.5。该酶能有效地抑制变异链球菌牙菌斑生物膜的形成。【结论】菌株KQ11右旋糖苷酶能够抑制变异链球菌牙菌斑生物膜的形成,可望用于漱口液等口腔护理产品中。     

Purification,characterization, and biocatalytic potential of a novel dextranase from <Emphasis Type="Italic">Chaetomium globosum</Emphasis>

Objective

We aimed to identify new high-yield dextranase strains and study the catalytic potential of dextranase from the strain in industrial applications.

Results

Dextranase-producing strains were screened from soil samples, and a potential strain was identified as Chaetomium globosum according to its phenotype, biochemical characteristics, and rDNA analysis. Crude dextranase was purified to reach 10.97-fold specific activity and 18.7% recovery. The molecular weight of the enzyme was 53 kDa with an optimum temperature and pH of 60 °C and 5.5, respectively. Enzyme activity was stable at pH 4.0–7.0 and displayed sufficient thermal stability at temperatures?<?50 °C. Mn²⁺ (10 mM) enhanced dextranase activity by 134.44%. The enzyme was identified as an endodextranase. It displayed very high hydrolytic affinity toward high-molecular weight dextran T2000, reaching 97.9% hydrolysis within 15 min at 2 U/mL.

Conclusion

Collectively, these results suggest that Chaetomium globosum shows higher production and specificity of dextranase than that from other reported strains. These findings may offer new insights into the potential of dextranase in the sugar, medical, and food industries.

Graphical abstract

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Sporosarcina siberiensis sp. nov., isolated from the East Siberian Sea

A Gram-stain positive, non-motile, rod-shaped bacterium, designated strain 1111S-42^T, was isolated from the East Siberian Sea. The organism was found to grow at 4–30 °C, pH 7.0–8.5 and in 0–8 % (w/v) NaCl, with optimal growth occurring at 28 °C, pH 7.5 and in 1 % NaCl. Based on 16S rRNA gene sequence similarity studies, strain 1111S-42^T was found to belong to the genus Sporosarcina and to be most closely related to Sporosarcina contaminans CCUG53915^T (97.3 %) and Sporosarcina soli I80^T (97.2 %). The main polar lipids were found to include diphosphatidylglycerol, phosphatidylglycerol and phosphatidylethanolamine. The predominant menaquinone was identified as MK-7. The major cellular fatty acids were identified as anteiso-C_15:0 (34.4 %), iso-C_15:0 (29.8 %) and anteiso-C_17:0 (22.4 %). The DNA G+C content of strain 1111S-42^T was determined to be 39 mol %. The values of DNA–DNA relatedness between the strain 1111S-42^T and related type strains of the genus Sporosarcina were less than 30 %. Based on the phylogenetic analysis, along with extensive physiological and chemotaxonomic testing, we conclude that the bacterium represents a novel species of the genus Sporosarcina, for which the name Sporosarcina siberiensis sp. nov. is proposed. The type strain is strain 1111S-42^T (=CGMCC 1.12516^T = LMG 27494^T).     

Purification, characterization and cloning of a thermotolerant isoamylase produced from Bacillus sp. CICIM 304

A novel thermostable isoamylase, IAM, was purified to homogeneity from the newly isolated thermophilic bacterium Bacillus sp. CICIM 304. The purified monomeric protein with an estimated molecular mass of 100 kDa displayed its optimal temperature and pH at 70 °C and 6.0, respectively, with excellent thermostability between 30 and 70 °C and pH values from 5.5 to 9.0. Under the conditions of temperature 50 °C and pH 6.0, the K _m and V _max on glycogen were 0.403 ± 0.018 mg/mg and 0.018 ± 0.001 mg/(min mg), respectively. Gene encoding IAM, BsIam was identified from genomic DNA sequence with inverse PCRs. The open reading frame of the BsIam gene was 2,655 base pairs long and encoded a polypeptide of 885 amino acids with a calculated molecular mass of 101,155 Da. The deduced amino acid sequence of IAM shared less than 40 % homology with that of microbial isoamylase ever reported, which indicated it was a novel isoamylase. This enzyme showed its obvious superiority in the industrial starch conversion process.     

Biodegradation and metabolic pathway of sulfamethoxazole by Pseudomonas psychrophila HA-4, a newly isolated cold-adapted sulfamethoxazole-degrading bacterium

Sulfamethoxazole is a common antibiotic that is frequently detected in wastewater and surface water. This study investigated the biodegradation and metabolic pathway of sulfamethoxazole by Pseudomonas psychrophila HA-4, a cold-adapted bacterium. Strain HA-4, which uses sulfamethoxazole as its sole source of carbon and energy, was isolated at a low temperature (10 °C) and identified as P. psychrophila by physico-biochemical characterization and 16S rRNA gene sequence analysis. Strain HA-4 removed sulfamethoxazole at temperatures ranging from 5.0 °C to 30 °C, with the maximal removal rate at 10 °C. The maximal removal rate of sulfamethoxazole by strain HA-4 was 34.30 % after 192 h at 10 °C. The highest percentage of unsaturated fatty acid was determined to be 23.03 % at 10 °C, which adheres to the characteristic for cold-adapted psychrophiles and psychrotrophs. At low concentrations of sulfamethoxazole, the growth kinetics correlated well with the Haldane model. The single-substrate parameter values of sulfamethoxazole on cell growth were determined to be μ _max?=?0.01 h^?1, K _s?=?20.91 mg/l and K _i?=?170.60 mg/l. Additionally, the major intermediates from sulfamethoxazole biodegradation by strain HA-4, including aniline, 3-amino-5-methylisoxazole, 4-aminothiophenol and sulfanilamide, were identified by GC-MS and high-resolution mass spectrometry (HR-MS) analysis. The results demonstrate that strain HA-4 has the potential to degrade sulfamethoxazole at low temperatures.     

Flavobacterium qiangtangensis sp. nov., Isolated from Qiangtang Basin in Qinghai-Tibetan Plateau,China

A flexirubin-type yellow-pigmented, non-gliding, non-flagellated, gram-negative bacterium strain, designated F3^T, was isolated from a drilling core sample of the Qiangtang basin, Qinghai-Tibetan plateau, China. Phylogenetic analysis based on 16S rRNA gene sequences showed that the strain F3^T belongs to the genus Flavobacterium, with the highest 16S rRNA gene sequence similarity to the Flavobacterium noncentrifugens CGMCC 1.10076^T (94.92 %). Strain F3^T grew optimally at temperature about 20 °C, at pH about 7.0–8.0, at NaCl concentration 0 % (w/v). The DNA G+C content of the isolate was 35.5 mol%. The major polar lipid was phosphatidylethanolamine, predominant cellular fatty acids of the strain was iso-C_15:0 (22.02 %), while the major menaquinone was menaquinone 6. Due to the phenotypic and genetic distinctiveness and several other characteristic studied in this article, we consider F3^T as a novel species of the genus Flavobacterium, and propose to name it Flavobacterium qiangtangensis sp. nov. The type strain is F3^T (=CGMCC 1.12706^T = JCM 19739^T).     

Isolation and lipid degradation profile of Raoultella planticola strain 232-2 capable of efficiently catabolizing edible oils under acidic conditions

The lipids (fats and oils) degradation capabilities of soil microorganisms were investigated for possible application in treatment of lipids-contaminated wastewater. We isolated a strain of the bacterium Raoultella planticola strain 232-2 that is capable of efficiently catabolizing lipids under acidic conditions such as in grease traps in restaurants and food processing plants. The strain 232-2 efficiently catabolized a mixture (mixed lipids) of commercial vegetable oil, lard, and beef tallow (1:1:1, w/w/w) at 20–35 °C, pH 3–9, and 1,000–5,000 ppm lipid content. Highly effective degradation rate was observed at 35 °C and pH 4.0, and the 24-h degradation rate was 62.5?±?10.5 % for 3,000 ppm mixed lipids. The 24-h degradation rate for 3,000 ppm commercial vegetable oil, lard, beef tallow, mixed lipids, and oleic acid was 71.8 %, 58.7 %, 56.1 %, 55.3?±?8.5 %, and 91.9 % at pH 4 and 30 °C, respectively. R. planticola NBRC14939 (type strain) was also able to efficiently catabolize the lipids after repeated subculturing. The composition of the culture medium strongly influenced the degradation efficiency, with yeast extract supporting more complete dissimilation than BactoPeptone or beef extract. The acid tolerance of strain 232-2 is proposed to result from neutralization of the culture medium by urease-mediated decomposition of urea to NH₃. The rate of lipids degradation increased with the rates of neutralization and cell growth. Efficient lipids degradation using strain 232-2 has been achieved in the batch treatment of a restaurant wastewater.