Biodegradation of 4-bromophenol by Arthrobacter chlorophenolicus A6 in batch shake flasks and in a continuously operated packed bed reactor

The present study investigated growth and biodegradation of 4-bromophenol (4-BP) by Arthrobacter chlorophenolicus A6 in batch shake flasks as well as in a continuously operated packed bed reactor (PBR). Batch growth kinetics of A. chlorophenolicus A6 in presence of 4-BP followed substrate inhibition kinetics with the estimated biokinetic parameters value of μ _max = 0.246 h^?1, K _i = 111 mg L^?1, K _s  = 30.77 mg L^?1 and K = 100 mg L^?1. In addition, variations in the observed and theoretical biomass yield coefficient and maintenance energy of the culture were investigated at different initial 4-BP concentration. Results indicates that the toxicity tolerance and the biomass yield of A. chlorophenolicus A6 towards 4-BP was found to be poor as the organism utilized the substrate mainly for its metabolic maintenance energy. Further, 4-BP biodegradation performance by the microorganism was evaluated in a continuously operated PBR by varying the influent concentration and hydraulic retention time in the ranges 400–1,200 mg L^?1 and 24–7.5 h, respectively. Complete removal of 4-BP was achieved in the PBR up to a loading rate of 2,276 mg L^?1 day^?1.     

The removal of ρ-chlorophenol in aqueous cultures with free and alginate-immobilized cells of the microalga Tetraselmis suecica

The present study aimed at evaluating the ability of some isolated cyanobacterial and microalgal strains for the removal of ρ-chlorophenol (ρ-CP), an environmentally harmful contaminant. To identify the most efficient species, a screening program was carried out using 15 microalgal and cyanobacterial strains. Among them, Tetraselmis suecica was able to remove 67 % of the ρ-chlorophenol at an initial concentration of 20 mg L^?1 from the medium within a 10-day period. The efficacy of the process was dependent on the ρ-chlorophenol concentration. At concentrations above 60 mg L^?1 of the pollutant, no removal was observed due to the inhibitory effect of ρ-chlorophenol on the T. suecica cells. The effect of cell immobilization in alginate beads on T. suecica removal capacity was also examined. Using this technique, the removal efficacy for the initial ρ-CP concentration of 20 mg L^?1 increased up to 94 %.     

Kinetics and pathway of biodegradation of dibutyl phthalate by Pleurotus ostreatus

Dibutyl phthalate (DBP) is a plasticizer, whose presence in the environment as a pollutant has attained a great deal of attention due to its reported association with endocrine system disturbances on animals. Growth parameters, glucose uptake, percentage of removal efficiency (%E) of DBP, biodegradation constant of DBP (k) and half-life of DBP biodegradation (t_1/2) were evaluated for Pleurotus ostreatus grown on media containing glucose and different concentrations of DBP (0, 500 and 1000 mg l^?1). P. ostreatus degraded 99.6 % and 94 % of 500 and 1000 mg of DBP l^?1 after 312 h and 504 h, respectively. The k was 0.0155 h^?1 and 0.0043 h^?1 for 500 and 1000 mg of DBP l^?1, respectively. t_1/2 was 44.7 h and 161 h for 500 and 1000 mg of DBP l^?1, respectively. Intermediate compounds of biodegraded DBP were identified by GC-MS and a DBP biodegradation pathway was proposed using quantum chemical calculation. DBP might be metabolized to benzene and acetyl acetate, the first would be oxidated to muconic acid and the latter would enter into the Krebs cycle. P. ostreatus has the ability to degrade DBP and utilizes it as source of carbon and energy.     

Nickel,manganese and copper removal by a mixed consortium of sulfate reducing bacteria at a high COD/sulfate ratio

The use of sulfate-reducing bacteria (SRB) in passive treatments of acidic effluents containing heavy metals has become an attractive alternative biotechnology. Treatment efficiency may be linked with the effluent conditions (pH and metal concentration) and also to the amount and nature of the organic substrate. Variations on organic substrate and sulfate ratios clearly interfere with the biological removal of this ion by mixed cultures of SRB. This study aimed to cultivate a mixed culture of SRB using different lactate concentrations at pH 7.0 in the presence of Ni, Mn and Cu. The highest sulfate removal efficiency obtained was 98 %, at a COD/sulfate ratio of 2.0. The organic acid analyses indicated an acetate accumulation as a consequence of lactate degradation. Different concentrations of metals were added to the system at neutral pH conditions. Cell proliferation and sulfate consumption in the presence of nickel (4, 20 and 50 mg l^?1), manganese (1.5, 10 and 25 mg l^?1) and copper (1.5, 10 and 25 mg l^?1) were measured. The presence of metals interfered in the sulfate biological removal however the concentration of sulfide produced was high enough to remove over 90 % of the metals in the environment. The molecular characterization of the bacterial consortium based on dsrB gene sequencing indicated the presence of Desulfovibrio desulfuricans, Desulfomonas pigra and Desulfobulbus sp. The results here presented indicate that this SRB culture may be employed for mine effluent bioremediation due to its potential for removing sulfate and metals, simultaneously.     

Biodegradation of 4-chlorophenol by <Emphasis Type="Italic">Arthrobacter chlorophenolicus A6</Emphasis>: effect of culture conditions and degradation kinetics

Among known microbial species, Arthrobacter chlorophenolicus A6 has shown very good potential to treat phenolic wastewaters. In this study, the levels of various culture conditions, namely initial pH, agitation (rpm), temperature (°C), and inoculum age (h) were optimized to enhance 4-chlorophenol (4-CP) biodegradation and the culture specific growth rate. For optimization, central composite design of experiments followed by response surface methodology (RSM) was applied. Results showed that among the four independent variables, i.e., pH, agitation (rpm), temperature (°C), and inoculum age (h) investigated in this study, interaction effect between agitation and inoculum age as well as that between agitation and temperature were significant on both 4-CP biodegradation efficiency and culture specific growth rate. Also, at the RSM optimized settings of 7.5 pH, 207 rpm, 29.6°C and 39.5 h inoculum age, 100% biodegradation of 4-CP at a high initial concentration of 300 mg l⁻¹ was achieved within a short span of 18.5 h of culture. The enhancement in the 4-CP biodegradation efficiency was found to be 23% higher than that obtained at the unoptimized settings of the culture conditions. Results of batch growth kinetics of A. chlorophenolicus A6 for various 4-CP initial concentrations revealed that the culture followed substrate inhibition kinetics. Biokinetic constants involved in the process were estimated by fitting the experimental data to several models available from the literature.     

Plant regeneration via callus culture and subsequent in vitro flowering of Dendrobium huoshanense

A protocol for regenerating and subsequent in vitro flowering of an economical important and endangered medicinal orchid, Dendrobium huoshanense, was established mainly via indirect protocorm-like body (PLB) formation. A four-step method was developed to induce successful plant regeneration on 1/2 MS medium supplemented with suitable plant growth regulators (PGRs). Step 1 (callus induction): the root tip explants (1 cm long) were cultured at 1 mg l^?1 2,4-D + 1 mg l^?1 TDZ for 3 months. Step 2 (callus proliferation): the calli were subcultured with a 1-month interval at 1 mg l^?1 2,4-D + 1 mg l^?1 TDZ. Step 3 (PLB induction): the calli were cultured at 2 mg l^?1 NAA + 1 mg l^?1 BA for 2 months. Step 4 (plantlet conversion): the 2-month-old PLBs were cultured at 0.1 mg l^?1 IBA for 4 months. It took at least 6 months to produce well-rooted regenerated plantlets with an average of 3.2 roots and 3.6 leaves from the initial callus. The 6-month-old rooted plantlets were transferred onto PGR-free 1/2 MS medium for 6 months, and then potted with Sphagnum moss for acclimatization. After 2 month of culture, the survival rate was 100 %. The in vitro flowers were obtained on the 8-month-old plantlets at 1 mg l^?1 IBA, 5 mg l^?1 IBA and 0.1 mg l^?1 NAA, but the flowers showed a lack of the gynandrium. The abnormity was overcome by the aid of 5 mg l^?1 TDZ, and subsequently, the capsules formed without artificial pollination. This protocol provides the basis for further investigation on cell suspension, micropropagation, in vitro flowering and breeding programs in Dendrobium huoshanense.     

Laboratory-scale biofiltration of acrylonitrile by <Emphasis Type="Italic">Rhodococcus rhodochrous</Emphasis> DAP 96622 in a trickling bed bioreactor

Acrylonitrile (ACN), a volatile component of the waste generated during the production of acrylamide, also is often associated with aromatic contaminants such as toluene and styrene. Biofiltration, considered an effective technique for the treatment of volatile hydrocarbons, has not been used to treat volatile nitriles. An experimental laboratory-scale trickling bed bioreactor using cells of Rhodococcus rhodochrous DAP 96622 supported on granular activated carbon (GAC) was developed and evaluated to assess the ability of biofiltration to treat ACN. In addition to following the course of treatability of ACN, kinetics of ACN biodegradation during both recycle batch and open modes of operation by immobilized and free cells were evaluated. For fed-batch mode bioreactor with immobilized cells, almost complete ACN removal (>95%) was achieved at a flow rate of 0.1 μl/min ACN and 0.8 μl/min toluene (TOL) (for comparative purposes this is equivalent to 6.9 mg l^?1 h^?1 ACN and 83.52 mg l^?1 h^?1 TOL). In a single-pass mode bioreactor with immobilized cells, at ACN inlet loads of 100–200 mg l^?1 h^?1 and TOL inlet load of ~400 mg l^?1 h^?1, with empty bed retention time (EBRT) of 8 min, ACN removal efficiency was ~90%. The three-dimensional structure and characteristics of the biofilm were investigated using confocal scanning laser microscopy (CSLM). CLSM images revealed a robust and heterogeneous biofilm, with microcolonies interspersed with voids and channels. Analysis of the precise measurement of biofilm characteristics using COMSTAT^® agreed with the assumption that both biomass and biofilm thickness increased along the carbon column depth.     

Efficacy of Acinetobacter sp. B9 for simultaneous removal of phenol and hexavalent chromium from co-contaminated system

The present study shows the feasibility of a newly isolated strain Acinetobacter sp. B9 for concurrent removal of phenol and Cr (VI) from wastewater. The experiments were conducted in a batch reactor under aerobic conditions. Initially, when mineral salt solution was used as the culture medium, the strain was found to utilize phenol as sole carbon and energy source while no Cr (VI) removal was observed. However, the addition of glucose as co-carbon source resulted in the removal of both toxicants. This co-removal efficiency of the strain was further improved with nutrient-rich media (NB). Optimum co-removal was determined at 188 mg L^?1 of phenol and 3.5 mg L^?1 of Cr (VI) concentrations at pH 7.0. Strain B9 followed the orthometabolic pathway for phenol degradation. Transmission electron microscopy (TEM) and Fourier transform infrared spectroscopy (FT-IR) studies showed sorption of chromium as one of the major mechanisms for Cr (VI) removal by B9 cells. Acinetobacter sp. B9 was later on checked for bioremediation of real tannery wastewater. After 96 h of batch treatment of tannery effluent containing an initial 47 mg L^?1 phenol and 16 mg L^?1 Cr (VI), complete removal of phenol and 87 % reduction of Cr (VI) were attained, showing high efficiency of the bacterial strain for potential application in industrial pollution control.     

Aerobic decolorization of Acid Brilliant Scarlet GR by microbial community and the community dynamics during sequencing batch processes

In this research, aerobic decolorization of Acid Brilliant Scarlet GR by microbial community was studied. Effects of conditions and dye concentraion on decolorization processes were investigated. Additionally, continuous decolorization was evaluated through sequencing batch tests and the microbial dynamics during this process was analyzed by polymerase chain reaction-denaturing gradient gel electrophoresis. The results showed that 100 mg l^?1 of the dye was completely decolorized within 12 h, which was mainly caused by biodegradation. The optimal decolorization conditions were as follows: inoculation size 2.07 g l^?1 (wet cell pellet), rotation speed 150 r min^?1, pH 5.0–7.0 and 30 °C. The processes were well described by zero-order kinetics, and more than 700 mg l^?1 of the dye would inhibit the activity of the consortium. Furthermore, the microbial community exhibited high efficiency in sequencing batch processes for continuous decolorization. Microbial community structure shifted obviously when exposed to higher concentration of the dye (500 mg l^?1), and all the dominant microorganisms were affiliated with four different phyla of Actinobacteria, Bacteroidetes, Proteobacteria and Firmicutes.     

Production of volatile phenols by Lactobacillus plantarum in wine conditions

Some lactic acid bacteria produce volatile phenols in culture medium but this activity has not been extensively studied in wine conditions. Red and white wines were mixed with MRS medium at different ratios to study the influence of wine on the metabolism of p-coumaric and ferulic acids by Lactobacillus plantarum. In MRS broth supplemented with these precursors at 10 mg l^?1, only 4-ethylphenol was produced (1 mg l^?1) while, in the presence of wine, 4-vinylphenol was also obtained. Both volatile phenols are produced in nearly equal amounts (1 mg l^?1) or almost only 4-vinylphenol depending on the MRS:wine ratio. Thus, wine favours the accumulation of 4-vinylphenol. Ferulic acid was not or was weakly metabolized in the conditions studied.     

Simultaneous removal of hexavalent chromium and <Emphasis Type="Italic">o</Emphasis>-dichlorobenzene by isolated <Emphasis Type="Italic">Serratia marcescens</Emphasis> ZD-9

Heavy metals–organics mixture pollution is increasingly concerned and simultaneous removal of organic pollutants and heavy metals is becoming significant. In this study, a strain was isolated from the sediment of a tannery effluent outfalls, which can remove o-dichlorobenzene (o-DCB) and Cr(VI) simultaneously. The bacterial isolate was identified as Serratia marcescens by the 16S rRNA gene sequences. The strain removed about 90% of o-DCB and more than 80% of Cr(VI) at the concentration of 1.29 g L^?1 o-DCB and 20 mg L^?1 Cr(VI). In the presence of concomitant pollutant o-DCB, the optimal pH (8.0) and temperature (30 °C) were determined for Cr(VI) removal. Changes of the bacterial cells and intracellular black Cr(III) sediments were observed by the TEM auxiliary analysis. The results of the FTIR spectroscopy analysis indicated that hydroxyl, amide and polysaccharides were involved in the process of Cr(VI) removal.     

Kinetics study of pyridine biodegradation by a novel bacterial strain,Rhizobium sp. NJUST18

Biodegradation of pyridine by a novel bacterial strain, Rhizobium sp. NJUST18, was studied in batch experiments over a wide concentration range (from 100 to 1,000 mg l^?1). Pyridine inhibited both growth of Rhizobium sp. NJUST18 and biodegradation of pyridine. The Haldane model could be fitted to the growth kinetics data well with the kinetic constants μ* = 0.1473 h^?1, K _s = 793.97 mg l^?1, K _i = 268.60 mg l^?1 and S _m = 461.80 mg l^?1. The true μ _max, calculated from μ*, was found to be 0.0332 h^?1. Yield coefficient Y _X/S depended on S _i and reached a maximum of 0.51 g g^?1 at S _i of 600 mg l^?1. V _max was calculated by fitting the pyridine consumption data with the Gompertz model. V _max increased with initial pyridine concentration up to 14.809 mg l^?1 h^?1. The q _S values, calculated from $V_{ \hbox{max} }$ , were fitted with the Haldane equation, yielding q _Smax = 0.1212 g g^?1 h^?1 and q* = 0.3874 g g^?1 h^?1 at S _m′ = 507.83 mg l^?1, K _s′ = 558.03 mg l^?1, and K _i′ = 462.15 mg l^?1. Inhibition constants for growth and degradation rate value were in the same range. Compared with other pyridine degraders, μ _max and S _m obtained for Rhizobium sp. NJUST18 were relatively high. High K _i and K _i′ values and extremely high K _s and K _s′ values indicated that NJUST18 was able to grow on pyridine within a wide concentration range, especially at relatively high concentrations.     

Effect of culture conditions, cytokinins, methyl jasmonate and salicylic acid on the biomass accumulation and production of withanolides in multiple shoot culture of Withania somnifera (L.) Dunal using liquid culture

The influence of cytokinins and culture conditions including medium volume, harvest time and elicitation with abiotic elicitors (SA/MeJ) have been studied for the optimal production of biomass and withanolides in the multiple shoot culture of Withania somnifera. Elicitation of shoot inoculum mass (2 g l^?l FW) with SA at 100 μM in the presence of 0.6 mg l^?l BA and 20 mg l^?l spermidine for 4 h exposure time at the 4th week in 20 ml liquid medium recorded higher withanolides production (withanolides A [8.48 mg g^?l DW], withanolides B [15.47 mg g^?l DW], withaferin A [29.55 mg g^?l DW] and withanone [23.44 mg g^?l DW]), which were 1.14 to 1.18-fold higher than elicitation with MeJ at 100 μM after 5 weeks of culture. SA-elicited cultures did not exhibit much variation in biomass accumulation when compared to control. This cytokinin induces and SA-elicited multiple shoot culture protocol provides a potential alternative for the optimal production of biomass and withanolides utilizing liquid culture.     

Embryogenesis,plant regeneration and cardiac glycoside determination in Digitalis ferruginea subsp. ferruginea L

The present study reports, for the first time, an efficient in vitro plant regeneration protocol for Digitalis ferruginea subsp. ferruginea L. (rusty foxglove). We have used different concentrations of gibberellic acid (GA₃) on Murashige and Skoog (MS) medium to assess the germination frequency of seeds. High frequency of germination was achieved on MS medium with 1.0 mg l^?1 GA₃. 6-Benzylaminopurine (BAP) combined with α-naphtaleneacetic acid (NAA) or 2, 4-dichlorophenoxy acetic acid (2, 4-D) in the induction MS medium induced both somatic embryogensis and shoot organogenesis. The highest percentage of callus growth (85 %) was obtained when hypocotyl explants were cultured on MS medium containing 0.5 mg l^?1 2, 4-D plus 1.0 mg l^?1 BAP. The maximum mean number of somatic embryos (7.3 ± 1.3 embryos) or shoots (12.0 ± 1.1 shoots) per callus was obtained when medium contained 0.25 mg l^?1 NAA plus 1.0 mg l^?1 BAP or 0.5 mg l^?1 NAA plus 2.0 mg l^?1 BAP. The regenerated shoots easily rooted on MS medium. Higher amounts of lanatoside C [13.2 ± 0.5 mg 100 g^?1 dry weight (dw)] and digoxin (2.93 ± 0.31 mg 100 g^?1 dw) accumulation were obtained when shoots were obtained by indirect regeneration. We also investigated derivatives of cardenolides, i.e., digitoxigenin (730 ± 180 mg 100 g^?1 dw), gitoxigenin (50 ± 20 mg 100 g^?1 dw) and digoxigenin (490 ± 170 mg 100 g^?1 dw) from natural samples.     

Effects of stationary phase elongation and initial nitrogen and phosphorus concentrations on the growth and lipid-producing potential of Chlorella sp. HQ

Higher lipid production and nutrient removal rates are the pursuing goals for synchronous biodiesel production and wastewater treatment technology. An oleaginous alga Chlorella sp. HQ was tested in five different synthetic water, and it was found to achieve the maximum biomass (0.27 g L^?1) and lipid yield (41.3 mg L^?1) in the synthetic secondary effluent. Next, the effects of the stationary phase elongation and initial nitrogen (N) and phosphorus (P) concentrations were investigated. The results show that the algal characteristics were affected apparently under different N concentrations but not P, which were verified by Logistic and Monod models. At the early stationary phase, the algal biomass, lipid and triacylglycerols (TAGs) yields, and P removal efficiency increased and reached up to 0.19 g L^?1, 46.7 mg L^?1, 14.3 mg L^?1, and 94.3 %, respectively, but N removal efficiency decreased from 86.2 to 26.8 % under different N concentrations. And the largest TAGs yield was only 6.4 mg L^?1 and N removal efficiency was above 71.1 % under different P concentrations. At the late stationary phase, the maximal biomass, lipid and TAGs yields, and P removal efficiencies primarily increased as the initial N and P concentrations increase and climbed up to 0.49 g L^?1, 99.2 mg L^?1, 54.0 mg L^?1, and 100.0 %, respectively. It is concluded that stationary phase elongation is of great importance and the optimal initial N/P ratio should be controlled between 8/1 and 20/1 to serve Chlorella sp. HQ for better biodiesel production and secondary effluent purification.     

Occurrence of 4-tert-butylphenol (4-t-BP) biodegradation in an aquatic sample caused by the presence of Spirodela polyrrhiza and isolation of a 4-t-BP-utilizing bacterium

Although 4-tert-butylphenol (4-t-BP) is a serious aquatic pollutant, its biodegradation in aquatic environments has not been well documented. In this study, 4-t-BP was obviously and repeatedly removed from water from four different environments in the presence of Spirodela polyrrhiza, giant duckweed, but 4-t-BP persisted in the environmental waters in the absence of S. polyrrhiza. Also, 4-t-BP was not removed from autoclaved pond water with sterilized S. polyrrhiza. These results suggest that the 4-t-BP removal from the environmental waters was caused by biodegradation stimulated by the presence of S. polyrrhiza rather than by uptake by the plant. Moreover, Sphingobium fuliginis OMI capable of utilizing 4-t-BP as a sole carbon and energy source was isolated from the S. polyrrhiza rhizosphere. Strain OMI degraded 4-t-BP via a meta-cleavage pathway, and also degraded a broad range of alkylphenols with linear or branched alkyl side chains containing two to nine carbon atoms. Root exudates of S. polyrrhiza stimulated 4-t-BP degradation and cell growth of strain OMI. Thus, the stimulating effects of S. polyrrhiza root exudates on 4-t-BP-degrading bacteria might have contributed to 4-t-BP removal in the environmental waters with S. polyrrhiza. These results demonstrate that the S. polyrrhiza–bacteria association may be applicable to the removal of highly persistent 4-t-BP from wastewaters or polluted aquatic environments.     

Transgenic <Emphasis Type="Italic">Eustoma grandiflorum</Emphasis> expressing the <Emphasis Type="Italic">bar</Emphasis> gene are resistant to the herbicide Basta<Superscript>®</Superscript>

Lisianthus (Eustoma grandiflorum) is a cut or ornamental flower that is popular all over the world. This ornamental crop, however, lacks an effective weed control method due to its susceptibility to herbicide. In this study, transgenic plants of a lisianthus cultivar were produced using Agrobacterium-mediated delivery of the plasmid pCAMBIA3300, which carried the bialaphos resistance (bar) gene under driven by the CaMV 35S promoter. The transgenic calli were derived from wounded edges of the leaves grown on a shoot regeneration medium containing 100 mg l^?1 cefotaxime and 2 mg l^?1 glufosinate ammonium for 4 weeks. The callus that was detached from the wounded edge of the leaf was transferred to the shoot regeneration medium with 100 mg l^?1 cefotaxime and 5 mg l^?1 glufosinate ammonium for 4 weeks for shoot regeneration. The bar gene integration and expression in the transgenic plants were confirmed by Southern and Northern blot analyses, respectively. Subsequently, the transgenic lines were assessed in vitro and under greenhouse conditions for their resistance to the commercial herbicide Basta^®, which contains glufosinate ammonium as the active component. Six transgenic lines showed high percentages (67–80%) of survival in vitro under the selection condition with glufosinate ammonium (up to 216 mg l^?1). Under greenhouse conditions, the plants from these six lines remained healthy and exhibited a normal phenotype after spraying with glufosinate ammonium (up to 1,350 mg l^?1). This is the first paper to provide a detailed survey of transgenic lisianthus expressing the bar gene and exhibiting herbicide-resistance under greenhouse conditions.     

In vitro propagation and micromorphological studies of Cleome gynandra: a C4 model plant closely related to Arabidopsis thaliana

We developed a micropropagation protocol for Cleome gynandra, a C₄ model plant with medicinal importance. Surface-sterilized nodal segments obtained from 1 to 2-month-old field grown plant were used as explants for culture establishment and plant regeneration. Multiple shoots differentiated through bud breaking on Murashige and Skoog (MS) medium with different concentrations of benzyladenine (BA) and kinetin (Kin). The optimum shoot differentiation occurred on medium with 1.5 mg l^?1 BA. Out of various concentrations and combinations of cytokinins and auxins, MS medium containing 0.5 mg l^?1 BA and 0.1 mg l^?1 IAA (indole-3-acetic acid) was found best for shoot multiplication. However, the differentiated shoots exhibited hyperhydration, leaf curling and early leaf fall during subculturing. To overcome these problems, regenerated shoots were transferred to the modified MS medium with reduced nitrates (825 mg l^?1 NH₄NO₃ and 950 mg l^?1 KNO₃) and 100 mg l^?1 (NH₄)₂SO₄. The micropropagated shoots were rooted (i) in vitro on one-fourth strength of MS salts with 0.25 mg l^?1 each of IBA (indole-3 butyric acid) and NOA (2-naphthoxyacetic acid) + 100 mg l^?1 activated charcoal, and (ii) ex vitro, by treating the shoot base(s) with 200 mg l^?1 of IBA for 3 min and transferred to soilrite moistened with one-fourth strength of MS macro salts in culture bottles. The plants were hardened in the greenhouse with 85 % survival rate. Micromorphological studies of the plants were conducted during hardening with reference to development and changes in vein spacing, glandular trichome and stomata. In comparison to leaves under in vitro condition, higher density of veins and glandular trichomes was observed in the leaves of hardened plants. In addition, stomata became functional during hardening which were non-functional under in vitro condition.     

Degradation of cationic surfactants using Pseudomonas putida A ATCC 12633 immobilized in calcium alginate beads

In this study, the degradation of tetradecyltrimethylammonium bromide (TTAB) by freely suspended and alginate-entrapped cells from the bacteria Pseudomonas putida (P. putida) A ATCC 12633 was investigated in batch cultures. The optimal conditions to prepare beads for achieving a higher TTAB degradation rate were investigated by changing the concentration of sodium alginate, pH, temperature, agitation rate and initial concentration of TTAB. The results show that the optimal embedding conditions of calcium alginate beads are 4 % w/v of sodium alginate content and 2 × 10⁸ cfu ml^?1 of P. putida A ATCC 12633 cells that had been previously grown in rich medium. The optimal degradation process was carried out in pH 7.4 buffered medium at 30 °C on a rotary shaker at 100 rpm. After 48 h of incubation, the free cells degraded 26 mg l^?1 of TTAB from an initial concentration of 50 mg l^?1 TTAB. When the initial TTAB concentration was increased to 100 mg l^?1, the free cells lost their degrading activity and were no longer viable. In contrast, when the cells were immobilized on alginate, they degraded 75 % of the TTAB after 24 h of incubation from an initial concentration of 330 mg l^?1 of TTAB. The immobilized cells can be stored at 4 °C for 25 days without loss of viability and can be reused without losing degrading capacity for three cycles.     

4-Chlorophenol degradation by a bacterial consortium: development of a granular activated carbon biofilm reactor

A bacterial consortium that can degrade chloro- and nitrophenols has been isolated from the rhizosphere of Phragmitis communis. Degradation of 4-chlorophenol (4-CP) by a consortium attached to granular activated carbon (GAC) in a biofilm reactor was evaluated during both open and closed modes of operation. During the operation of the biofilm reactor, 4-CP was not detected in the column effluent, being either adsorbed to the GAC or biodegraded by the consortium. When 4-CP at 100 mg l⁻¹ was fed to the column in open mode operation (20 mg g⁻¹ GAC total supply), up to 27% was immediately available for biodegradation, the rest being adsorbed to the GAC. Biodegradation continued after the system was returned to closed mode operation, indicating that GAC bound 4-CP became available to the consortium. Biofilm batch cultures supplied with 10–216 mg 4-CP g⁻¹ GAC suggested that a residual fraction of GAC-bound 4-CP was biologically unavailable. The consortium was able to metabolise 4-CP after perturbations by the addition of chromium (Cr VI) at 1–5 mg l⁻¹ and nitrate at concentrations up to 400 mg l⁻¹. The development of the biofilm structure was analysed by scanning electron microscopy and confocal laser scanning microscopy (CLSM) techniques. CLSM revealed a heterogeneous structure with a network of channels throughout the biofilm, partially occupied by microbial exopolymer structures. Received: 17 March 1999 / Received revision: 27 May 1999 / Accepted: 28 May 1999