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1.
The objective of this study was to assess the efficacy of a bench-scale, acetate-fed, packed bed bioreactor (PBR) to treat low concentrations (>1 mg L?1) of perchlorate (ClO4 ?) in groundwater collected from an impacted site. The PBR consisted of a cylindrical plexiglass column packed with Celite, a diatomaceous earth product, as a solid support medium. The reactor was inoculated with a ClO?4 ?-reducing bacterial isolate, perclace. Results showed that with influent ClO4 ? concentrations of approximately 800 μg L?1, nondetectable effluent concentrations (>4 μg L?1) were achieved with the PBR/perclace system at residence time as low as 0.3 h. Influent acetate concentrations of less than 500 mg L?1 yielded nondetectable effluent ClO? 4 concentrations, and acetate concentrations generally less than 50 mg L?1 were present in the effluent. Nitrate (NO? ?3) was also removed in this system, while sulfate (SO4 2?) reduction was not observed. The pH remained relatively constant during the process. 相似文献
2.
Microorganism with simultaneous nitrification and denitrification ability plays a significant role in nitrogen removal process, especially in the eutrophic waters with excessive nitrogen loads. The nitrogen removal capacity of microorganism may suffer from low temperature or nitrite nitrogen source. In this study, a hypothermia aerobic nitrite-denitrifying bacterium, Pseudomonas tolaasii strain Y-11, was selected to determine the simultaneous nitrification and denitrification ability with mixed nitrogen source at 15 °C. The sole nitrogen removal efficiencies of strain Y-11 in simulated wastewater were obtained. After 24 h of incubation at 15 °C, the ammonium nitrogen fell below the detection limit from an initial value of 10.99 mg/L. Approximately 88.0 ± 0.33% of nitrate nitrogen was removed with the initial concentration of 11.78 mg/L and the nitrite nitrogen was not detected with the initial concentration of 10.75 mg/L after 48 h of incubation at 15 °C. Additionally, the simultaneous nitrification and denitrification nitrogen removal ability of P. tolaasii strain Y-11 was evaluated using low concentration of mixed NH4+-N and NO3?–N/NO2?–N (about 5 mg/L-N each) and high concentration of mixed NH4+–N and NO3?–N/NO2?–N (about 100 mg/L-N each). There was no nitrite nitrogen accumulation at the time of evaluation. The results demonstrated that P. tolaasii strain Y-11 had higher simultaneous nitrification and denitrification capacity with low concentration of mixed inorganic nitrogen sources and may be applied in low temperature wastewater treatment. 相似文献
3.
S. Wuertz S.G.E. Schulze U. Eberhardt C. Schulz J.P. Schroeder 《Comparative biochemistry and physiology. Toxicology & pharmacology : CBP》2013,157(4):352-360
Pike-perch Sander lucioperca is currently considered as one of the most promising candidates for production in freshwater recirculation aquaculture systems (RAS). Here, due to the lack of studies on nitrite (NO2?) toxicity in pike-perch, a flow-through exposure at 0, 0.44, 0.88, 1.75, 3.5, 7, 14 and 28 mg/L NO2?–N was carried out to determine the acute and chronic toxicity over a period of 32 days. In juvenile pike-perch, 120 h LC50 was 6.1 mg/L NO2?–N and at ≥ 14 mg/L NO2?–N all fish had died within 24 h. Chronic exposure revealed a significant build up of NO2? in the plasma as well as in the muscles at ≥ 0.44 mg/L NO2?–N peaking in fish exposed to the highest concentration of 3.5 mg/L NO2?–N after 32 days. Still, due to high individual variation methemoglobin (MetHb) was only significantly increased (p < 0.01) at 3.5 mg/L NO2?–N. No adverse effects on red blood cells (RBC) and hematocrit were observed in any of the treatments. In a second experiment, compensation of NO2? toxicity at increasing chloride concentrations (40 (freshwater), 65, 90, 140, 240, 440 mg/L Cl?) was observed at a constant exposure of 10 mg/L NO2?–N for 42 days. At ≥ 240 mg/L Cl?, NO2? build-up in blood plasma and muscle was completely inhibited. At lower Cl? concentrations (≤ 140 mg/L), NO2? was significantly increased in plasma, but only insignificantly elevated in muscle due to high individual variation. MetHb was increased significantly difference only at 40 mg/L Cl? (freshwater control) compared to the control. Again, high individual variations were observed. As a conclusion, S. lucioperca is moderately sensitive towards NO2? and acceptable levels in RAS should hence not exceed 1.75 mg/L NO2?–N to avoid MetHb formation. However, based on the 120 h LC50 and a factor of 0.01 according to Sprague (1971), a NO2? concentration of ≤ 0.061 mg/L NO2?–N is considered as “safe.” Thereby, no NO2? should accumulate in the plasma or muscle tissue during chronic exposure. For 10 mg/L NO2?–N, ≥ 240 mg/L chloride compensates for NO2? uptake in plasma and muscle. 相似文献
4.
Celia E. Deane-Drummond 《Physiologia plantarum》1986,66(1):115-121
Net nitrate uptake, 36ClO?3/NO?3 influx and 36Cl? influx into Pisum sativum L. cv. Feltham First seedlings have been examined following growth in culture medium containing different combinations of chloride and nitrate. When young (6 days old) seedlings, that had been grown in the absence of N were used, nitrate accumulation stimulated net nitrate uptake and 36ClO?3/NO?3 influx (r2= 0.99) while chloride accumulation inhibited nitrate uptake and 36ClO?3/NO?3 influx (r2= 0.65). When nitrate was provided during growth there was no effect of chloride pretreatment on net nitrate uptake and there was little effect of total [NO?3+ Cl?]i on 36ClO?3/NO?3 influx (r2= 0.26). A direct effect of Cl? on 36ClO?3/NO?3 influx was only found when seedlings had been starved of N for more prolonged periods (14 days). When moderate chloride was supplied during growth, 36Cl? influx was insensitive to nitrate or chloride accumulated, but significantly correlated with loge [NO?3+ Cl?]i (r2= 0.75). When trace amounts of Cl? were supplied during growth 36Cl? influx was inhibited by (a) NO?3 in the external medium and (b) Cl? pretreatment, but was insensitive to NO?3 pretreatment. The sensitivity of 36Cl? influx to external nitrate was not found following Cl? pretreatment in the absence of nitrate. The possibility that there are two populations of chloride carriers which differ in their sensitivity to external nitrate is discussed. Tentative schematic models to account for the regulation of nitrate and chloride uptake are proposed in the context of current hypotheses for regulation of ion transport and control systems theory. 相似文献
5.
Hongyu Wang Yaping Ye Jinni Sai Dan Chen Kai Yang Jun Zhou 《Geomicrobiology journal》2016,33(6):505-512
Static experiments were conducted to investigate the effects of environmental factors on nitrate (NO3?-N)-removal efficiency, such as NO3?-N loading, pH value, C/N ratio and temperature in activated sludge using Fe (II) as electron donor. The results demonstrated that the average denitrification rate increased from 1.25 to 2.23 mg NO3?-N/(L·h) with NO3?-N loading increased from 30 to 60 mg/L. When pH increased from 7 to 8, the concentration of NO3?-N and nitrite (NO2?-N) in effluent were all maintained at quite low levels. C/N ratio had little impact on denitrification process, i.e., inorganic carbon (C) source could still be enough for denitrification process with C/N ratio as low as 5. Temperature had a significant effect on the denitrification efficiency, and NO3?-N removal efficiency of 92.03%, 96.77%, 97.67% and 98.23% could be obtained with temperature of 25°C, 30°C, 35°C and 40°C, respectively. SEM, XRD and XRF analysis was used to investigate microscopic surface morphology and chemical composition of the denitrifying activated sludge, and mechanism of the nitrate-dependent anaerobic ferrous oxidation (NAFO) bacterias could be explored with this research. 相似文献
6.
Plínio S. Furtado Manuel A. J. Valenzuela Gabriela Rodriguez-Fuentes Bruno R. Campos Wilson Wasielesky Jr. Gabriela Gaxiola 《Marine and Freshwater Behaviour and Physiology》2016,49(3):201-211
The aim of this study was to determine the effects of nitrite on the growth and survival of the white shrimp L. vannamei in two different salinities. Nitrite concentrations tested in salinity 8 g/L were 0 (control), 2.5, 5.0, 10.0, and 20.0 mg NO2?-N/L, and in salinity 24 g/L were 0 (control), 5.0, 10.0, 20.0, and 40.0 mg NO2?-N/L. For these experiments, 30 experimental units with 30?L of useful volume were stocked with 20 juvenile L. vannamei (8.0 ± 0.50 g), corresponding to a stocking density of 100 shrimp/m2, and cultivated for an experimental period of 30 days. A significant difference was found between the control and treatment groups with respect to growth and survival. The 2.5 mg NO2?-N/L treatment showed the best performance indexes in salinity 8 g/L, while the best growth performance indexes were found in the control and 5.0 mg NO2?-N/L treatments in salinity 24 g/L. Total mortality was observed in the 10 and 20 mg NO2?-N/L treatment groups from salinity 8 g/L and in the 40 mg NO2?-N/L treatment group in salinity 24 g/L. This study determined that concentrations of nitrite of up to 2.5 and 10 mg/L are acceptable for the rearing of L. vannamei in salinities of 8 and 24 g/L, respectively. 相似文献
7.
The Wei River is the largest tributary of the Yellow River in China. To understand the sources and cycling of nitrate in the Wei River, we determined the concentrations and nitrogen and oxygen isotopic values of nitrate from water samples. Our results revealed that NO3?-N dominated the inorganic N and ranged from 0.1 to 8.8 mg/L (averaging 3.3 mg/L). Although this NO3?-N concentration does not exceed the World Health Organization's drinking water standard of 10 mg/L, the NO3?-N content of most water samples exceeded 3 mg/L, indicating poor water quality. The NO3?-N concentrations and δ15N-NO3? values demonstrate that there are significant differences in the spatial distribution of nitrogen between the tributaries and the main stream of the Wei River. In addition, a negative linear relationship (r2 = 0.63) between NO3?-N concentrations and δ18O-NO3? values suggests mixing between two distinct sources (fertilizer and manure or sewage). Furthermore, we infer that the main source of nitrate is not manure or sewage itself, but rather the nitrification of NH4+ in manure and sewage. Finally, no obvious denitrification processes were observed. These results expand our understanding of sewage as a major source of nitrate to the Wei River, emphasizing the role of nitrification. 相似文献
8.
Kyoden Yasumoto Kimikazu Iwami Toshihiko Yonezawa Hisateru Mitsuda 《Phytochemistry》1977,16(9):1351-1354
Activation by different anions of γ-glutamyltransferase obtained in a. particulate form from fruiting bodies of Lentinus edodes has been studied using either L-γ-glutamyl-p-nitroanlide or lentinic acid as substrate. The mushroom transferase was activated by SCN?, NO3?, Cl?, Br?, ClO3?, Bro3?, N3?, I? and F?, but not those alkali and earth cations previously believed to activate the animal transferase, nor by citrate, claimed to be effective for the kidney bean transferase. Among anions proved hardly to activate the transferase were ClO4?, NO2?, HCO3?, H2PO4?, SO32? and SO42?. A high concentration of these anions more or less impeded the halide activation. Kinetic studies revealed that halides function as activators of increasing Vmax while keeping Km constant. These observations appeared least compatible with the possibility that the anion activation might involve a non-specific effect of high solute concentration, viz. dissociation of the enzyme from the supporting structure in the particulates. The activating effect of halides described here probably extends also to the animal enzymes. 相似文献
9.
Abstract Depending on the applied concentration, nitrate can either stimulate or inhibit germination of dormant seeds of Sinapis arvensis L. (wild mustard). Seed NO?3 levels that corresponded to the maximal germination frequencies recorded, ranged from 0.3 to 4.4 nmol seed?1 for applied NO?3 concentrations between 2.5 and 20 mol m?3. Germination was significantly lower in seeds containing more than 5 nmol NO?3. Although the presence of NO?3 within the seed was required to promote germination, seed NO?3 levels were 5 to 15 times less than levels calculated from the volume of solution taken up by the seeds. Seed NO?3 levels also responded in a passive fashion to the external NO3? concentration when the seeds were incubated on filter paper or in soil. In a representative soil containing 26 mg NO?3-N per kg dry weight and 8 to 16% water by weight, the amount of NO?3 taken up by the seeds reached inhibitory levels. Our results suggest that levels of inorganic nitrogen (NO?3, NH+4) in managed soils may play an important role in regulating germination of dormant S. arvensis seeds. 相似文献
10.
Jun Feng Su Sheng Chen Zheng Ting Lin Huang Fang Ma Si Cheng Shao Shao Fei Yang 《Geomicrobiology journal》2016,33(7):586-591
A novel bacterium, strain SZ28, identified as Acinetobacter sp., showed anaerobic denitrification ability using Mn(II) as the electron donor. Nitrate-nitrogen concentration decreased from nearly 16.52–mg L?1 to 4.4–mg L?1, without accumulation of nitrite as an intermediate, with a maximum of 0.063–mg NO3?-N L?1 h?1, reaching a peak of 0.085–mg NO3?-N L?1 h?1 in sodium acetate. The nitrate removal rate reached 0.067–mg NO3?-N L?1 h?1, 0.059–mg NO3?-N L?1 h?1, and 0.078 mg NO3?-N L?1 h?1 using Mn(II), S(II), and Fe(II) as electron donors, respectively. The optimum pH was 6.0, with a removal rate of 0.063–mg NO3?-N L?1 h?1 相似文献
11.
Complexes of dioxouranium(VI) with the amino acid L-arginine have been prepared and studied by ir and pmr measurements. The results indicate the formation of UO2L XnH2O (L = Arg?; X = NO3?, CH3COO?, or ClO4?; n = 2 or 3). The bonding involves carboxylato-, amino-, and probably guanido groups of the ligand. The coordination sphere of dioxouranium(VI) also includes nitrate or acetate, but not the perchlorate group. Uranyl(VI) may reach the coordination number of 5 in the equatorial plane by the coordination of moleclues of H2O. 相似文献
12.
Wijaya Altekar 《Biopolymers》1977,16(2):341-368
The effects of anions of neutral salts on the fluorescence emission of six proteins as well as on tryptophan and tyrosine were studied in relation to the structure of proteins. Most anions are good quenchers of tryptophyl and tyrosyl fluorescence, free or in proteins. The results with tryptophan and tyrosine indicate involvement of a collisional quenching mechanism due to agreement with Stern–Volmer law. The deactivation of fluorescence probably occurs because of the transition from singlet state to triplet state. Lehrer's modification of Stern–Volmer law was applied to proteins. The effective quenching constants ([KQ]eff) and the fraction of fluorescence available ([fa]eff) to the quencher are also calculated. In contrast to its effect on tryptophan, CH3COO? quenches tyrosyl fluorescence and ClO4? does not. The effects on fluorescence of ribonuclease and free tyrosine are similar and without any changes in emission maximum. The anions are divided into three groups based on the effect they have on tryptophan-containing proteins. (1) NO3?, NO2?, Br?, and I? have high [KQ]eff values and readily quench tryptophyl fluorescence of proteins causing a shift of emission maximum to a shorter wavelength. This change is due to the specific quenching of “exposed” tryptophan residues which are accessible to quenchers and the observed residual fluorescence is from the “buried” tryptophyls. (2) ClO4? and SCN? also quench fluorescence of tryptophan in proteins and have lower ([KQ]eff) values. In their presence the fluorescence maximum is shifted to a longer wavelength, which indicates the unfolding of a protein with [(fa)eff] = 1. (3) Cl?, CH3COO?, and SO4? do not have a direct effect on the fluorescence of tryptophan. Besides the “direct” effects, “indirect” effects on fluorophors in protein are also seen, pointing out that the neutral salts can interact in more than one manner with proteins. The effectiveness of anions in quenching fluorescence of proteins follows similar sequences which almost resemble the Hofmeister series, viz., SO4=, CH3COO? ? Cl? < ClO4? < SCN? < Br? < I? < NO3? < NO2?. 相似文献
13.
Effect of influent substrate ratio on anammox process was studied in sequencing batch reactor. Operating temperature was fixed at 35 ± 1 °C. Influent pH and hydraulic retention time were 7.5 and 6 h, respectively. When influent NO2 ?-N/NH4 +-N was no more than 2.0, total nitrogen removal rate (TNRR) increased whereas NH4 +-N removal rate stabilized at 0.32 kg/(m3 d). ΔNO2 ?-N/ΔNH4 +-N increased with enhancing NO2 ?-N/NH4 +-N. When NO2 ?-N/NH4 +-N was 4.5, ΔNO2 ?-N/ΔNH4 +-N was 1.98, which was much higher than theoretical value (1.32). The IC50 of NO2 ?-N was 289 mg/L and anammox activity was inhibited at high NO2 ?-N/NH4 +-N ratio. With regard to influent NH4 +-N/NO2 ?-N, the maximum NH4 +-N removal rate was 0.36 kg/(m3 d), which occurred at the ratio of 4.0. Anammox activity was inhibited when influent NH4 +-N/NO2 ?-N was higher than 5.0. With influent NO3 ?-N/NH4 +-N of 2.5–6.5, NH4 +-N removal rate and NRR were stabilized at 0.33 and 0.40 kg/(m3 d), respectively. When the ratio was higher than 6.5, nitrogen removal would be worsened. The inhibitory threshold concentration of NO2 ?-N was lower than NH4 +-N and NO3 ?-N. Anammox bacteria were more sensitive to NO2 ?-N than NH4 +-N and NO3 ?-N. TNRR would be enhanced with increasing nitrogen loading rate, but sludge floatation occurred at high nitrogen loading shock. The Han-Levenspiel could be applied to simulate nitrogen removal resulting from NO2 ?-N inhibition. 相似文献
14.
NH4+ and NO3? uptake were measured by continuous sampling with an autoanalyzer. For Hypnea musciformis (Wulfen) Lamouroux, NO3?up take followed saturable kinetics (K2=4.9 μg-at N t?1, Vmax= 2.85 μg- at N, g(wet)?1. h?1. The ammonium uptake data fit a trucatd hyperbola, i.e., saturation was not reach at the concentrations used. NO3? uptake was reduced one-half in the presence of NH4+, but presence of NO3? had no effect on NH4+ uptake. Darkness reduced both NO3? and NH4+ uptake by one-third to one-half. For Macrocystis pyrufera (L) C. Agardh, NO3? uptake followed saturable kinetices: K2=13.1 μg-at N. l?1. Vmax=3.05 μg-at N. g(wet)?1. h?1.NH4+ uptake showed saturable kinetics at concentration below 22 μg-at N l -1 (K2=5.3 μg-at N.1–1, Vmax= 2.38 μg-at N G (wet)?1.h?1: at higher concentration uptake increased lincarly with concentrations. NO3?and NH4+ were taken up simulataneously: presence of one form did not affect uptake of the other. 相似文献
15.
Denitrification and nitrous oxide effluxes in boreal, eutrophic river sediments under increasing nitrate load: a laboratory microcosm study 总被引:1,自引:0,他引:1
Hanna Silvennoinen Anu Liikanen Jouni Torssonen C. Florian Stange Pertti J. Martikainen 《Biogeochemistry》2008,91(2-3):105-116
Intact sediment cores from rivers of the Bothnian Bay (Baltic Sea) were studied for denitrification based on benthic fluxes of molecular nitrogen (N2) and nitrous oxide (N2O) in a temperature controlled continuous water flow laboratory microcosm under 10, 30, 100, and 300 μM of 15N enriched nitrate (NO3 ?, ~98 at. %). Effluxes of both N2 and N2O from sediment to the overlying water increased with increasing NO3 ? load. Although the ratio of N2O to N2 increased with increasing NO3 ? load, it remained below 0.04, N2 always being the main product. At the NO3 ? concentrations most frequently found in the studied river water (10–100 μM), up to 8% of the NO3 ? was removed in denitrification, whereas with the highest concentration (300 μM), the removal by denitrification was less than 2%. However, overall up to 42% of the NO3 ? was removed by mechanisms other than denitrification. As the microbial activity was simultaneously enhanced by the NO3 ? load, shown as increased oxygen consumption and dissolved inorganic carbom efflux, it is likely that a majority of the NO3 ? was assimilated by microbes during their growth. The 15N content in ammonium (NH4 +) in the efflux was low, suggesting that reduction of NO3 ? to NH4 + was not the reason for the NO3 ? removal. This study provides the first published information on denitrification and N2O fluxes and their regulation by NO3 ? load in eutrophic high latitude rivers. 相似文献
16.
17.
A monitoring network of nine sites was established to determine the spatial and temporal variation of atmospheric nitrogen (N) deposition in the North China Plain (NCP) over a two-year period. The annual bulk deposition of inorganic N in the North China Plain ranged from 18.4 to 38.5 kg/hm2 and averaged 28.0 kg/hm2. The concentration of NH4+-N and NO3?-N in rainwater averaged 3.76 and 1.85 mg/L, respectively, which were significantly higher than the values at background sites in China (normally less than 0.5 mg/L). Annual bulk deposition of inorganic N in the Beijing area (32.5 kg/hm2) was higher than that in Shandong and Hebei provinces (21.2 kg/hm2 on an average). Also bulk N deposition was much greater in Dongbeiwang and Fangshan than in Yanqing and Shunyi counties. Significant spatial variation of bulk deposition was observed in the Beijing area because of variation of precipitation, and 60% of bulk deposition occurred from June to September. Bulk deposition of NH4+-N was 2.0 times that of NO3?-N deposition at the rural monitoring sites. However, the situation was reversed at the Beijing Academy of Agricultural-Forestry Sciences (BAAFS), the unique urban monitoring site. The results suggest that reduced N in precipitation is dominant in rural regions, but oxidized N is the major form in urban regions. The positive relationship between inorganic N deposition and precipitation can be fitted by a power equation (r2= 0.67), showing an increase of NH4+-N and NO3?-N inputs with increased precipitation. Wet deposition of N accounted for 73% of the bulk deposition, implying that dry deposition of N, particularly NH4+-N from dust, is important in the North China Plain. 相似文献
18.
Sarah H. Ledford Laura K. Lautz Philippe G. Vidon John C. Stella 《Biogeochemistry》2017,133(3):317-331
Nitrate (NO3 ?) dynamics in urban streams differ from many natural streams due to stormwater runoff, sewage inputs, decreased groundwater discharge, often limited hyporheic exchange, increased primary productivity, and limited carbon input. We investigated NO3 ? dynamics in a first-order urban stream in Syracuse, NY, which has urbanized headwaters and a geomorphologically natural downstream section. Twice-monthly water sampling, NO3 ? injection tests, NO3 ? isotopic analysis, filamentous algae mat density, and riparian shading were used to identify processes regulating NO3 ? dynamics in the stream over a 12-month period. The urban headwater reach had low NO3 ? (0.006–0.2 mg N/L) in the spring through fall, with a minimum uptake length of 900 m, no canopy cover, and high algae mat density. The downstream natural reach (100% canopy cover during the summer and low algae mat density) had nitrate concentrations between 0.6 and 1.2 mg N/L from winter to summer, which decreased during autumn leaf-off. In the urban reach, autotrophic uptake by filamentous green algae is a major NO3 ? sink in summer. In the natural reach, the addition of organic matter to the stream at leaf-off led to a decrease in NO3 ? concentration followed by an increase in NO3 ? concentration in winter as gross primary productivity decreased. This study shows that the balance between autotrophy and heterotrophy in urban streams is variable and depends on an interplay of drivers such as temperature, light, and carbon inputs that are mediated by the riparian ecosystem. 相似文献
19.
Heterotrophic nitrification and aerobic denitrification by a novel Halomonas campisalis 总被引:1,自引:0,他引:1
Yan Guo Xuemei Zhou Yuguang Li Ke Li Caixia Wang Jianfei Liu Daojiang Yan Yilan Liu Dehui Yang Jianmin Xing 《Biotechnology letters》2013,35(12):2045-2049
A novel halophilic strain that could carry out heterotrophic nitrification and aerobic denitrification was isolated and named as Halomonas campisalis ha3. It removed inorganic nitrogen compounds (e.g. NO3 ?, NO2 ? and NH4 +) simultaneously, and grew well in the medium containing up to 20 % (w/v) NaCl. PCR revealed four genes in the genome of ha3 related to aerobic denitrification: napA, nirS, norB and nosZ. The optimal conditions for aerobic denitrification were pH 9.0, at 37 °C, with 4 % (w/v) NaCl and sodium succinate as carbon source. The nitrogen removal rate was 87.5 mg NO3 ?–N l?1 h?1. Therefore, this strain is a potential aerobic denitrifier for the treatment of saline wastewater. 相似文献
20.
A protocol for in vitro propagation of cineraria (Senecio cruentus) was developed. The highest frequency of shoot proliferation was obtained from nodal explants cultured on Murashige and Skoog (MS) medium supplemented with 2.0?mg L?1 6-benzyladenine (BA) and 0.5?mg L?1 ??-naphthalene acetic acid (NAA), with a mean number of 14 shoots per explant. A high concentration of BA (4.0?mg L?1) and repeated subcultures resulted in hyperhydric shoots. Decreasing the BA concentration to 1.0?mg L?1 in the culture medium eliminated hyperhydricity. The concentration of ammonium nitrate (NH4NO3) and temperature had marked effects on somaclonal variation. Variation was observed when the cultures were maintained at 15?°C but not at 25?°C. Variants with blue-colored leaves and stems were identified; whereas, normal plants maintained their green-colored leaves and stems. The highest frequency of variation (67.5?%), with a mean number of 3.0 variant shoots per explants, was obtained on shoot proliferation medium (MS?+?2.0?mg L?1 BA and 0.5?mg L?1 NAA) devoid of NH4NO3. The best rooting (100?%), with the highest number of roots per shoot (10.8) and the greatest root length (6.8?cm) was obtained on medium supplemented with 0.1?mg L?1 NAA. In vitro-grown plantlets were successfully acclimatized in a greenhouse, and transferred to the field. 相似文献