Chromium(VI) resistance and removal by <Emphasis Type="Italic">Acinetobacter haemolyticus</Emphasis>

The present work highlighted the studies on Cr(VI) reduction by cells of Acinetobacter haemolyticus (A. haemolyticus). The strain tolerated 90 mg Cr(VI) l⁻¹ in LB broth compared to only 30 mg Cr(VI) l⁻¹ in LB agar. From the FTIR analysis, the Cr(III) species formed was also most likely to form complexes with carboxyl, hydroxyl, and amide groups from the bacteria. A TEM study showed the absence of precipitates on the cell wall region of the bacteria. Instead, microprecipitates were observed in the cytoplasmic region of the cells, suggesting the transportation of Cr(VI) into the cells. Intracellular reduction of Cr(VI) was supported by a reductase test using soluble crude cell-free extracts. The specific reductase activity obtained was 0.52 μg Cr(VI) reduced per mg of protein an hour at pH 7.2 and 37°C. Our results indicated that A. haemolyticus can be used as a promising microorganism for Cr(VI) reduction from industrial wastewaters.     

Isolation and characterization of a chromium-resistant bacterium Serratia sp. Cr-10 from a chromate-contaminated site

A novel bacterium, Cr-10, was isolated from a chromium-contaminated site and capable of removing toxic chromium species from solution by reducing hexavalent chromium to an insoluble precipitate. Sequence analysis of 16S rRNA gene of strain Cr-10 showed that it was most closely related to Serratia rubidaea JCM 1240^T (97.68%). Physiological and chemotaxonomic data also supported that strain Cr-10 was identified as Serratia sp., a genus which was never specially reported chromate-resistant before. Serratia sp., Cr-10 was tolerant to a concentration of 1,500 mg Cr(VI) L⁻¹, which was the highest level reported until now. The optimum pH and temperature for reduction of Cr(VI) by Serratia sp. Cr-10 were found to be 7.0 and 37 °C, respectively. The Cr(VI) reduction was significantly influenced by additional carbon sources, and among them fructose and lactose offered maximum reduction, with a rate of 0.28 and 0.25 mg Cr(VI) L⁻¹ h⁻¹, respectively. The cell-free extracts and filtrate of the culture were able to reduce Cr(VI) while concentration of total chromium remained stable in the process, indicating that the enzyme-catalyzed mechanism was applied in Cr(VI) reduction by the isolate. Additionally, it was found that there was hardly any chromium on the cell surface of the strain, further supporting that reduction, rather than bioadsorption, plays a major role in the Cr(VI) removal.     

Chromate reduction by PVA-alginate immobilized <Emphasis Type="Italic">Streptomyces griseus</Emphasis> in a bioreactor

Microbial reduction of toxic Cr⁶⁺ to the less toxic Cr³⁺ is potentially a useful bioremediation process. Among the matrices tested for whole cell immobilization of an efficient chromate-reducing Streptomyces griseus strain, PVA-alginate was the most effective and was used for reduction of Cr(VI) in a bioreactor. Cr⁶⁺ reduction efficiency decreased as Cr⁶⁺ was increased from 2 to 12 mg l⁻¹ but increased with an increase in biomass concentration. However, increasing the flow rate from 2 to 8 ml h⁻¹ did not significantly affect Cr⁶⁺ reduction. The reduction was faster in simulated effluent than in synthetic medium and complete removal of 8 mg Cr⁶⁺ l⁻¹ from effluent and synthetic medium occurred in 2 and 12 h, respectively. Our results indicate that immobilized S. griseus cells could be applied for the large-scale bioremediation of chromate-containing effluents and wastewaters.     

Enhancement of hexavalent chromium reduction and electricity production from a biocathode microbial fuel cell

Enhancement of Cr (VI) reduction rate and power production from biocathode microbial fuel cells (MFCs) was achieved using indigenous bacteria from Cr (VI)-contaminated site as inoculum and MFC architecture with a relatively large cathode-specific surface area of 340–900 m² m⁻³. A specific Cr (VI) reduction rate of 2.4 ± 0.2 mg g⁻¹VSS h⁻¹ and a power production of 2.4 ± 0.1 W m⁻³ at a current density of 6.9 A m⁻³ were simultaneously achieved at an initial Cr (VI) concentration of 39.2 mg L⁻¹. Initial Cr (VI) concentration and solution conductivity affected Cr (VI) reduction rate, power production and coulombic efficiency. These findings demonstrate the importance of inoculation and MFC architecture in the enhancement of Cr (VI) reduction rate and power production. This study is a beneficial attempt to improve the efficiency of biocathode MFCs and provide a good candidate of bioremediation process for Cr (VI)-contaminated sites.     

Encapsulation of <Emphasis Type="Italic">Pannonibacter phragmitetus</Emphasis> LSSE-09 in alginate–carboxymethyl cellulose capsules for reduction of hexavalent chromium under alkaline conditions

Cr(VI) was efficiently reduced to Cr(III) by Pannonibacter phragmitetus LSSE-09 encapsulated in liquid-core alginate–carboxymethyl cellulose capsules under alkaline conditions. Taking into account the physical properties of the capsules, the activity of encapsulated cells, and total Cr(III) concentration in the supernatant, optimal conditions (0.5% w/v sodium alginate; 2% w/v sodium carboxymethyl cellulose; 0.1 M CaCl₂; 30-min gelation time) for LSSE-09 encapsulation were determined. At optimal conditions, a relatively high reduction rate of 4.20 mg g_{(dry weight)}⁻¹  min⁻¹ was obtained. Total Cr(III) concentration in the supernatant was significantly decreased after reduction, because 63.7% of the formed soluble organo-Cr(III) compounds compared with those of free cells were captured by the relatively smaller porous structure of alginate capsules. The optimal pH value (9.0) for Cr(VI) reduction was not changed after encapsulation. In addition, encapsulated LSSE-09 showed no appreciable loss in activity after eight repeated cycles at 37°C, and 85.7% of its initial activity remained after 35-day storage at 4°C. The results suggest that encapsulated LSSE-09 in alginate–carboxymethyl cellulose capsules has potential biotechnological applications for the detoxification of Cr(VI)-contaminated wastewater.     

Bioreduction of Cu(II) by Cell-Free Copper Reductase from a Copper Resistant <Emphasis Type="Italic">Pseudomonas</Emphasis> sp. NA

Environmental copper contamination is a serious human health problem. Copper reductase is produced by microorganisms to facilitate copper uptake by ATPases into the cells increasing copper biosorption. This study assessed the reduction of Cu(II) by cell-free extracts of a highly copper-resistant bacterium, Pseudomonas sp. strain NA, isolated from vineyard soil contaminated with copper. Both intact cells and cell-free extract of Pseudomonas sp. strain NA displayed substantial reduction of Cu(II). Intact cells reduced more then 80 mg L⁻¹ of Cu(II) from medium amended with 200 mg L⁻¹ of copper after 24 h of incubation. Cell-free extract of the isolate reduced more than 65% of the Cu(II) at initial copper concentration of 200 mg L⁻¹ after 24 h. Soluble protein production was high at 72 h of incubation at 100 mg L⁻¹ of copper, with more then 60 μg L⁻¹ of total soluble protein in cell-free extract recorded. Cu(II) reduction by isolate NA was increased when copper concentration increased for both intact cells and cell-free extract. Results indicate that Pseudomonas sp. strain NA produces copper reductase enzyme as the key mechanism of copper biotransformation.     

Efficacy of bacterial consortium-AIE2 for contemporaneous Cr(VI) and azo dye bioremediation in batch and continuous bioreactor systems, monitoring steady-state bacterial dynamics using qPCR assays

Bacterial consortium-AIE2 with a capability of contemporaneous Cr(VI) reduction and azo dye RV5 decolourization was developed from industrial wastewaters by enrichment culture technique. The 16S rRNA gene based molecular analyses revealed that the consortium bacterial community structure consisted of four bacterial strains namely, Alcaligenes sp. DMA, Bacillus sp. DMB, Stenotrophomonas sp. DMS and Enterococcus sp. DME. Cumulative mechanism of Cr(VI) reduction by the consortium was determined using in vitro Cr(VI) reduction assays. Similarly, the complete degradation of Reactive Violet 5 (RV5) dye was confirmed by FTIR spectroscopic analysis. Consortium-AIE2 exhibited simultaneous bioremediation efficiencies of (97.8 ± 1.4) % and (74.1 ± 1.2) % in treatment of both 50 mg l⁻¹ Cr(VI) and RV5 dye concentrations within 48 h of incubation at pH 7 and 37°C in batch systems. Continuous bioreactor systems achieved simultaneous bioremediation efficiencies of (98.4 ± 1.5) % and (97.5 ± 1.4) % after the onset of steady-state at 50 mg l⁻¹ input Cr(VI) and 25 mg l⁻¹ input RV5 concentrations, respectively, at medium dilution rate (D) of 0.014 h⁻¹. The 16S rRNA gene copy numbers in the continuous bioreactor as determined by real-time PCR assay indicated that Alcaligenes sp. DMA and Bacillus sp. DMB dominated consortium bacterial community during the active continuous bioremediation process.     

Combination effect of lactoalbumin hydrolysate and methyl jasmonate on ginsenoside and polysaccharide production in <Emphasis Type="Italic">Panax quinquefolium</Emphasis> L. cells cultures

Lactoalbumin hydrolysate (LH) at 100 mg L⁻¹ with methyl jasmonate (MJ) at 2 mg L⁻¹ synergistically stimulated ginsenoside accumulation in Panax quinquefolium cells compared with 100 mg L⁻¹ LH. Combination elicitors led to higher ginsenoside productivity (45.93 mg L⁻¹) than single treatment of 100 mg L⁻¹ LH (31.37 mg L⁻¹). This present result will be helpful in providing a tool for enhancing the productivity of ginsenoside by Panax quinquefolium cell cultures on a commercial scale.     

A bioprocessing strategy that allows for the selection of Cr(VI)-reducing bacteria from soils

Anaerobic bacteria that reduce hexavalent chromium [Cr(VI)] to trivalent [Cr(III)] are common in soils and were used to develop a bioprocess employing a selection strategy. Indigenous Cr(VI)-reducers were enriched from Cr(VI)-contaminated soil under anaerobic conditions. The mixed culture was then tested for Cr(VI)-reducing activity in a chemostat, followed by transfer to a 1-L packed-bed bioreactor operated at 30°C for additional study. The support material used in the reactor consisted of 6-mm porcelain saddles. Cr(VI) concentrations in the liquid ranged from 140–750 mg L⁻¹. Cr(VI)-reducing bacteria were the dominant population with Cr(VI)-reduction rates of approximately 0.71 mg g⁻¹ dry cells h⁻¹ achieved at Cr(VI) concentrations of 750 mg L⁻¹. These results indicate a potential for selecting and maintaining indigenous Cr(VI)-reducers in a bioreactor for Cr(VI)-remediation of groundwater or soil wash effluents. Received 09 January 1996/ Accepted in revised form 15 November 1996     

Bioremoval of hexavalent chromium from water by a salt tolerant bacterium, Exiguobacterium sp. GS1

Pollution of terrestrial surfaces and aquatic systems by hexavalent chromium, Cr(VI), is a worldwide public health problem. A chromium resistant bacterial isolate identified as Exiguobacterium sp. GS1 by 16S rRNA gene sequencing displayed high rate of removal of Cr(VI) from water. Exiguobacterium sp. GS1 is 99% identical to Exiguobacterium acetylicum. The isolate significantly removed Cr(VI) at both high and low concentrations (1–200 μg mL⁻¹) within 12 h. The Michaelis–Menten K _m and V _max for Cr(VI) bioremoval were calculated to be 141.92 μg mL⁻¹ and 13.22 μg mL⁻¹ h⁻¹, respectively. Growth of Exiguobacterium sp. GS1 was indifferent at 1–75 μg mL⁻¹ Cr(VI) in 12 h. At initial concentration of 8,000 μg L⁻¹, Exiguobacterium sp. GS1 displayed rapid bioremoval of Cr(VI) with over 50% bioremoval in 3 h and 91% bioremoval in 8 h. Kinetic analysis of Cr(VI) bioremoval rate revealed zero-order in 8 h. Exiguobacterium sp. GS1 grew and significantly reduced Cr(VI) in cultures containing 1–9% salt indicating high salt tolerance. Similarly the isolate substantially reduced Cr(VI) over a wide range of temperature (18–45  °C) and initial pH (6.0–9.0). The T _opt and initial pH_opt were 35–40  °C and 7–8, respectively. Exiguobacterium sp. GS1 displayed a great potential for bioremediation of Cr(VI) in diverse complex environments.     

Evaluation of chromium(VI) removal behaviour by two isolates of Synechocystis sp. in terms of exopolysaccharide (EPS) production and monomer composition

Chromium(VI) removal and its association with exopolysaccharide (EPS) production in cyanobacteria were investigated. Synechocystis sp. BASO670 produced higher EPS (548 mg L⁻¹) than Synechocystis sp. BASO672 (356 mg L⁻¹). While the EC₅₀ of the Cr(VI) for Synechocystis sp. BASO670 and Synechocystis sp. BASO672 were determined as 11.5 mg L⁻¹, and 2.0 mg L⁻¹, respectively, there was no relation between Cr(VI) removal and EPS production. Synechocystis sp. BASO672, which has higher EPS value, removed (33%) more Cr(VI) than Synechocystis sp. BASO670. Monomer compositions of EPS of each of the isolates were determined differently. Synechocystis sp. BASO672 which removed higher Cr(VI), had higher values of uronic acid and glucuronic acid (192 μg/mg and 89%, respectively). Our results showed that EPS might play a role in Cr(VI) tolerance. Monomer composition, especially uronic acid and glucuronic acid content of EPS may have enhanced Cr(VI) removal.     

Anaerobic co-reduction of chromate and nitrate by bacterial cultures of Staphylococcus epidermidis L-02

Industrial wastewater is often polluted by Cr(VI) compounds, presenting a serious environmental problem. This study addresses the removal of toxic, mutagenic Cr(VI) by means of microbial reduction to Cr(III), which can then be precipitated as oxides or hydroxides and extracted from the aquatic system. A strain of Staphylococcus epidermidis L-02 was isolated from a bacterial consortium used for the remediation of a chromate-contaminated constructed wetland system. This strain reduced Cr(VI) by using pyruvate as an electron donor under anaerobic conditions. The aims of the present study were to investigate the specific rate of Cr(VI) reduction by the strain L-02, the effects of chromate and nitrate (available as electron acceptors) on the strain, and the interference of chromate and nitrate reduction processes. The presence of Cr(VI) decreased the growth rate of the bacterium. Chromate and nitrate reduction did not occur under sterile conditions but was observed during tests with the strain L-02. The presence of nitrate increased both the specific Cr(VI) reduction rate and the cell number. Under denitrifying conditions, Cr(VI) reduction was not inhibited by nitrite, which was produced during nitrate reduction. The average specific rate of chromate reduction reached 4.4 μmol Cr 10¹⁰ cells⁻¹ h⁻¹, but was only 2.0 μmol Cr 10¹⁰ cells⁻¹ h⁻¹ at 20 °C. The maximum specific rate was as high as 8.8–9.8 μmol Cr 10¹⁰ cells⁻¹ h⁻¹. The role of nitrate in chromate reduction is discussed.     

In vitro hormone-regulated growth and floral induction of <Emphasis Type="Italic">Cuscuta reflexa</Emphasis>: a parasitic angiosperm

The role of different growth regulators in callus induction, shoot regeneration, floral induction and chlorophyll content of the obligatory parasitic plant Cuscuta reflexa has been studied. Callus development was excellent from the nodal part of the shoot explants in modified Murashige and Skoog (MMS) media supplemented with 2 mg L⁻¹ benzyl adenine (MMS1c). Supplementation of 2 mg L⁻¹ naphthalene acetic acid (NAA) along with MMS1c (MMS2c) was responsible for estimable shoot induction and development in callus. 2,4-Dichloro acetic acid (2,4-D) played a crucial role in the floral induction of C. reflexa in vitro. MMS supplemented with 2 mg L⁻¹ NAA and 2 mg L⁻¹ 2,4-D (MMS3b) supported floral induction after shooting in vitro. MMS supplemented with 3 mg L⁻¹ 2,4-D (MMS4a) rapidly induced flower directly from the stem explants without showing any elongation of shoot. MMS1c along with MMS3b (MMS5a) showed callus proliferation followed by shoot elongation and floral induction. In vitro MMS5a grown plants show a sharp increase in the chlorophyll contents. Cytokinin treatment further increases the chlorophyll level of the plant.     

Benzyladenine Promotes Flowering in <Emphasis Type="Italic">Doritaenopsis</Emphasis> and <Emphasis Type="Italic">Phalaenopsis</Emphasis> Orchids

Two experiments were performed to determine how application of the cytokinin benzyladenine (BA) influenced flowering in Doritaenopsis and Phalaenopsis orchid clones. In the first experiment, two vegetative orchid clones growing in 15-cm pots were transferred from a 28°C greenhouse that inhibited flowering to a 23°C greenhouse for flower induction (day 0). A foliar spray (0.2 L m⁻²) containing BA at 100, 200, or 400 mg L⁻¹ or 25, 50, or 100 mg L⁻¹ each of BA and gibberellins A₄ + A₇ (BA+GA) was applied on days 0, 7, and 14. Plants treated with BA alone at 200 or 400 mg L⁻¹ had a visible inflorescence 3–9 days earlier and had a mean of 0.7–3.5 more inflorescences and 3–8 more flowers per plant than nontreated plants. The application of BA+GA had no effect on inflorescence number and total flower number at the rates tested. In the second experiment, three orchid clones received a single foliar spray of BA at 200 mg L⁻¹ at six time points relative to time of transfer from 29°C to 23°C (−1, 0, +1, +2, +4, or +6 weeks). A separate group of plants received a BA application at week 0 but was maintained at 29°C. Inflorescence number was greatest in all three orchid clones when plants were treated with BA 1 week after the temperature transfer. Plants that were sprayed with BA and maintained at 29°C did not initiate inflorescences. The promotion of flowering by the application of BA suggests that cytokinins at least partially regulate inflorescence initiation of Doritaenopsis and Phalaenopsis, but its promotion is conditional and BA application cannot completely substitute for an inductive low temperature.     

Evaluation of the bioremoval of Cr(VI) and TOC in biofilters under continuous operation using response surface methodology

In the present study, the bioremoval of Cr(VI) and the removal of total organic carbon (TOC) were achieved with a system composed by an anaerobic filter and a submerged biofilter with intermittent aeration using a mixed culture of microorganisms originating from contaminated sludge. In the aforementioned biofilters, the concentrations of chromium, carbon, and nitrogen were optimized according to response surface methodology. The initial concentration of Cr(VI) was 137.35 mg l⁻¹, and a bioremoval of 85.23% was attained. The optimal conditions for the removal of TOC were 4 to 8 g l⁻¹ of sodium acetate, >0.8 g l⁻¹ of ammonium chloride and 60 to 100 mg l⁻¹ of Cr(VI). The results revealed that ammonium chloride had the strongest effect on the TOC removal, and 120 mg l⁻¹ of Cr(VI) could be removed after 156 h of operation. Moreover, 100% of the Cr(VI) and the total chromium content of the aerobic reactor output were removed, and TOC removals of 80 and 87% were attained after operating the anaerobic and aerobic reactors for 130 and 142 h, respectively. The concentrations of cells in both reactors remained nearly constant over time. The residence time distribution was obtained to evaluate the flow through the bioreactors.     

Rhizoremediation of diesel-contaminated soil using the plant growth-promoting rhizobacterium <Emphasis Type="Italic">Gordonia</Emphasis> sp. S2RP-17

A plant growth-promoting rhizobacterium (PGPR) was isolated and identified as Gordonia sp. S2RP-17, which showed ACC deaminase and siderophore synthesizing activities. Its maximum specific growth rate was 0.54 ± 0.12 d⁻¹ at 5,000 mg L⁻¹ of total petroleum hydrocarbon (TPH), and its maximum diesel degradation rate was 2,434.0 ± 124.4 mg L⁻¹ d⁻¹ at 20,000 mg L⁻¹ of TPH. The growth of Zea mays was significantly promoted by the inoculation of Gordonia sp. S2RP-17 in the diesel-contaminated soil. Measured TPH removal efficiencies by various means were 13% by natural attenuation, 84.5% by planting Zea mays, and 95.8% by the combination of Zea mays and Gordonia sp. S2RP-17. The S2RP-17 cell counts were maintained at 1 × 10⁶ CFU g-soil⁻¹ during the remediation period, although they slightly decreased from their initial numbers (2.94 × 10⁷ CFU g-soil⁻¹). These results indicate that rhizoremediation using both Zea mays and Gordonia sp. S2RP-17 is a promising strategy for enhancing remediation efficiency of diesel-contaminated soils.     

Chromate reduction by a chromate-resistant bacterium, <Emphasis Type="Italic">Microbacterium</Emphasis> sp.

Heavy-metal chromium [Cr(VI)] is a ubiquitous environmental pollutant. Comparing with chemical reduction, microbiological reduction is considered to be a friendly and cheaper way to decrease the damage caused by chromate. A bacterial strain, CR-07, which is resistant to and capable of reducing chromate was isolated from a mud sample of iron ore and identified as a Microbacterium sp. The bacterium had a high degree of tolerance to chromate, and could grow in LB medium containing 4.08 mM of K₂Cr₂O₇. It also had a degree of resistance to other heavy metals, e.g. Cd²⁺, Pb²⁺, Zn²⁺, Cu²⁺, Co²⁺, Hg²⁺ and Ag⁺. The bacterium could remove 1.02 mM of Cr(VI) from LB medium within 36 h of incubation. Chromate removal was achieved in the supernatant from the bacterial cultures, and corresponded to chromate reduction. The activity of chromate reduction by the bacterium was not related to enzymes or reducing sugars, while fluorometric assay suggested that glutathione, a chromate-reducing substance which was produced by the bacterium, was one of the factors that contributed to the reduction of Cr(VI).     

Evaluation of green fluorescent protein as a reporter gene and phosphinothricin as the selective agent for achieving a higher recovery of transformants in cucumber (<Emphasis Type="Italic">Cucumis sativus</Emphasis> L. cv. Poinsett76) via <Emphasis Type="Italic">Agrobacterium tumefaciens</Emphasis>

Efficient Agrobacterium tumefaciens-mediated transformation and a higher recovery of transformed plants of cucumber cv. Poinsett76 were achieved via direct organogenesis from cotyledon explants. Stable transformants were obtained by inoculating explants with A. tumefaciens strains EHA105 or LBA4404, both harboring the binary vector pME508, which contains the neomycin phosphotransferase II (nptII) and phosphinothricin resistance genes (bar) conferring resistance to kanamycin and PPT, respectively, as selectable markers and the sgfp-tyg gene for the green fluorescent protein (GFP) as a visual marker driven by the constitutive CaMV35S promoter in the presence of acetosyringone (50 μM). Transformed shoots were obtained on MS Murashige and Skoog (Plant Physiol. 15: 473–497, 1962) medium supplemented with 1 mg L⁻¹ benzyladenine (BA), 20 mg L⁻¹ l-glutamine and 2 mg L⁻¹ phosphinothricin (PPT) or 100 mg L⁻¹ kanamycin. The regenerated shoots were examined in vivo using a hand-held long wave UV lamp for GFP expression. The GFP screening helped identify escapes and chimeric shoots at regular intervals to increase the growth of transformed shoots on cotyledon explants. Elongation and rooting of putative transformants were achieved on PPT (2 mg L⁻¹) containing MS media with 0.5 mg L⁻¹ gibberellic acid (GA₃) and 0.6 mg L⁻¹ indole butyric acid (IBA), respectively. PCR and Southern analyses confirmed the integration of the sgfp gene into the genome of T₀ and the progenies. T₁ segregation of transgenic progeny exhibited Mendelian inheritance of the transgene. The use of EHA105 resulted in 21% transformation efficiency compared to 8.5% when LBA4404 was used. This higher rate was greatly facilitated by PPT selection coupled with effective screening of transformants for GFP expression, thus making the protocol highly useful for the recovery of a higher number of transgenic cucumber plants.     

Production of saponions and polysaccharide in the presence of lactoalbumin hydrolysate in <Emphasis Type="Italic">Panax quinquefolium</Emphasis> L. cells cultures

In this article, ginsenosides and polysaccharide contents in suspension cells and native roots of Panax quinquefolium L. were studied. In order to enhance the contents of ginsenosides and polysaccharide in P. quinquefolium suspension cells, we tested the effects of lactoalbumin hydrolysate on the growth of P. quinquefolium suspension cell, synthesis of ginsenosides and polysaccharide in flask and bioreactor. In flask culture, cells growth ratio was significantly enhanced by the addition of lower concentration of lactoalbumin hydrolysate. Addition of 100 mg L⁻¹ lactoalbumin hydrolysate significantly enhanced the contents of total saponins (5.44 mg g⁻¹ DW) and the contents were 3.89-fold over the control group. Addition of lactoalbumin hydrolysate significantly promoted the accumulation of polysaccharide, except 200 mg L⁻¹ lactoalbumin hydrolysate. The highest total saponins yield (36.72 mg L⁻¹ DW) and polysaccharide yield (0.83 g L⁻¹ DW) were obtained at 100 mg L⁻¹ lactoalbumin hydrolysate. In a 5-L stirred tank bioreactor, the highest contents of total saponins and TRb group ginsenosides were achieved on day 26, while the effect of lactoalbumin hydrolysate on the contents of TRg group ginsenosides were insignificant. This result suggests that lactoalbumin hydrolysate might have triggered the enzyme activities for the synthesis of TRb group ginsenosides. Overall, the highest total saponins yield (31.37 mg L⁻¹ DW) and polysaccharide yield (1.618 g L⁻¹ DW) were obtained on day 26 and day 24 respectively and the polysaccharide yield was 1.95-fold higher than the shake flask culture (0.83 g L⁻¹ DW). These results provided theoretical reference for two-stage culture in suspension cells of P. quinquefolium in bioreactor.     

Effects of Artemisinin Derivative on the Growth Metabolism of <Emphasis Type="Italic">Tetrahymena thermophila</Emphasis> BF5 Based on Expression of Thermokinetics

The toxic effects of artesunate and dihydroartemisinin on the growth metabolism of Tetrahymena thermophila BF5 were studied by microcalorimetry. The results showed that: (1) low concentrations of artesunate (≤1 mg L⁻¹) and dihydroartemisinin (≤ 2 mg L⁻¹) promoted the growth metabolism of T. thermophila BF5, whereas high concentrations of artesunate (1–60 mg L⁻¹) and dihydroartemisinin (2–60 mg L⁻¹) inhibited its growth; (2) the half inhibition concentrations IC₅₀ of artesunate and dihydroartemisinin were 17.5817 and 9.5089 mg L⁻¹, respectively. It was concluded that the inhibition of dihydroartemisinin was stronger than that of artesunate.