Ascorbate-Glutathione Cycle and Ultrastructural Analyses of Two Kenaf Cultivars (Hibiscus cannabinus L.) under Chromium Stress.

Kenaf (Hibiscus cannabinus L.) with high tolerance to chromium (Cr) can be used in the phytoremediation of chromium-contaminated soil. However, the mechanisms of chromium accumulation and tolerance in kenaf are still unclear. A hydroponic experiment was taken to screen two kenaf cultivars with Cr tolerance among nine kenaf cultivars via a tolerance index. This is first time the ascorbate-glutathione (AsA-GSH) cycle and chloroplast structural changes involved in Cr tolerance of two kenaf cultivars are explored. This study indicated that enhancement of chromium concentrations reduced nine kenaf growth rates and plant biomass. In addition, in all the nine cultivars, the roots had higher Cr accumulation than the shoots. Cr-tolerant cultivar Zhe70-3 with the maximum tolerant index had the significantly higher enzymatic activities of ascorbate peroxidase (APX), glutathione reductase (GR), dehydroascorbate reductase (DHAR) and mono- dehydroascorbate reductase (MDHAR) in non-enzymatic antioxidant system compared to Cr-sensitive cultivar Zhe77-1. In addition, higher GSH and AsA contents and lower damages of chloroplast ultrastructure were observed in Zhe70-3 under Cr treatment. In conclusion, Cr stress can cause less oxidative stress and destruction of chloroplast ultrastructure in Cr-tolerant cultivar Zhe70-3, and the AsA-GSH cycle may play a crucial role in kenaf Cr tolerance.

Physicochemical Properties and Oxidative Storage Stability of Milled Roselle (Hibiscus sabdariffa L.) Seeds.

Milled Roselle (Hibiscus sabdariffa L.) seeds of the UMKL cultivar were analyzed for proximate composition, water and oil absorption capacity, and the influence of storage conditions on storage stability. The storage stability was determined under four types of conditions: light/oxygen (air) (LO), light/nitrogen (LN), darkness/oxygen (air) (DO), and darkness/nitrogen (DN) while monitoring for seven consecutive months. During the storage period, the formation of volatiles was determined using dynamic headspace sampling and Gas Chromatography-Mass Spectrometry (GC-MS) analysis. In total, 85 volatiles were identified, mainly aldehydes, alcohols, ketones, furans, and acids indicating lipid oxidation. It is recommended that milled Roselle seeds should be flushed with nitrogen and stored in darkness. Under these conditions, the seeds can be stored for at least three months without changes in volatile profile. This is important to ensure the good quality of milled Roselle seeds for further commercialization.

Hibiscus rosa sinensis mediate anxiolytic effect via modulation of ionotropic GABA-A receptors: possible mechanism of action.

The current study was designed with the aim to investigate anti-anxiety potential of Hibiscus rosa sinensis roots and its possible mechanism of action. For this purpose hole board test, elevated plus maze test and light/dark exploration test were employed. The ethanol extract of plant was administered orally at two different doses i.e. 100 and 500 mg/kg for consecutive 14 days. The results of present investigation indicate that plant extract significantly (p < 0.05) increased the number of head dips and rearings as compared to control on respective days of observation. The extract increased the time of permanence in open arms and the number of head dips in elevated plus maze. In light/dark test, our study indicate that Hibiscus rosa sinensis significantly (p < 0.05) increased the time spent in light compartment and number of entries as compared to control. In addition the anxiolytic effects of HRS at highest tested dose were blocked by flumazenil, a GABA-A receptor antagonist that indicate that Hibiscus rosa sinensis potentiated the GABAergic actions. The results propose that the ethanol extract of Hibiscus rosa sinensis has prospective anxiolytic effect in mice via inhibition of ionotropic GABA receptors, using different behavioral paradigms.

Physico-chemical changes in karkade (Hibiscus sabdariffa L.) seedlings responding to salt stress.

Salinity is one of the major abiotic stress factors affecting series of morphological, physiological, metabolic and molecular changes in plant growth. The effect of different concentrations (0, 25, 50, 100 and 150 mM) of NaCl on the vegetative growth and some physiological parameters of karkade (Hibiscus sabdariffa var. sabdariffa) seedling were investigated. NaCl affected the germination rate, delayed emergence and retarded vegetative growth of seedlings. The length of seedling as well as the leaf area was significantly reduced. The fresh weight remained lower in NaCl treated seedlings compared to control. NaCl at 100 and 150 mM concentrations had significant effect on the dry matter contents of the treated seedlings. The chloroplast pigments in the treated seedlings were affected, suggesting that the NaCl had a significant effect on the chlorophyll and carotenoid biosynthesis. The results showed that the salt treatments induced an increase in proline concentration of the seedlings. The osmotic potential (ψs) of NaCl treated seedlings decreased with increasing NaCl concentrations. Salt treatments resulted in dramatic quantitative reduction in the total sterol percent compared with control ones. Salt stress resulted in increase and decrease of Na+ and K+ ions, respectively. NaCl salinity increased lipid peroxidation. SDS-PAGE was used to evaluate protein pattern after applying salt stress. High molecular weight proteins were intensified, while low molecular weight proteins were faint. NaCl at 100 and 150 mM concentration distinguished with new protein bands. Salt stress induced a new peroxidase bands and increased the band intensity, indicating the protective role of peroxidase enzyme.

Physiological responses and tolerance of kenaf (Hibiscus cannabinus L.) exposed to chromium.

Selection of kenaf species with chromium (Cr) tolerance and exploring the physiological mechanisms involved in Cr tolerance are crucial for application of these species to phyto-remediation. In the present study, a hydroponic experiment was conducted to investigate the variation in two kenaf cultivars, K39-2 and Zhe50-3 under Cr stress. At the same Cr concentration, the tolerance index (TI) of K39-2 was higher than that of Zhe50-3, indicating that K39-2 may be more tolerant to Cr than Zhe50-3. It was also observed that high concentration of chromium was accumulated both in the shoots and the roots of Hibiscus cannabinus L. The leaves of K39-2 accumulated 4760.28mgkg(-1) of dry weight under 1.50mM Cr stress, and the roots accumulated 11,958.33mgkg(-1). Physiological response shows that the antioxidant enzymes' superoxide dismutase (SOD), catalase activity (CAT) and peroxidase (POD) activities increased in the leaves and decreased in roots of the Cr-stressed plants nearly compared to the control. Moreover, the variation of antioxidant enzymes activities indicated Zhe50-3 was more vulnerable than K39-2, and the contents of the non-protein thiol pool (GSH, NPT and PCs) were higher in K39-2 than Zhe50-3 with the increased Cr concentration. Based on the observations above, it can be concluded that the well-coordinated physiological changes confer a greater Cr tolerance to K39-2 than Zhe50-3 under Cr exposure, and Hibiscus cannabinus L. has a great accumulation capacity for chromium.

Short-term regulation and alternative pathways of photosynthetic electron transport in Hibiscus rosa-sinensis leaves.

In this work, using the EPR and PAM-fluorometry methods, we have studied induction events of photosynthetic electron transport in Hibiscus rosa-sinensis leaves. The methods used are complementary, providing efficient tools for in situ monitoring of P700 redox transients and photochemical activity of photosystem II (PSII). The induction of P700(+) in dark-adapted leaves is characterized by the multiphase kinetics with a lag-phase, which duration elongates with the dark-adaptation time. Analyzing effects of the uncoupler monensin and artificial electron carrier methylviologen (MV) on photooxidation of P700 and slow induction of chlorophyll a fluorescence (SIF), we could ascribe different phases of transient kinetics of electron transport processes in dark-adapted leaves to the following regulatory mechanisms: (i) acceleration of electron transfer on the acceptor side of PSI, (ii) pH-dependent modulation of the intersystem electron flow, and (iii) re-distribution of electron fluxes between alternative (linear, cyclic, and pseudocyclic) pathways. Monensin significantly decreases a level of P700(+) and inhibits SIF. MV, which mediates electron flow from PSI to O2 with consequent formation of H2O2, promotes a rapid photooxidation of P700 without any lag-phase peculiar to untreated leaves. MV-mediated water-water cycle (H2O→PSII→PSI→MV→O2→H2O2→H2O) is accompanied by generation of ascorbate free radicals. This suggests that the ascorbate peroxidase system of defense against reactive oxygen species is active in chloroplasts of H. rosa-sinensis leaves. In DCMU-treated chloroplasts with inhibited PSII, the contribution of cyclic electron flow is insignificant as compared to linear electron flow. For analysis of induction events, we have simulated electron transport processes within the framework of our generalized mathematical model of oxygenic photosynthesis, which takes into account pH-dependent mechanisms of electron transport control and re-distribution of electron fluxes between alternative pathways. The model adequately describes the main peculiarities of P700(+) induction and dynamics of the intersystem electron transport.

Physicochemical and microbiological characterization of cement kiln dust for potential reuse in wastewater treatment.

Cement kiln dust (CKD), a byproduct of cement manufacturing process, was collected from Misr Cement Co. at Qena, Egypt. CKD was characterized by X-ray diffraction and FTIR analysis. This byproduct was investigated for its physical-chemical characters, antibacterial activities on sewage water and the presence of nematode, parasites and algae in the treated water. The efficiency of CKD-treated water was also examined on Hibiscus sabdarriffa seed germination. Total bacteria, total and fecal coliform, as well as fecal streptococci were completely inhibited by CKD. Interestingly, zinc, manganese, iron, nickel and lead were completely absent from sewage water as these metals precipitated after treatment with 10gl(-1) CKD. On the other hand, among all the tested plant criteria, only root length was significantly reduced by 55% and 15% after zero and 3 days of CKD addition respectively compared to control. Furthermore, plant lipid peroxidation showed no significant differences between treated sewage water and control after zero and 3 days time addition of CKD. Catalase enzyme activity showed significant decrease by 56% and 64%, while peroxidase activity significantly increased up to 49% and 63% compared to untreated sewage after zero and 3 days of treatment, respectively. The absorption of lead, iron and copper by treated and untreated plants showed no significant differences. Chromium ions were highly absorbed (0.075mgl(-1)) by plants irrigated only with treated sewage water at zero time, and decreased gradually to 0.018mgl(-1) after 3 days of CKD addition. This study highlighted the efficiency of cement kiln dust as an antibacterial agent and its ability of scavenging heavy metals leading to the use of treated sewage water in activities such as crop irrigation.

Determination of suitable microspore stage and callus induction from anthers of kenaf (Hibiscus cannabinus L.).

Kenaf (Hibiscus cannabinus L.) is one of the important species of Hibiscus cultivated for fiber. Availability of homozygous parent lines is prerequisite to the use of the heterosis effect reproducible in hybrid breeding. The production of haploid plants by anther culture followed by chromosome doubling can be achieved in short period compared with inbred lines by conventional method that requires self pollination of parent material. In this research, the effects of the microspore developmental stage, time of flower collection, various pretreatments, different combinations of hormones, and culture condition on anther culture of KB6 variety of Kenaf were studied. Young flower buds with immature anthers at the appropriate stage of microspore development were sterilized and the anthers were carefully dissected from the flower buds and subjected to various pretreatments and different combinations of hormones like NAA, 2,4-D, Kinetin, BAP, and TDZ to induce callus. The best microspore development stage of the flower buds was about 6-8 mm long collected 1-2 weeks after flower initiation. At that stage, the microspores were at the uninucleate stage which was suitable for culture. The best callus induction frequency was 90% in the optimized semisolid MS medium fortified with 3.0 mg/L BAP + 3.0 mg/L NAA.

Effect of zinc on nectar secretion of Hibiscus rosa -sinensis L.

Zinc toxicity in secretory cells caused a range of effects, mainly depending on metal concentration. Low concentrations activated nectary function increasing nectar secretion but secretion was greatly inhibited or stopped entirely by ongoing concentration. Water loss rate of zinc treated flower parts was significantly reduced whereas green sepals were dehydrated more rapidly in comparison to colored petals. The content of zinc, calcium, magnesium and manganese increased mainly in sepals under excess of zinc, but in the secreted nectar this metal was not evident. Morphological changes were observed in mucilage cells concerning the mucilage structure and appearance. The parenchymatic, subglandular cells displayed an early vacuolarization and cytoplasm condensation. Secretory hairs appeared to be thinner, the apical cell folded inwards and plasmolytic shrinkage became severe in all cells. The waxy cuticula showed an increased electron density. A plasmalemma detachment from the external cell walls was observed creating a gap between cell wall and plasmalemma. ER cisterns of all treated nectary hairs dominated the cytoplasm and electron dense deposits were seen within its profiles. A great number of other organelles were also present, showing electron dense deposits in their membranes as well. The vacuome was drastically reduced in all cells, except in the subglandular ones and electron dense membrane remnants were observed.

An improved TCF sequence for biobleaching kenaf pulp: influence of the hexenuronic acid content and the use of xylanase.

Enzymatic delignification with laccase from Trametes villosa used in combination with chemical mediators (acetosyringone, acetovanillone and 1-hydroxybenzotriazole) to improve the totally chlorine-free (TCF) bleaching of kenaf pulp was studied. The best final pulp properties were obtained by using an LHBTQPo sequence developed by incorporating a laccase-mediator stage into an industrial bleaching sequence involving chelation and peroxide stages. The new sequence resulted in increased kenaf pulp delignification (90.4%) and brightness (77.2%ISO) relative to a conventional TCF chemical sequence (74.5% delignification and 74.5% brightness). Also, the sequence provided bleached kenaf fibers with high cellulose content (pulp viscosity of 890 g·mL(-1) vs 660 g·mL(-1)). Scanning electron micrographs revealed that xylanase altered fiber surfaces and facilitated reagent access as a result. However, the LHBTX (xylanase) stage removed 21% of hexenuronic acids in kenaf pulp. These recalcitrant compounds spent additional bleaching reagents and affected pulp properties after peroxide stage.

Phytoextraction of As and Fe using Hibiscus cannabinus L. from soil polluted with landfill leachate.

Terrestrial plants as potential phytoremediators for remediation of surface soil contaminated with toxic metals have gained attention in clean-up technologies. The potential of kenaf (Hibiscus cannabinus L.) to offer a cost-effective mechanism to remediate Fe and As from landfill leachate-contaminated soil was investigated. Pot experiment employing soil polluted with treatments of Jeram landfill leachate was conducted for 120 days. Plants were harvested after 8th, 12th, and 16th weeks of growth. Accumulation of Fe and As was assessed based on Bioconcentration Factor and Translocation Factor. Results showed sequestration of 0.06-0.58 mg As and 66.82-461.71 mg Fe per g plant dry weight in kenaf root, which implies that kenaf root can be an bioavailable sink for toxic metals. Insignificant amount of Fe and As was observed in the aerial plant parts (< 12% of total bioavailable metals). The ability of kenaf to tolerate these metals and avoid phytotoxicity could be attributed to the stabilization of the metals in the roots and hence reduction of toxic metal mobility (TF < 1). With the application of leachate, kenaf was also found to have higher biomass and subsequently recorded 11% higher bioaccumulation capacity, indicating its suitability for phytoextraction of leachate contaminated sites.

Investigating the structure-effect relationships of various natural phenols used as laccase mediators in the biobleaching of kenaf and sisal pulps.

Nine phenol derivatives, p-coumaric acid (PC), vanillin (V), acetovanillone (AV), acetosyringone (AS), syringaldehyde (SA), coniferaldehyde (CLD), ferulic acid (FRC), sinapic acid (SNC), and sinapyl aldehyde (SLD) were assayed as laccase redox mediators in the biobleaching of kenaf and sisal pulps. As a general behaviour, the phenolic mediators increased the kappa number (KN) and reduced the brightness of pulps. In particular, these changes were found to depend in a linear manner on the energy of the highest occupied molecular orbital (E(HOMO)) of the mediators. The phenolic mediator with the lowest E(HOMO) (PC) led to the highest increase of KN and the lowest reduction of brightness. On the contrary, syringyl derivatives (i.e. SA) with high E(HOMO) values caused small KN increases and significant losses of brightness. This behaviour was explained on the basis of a competition between grafting and polymerisation processes. The former basically affects KN, whereas the latter affects pulp brightness.

Enhanced enzymatic saccharification of kenaf powder after ultrasonic pretreatment in ionic liquids at room temperature.

This study demonstrates for the first time that the enzymatic hydrolysis of cellulose is drastically enhanced following ultrasonic pretreatment of lignocellulosic material in ionic liquids (ILs) when compared to conventional thermal pretreatment. Five types of ILs, 1-buthyl-3-methylimidazolium chloride (BmimCl), 1-allyl-3-methylimidazolium chloride (AmimCl), 1-ethyl-3-methylimidazolium chloride (EmimCl), 1-ethyl-3-methylimidazolium diethyl phosphate (EmimDep), and 1-ethyl-3-methylimidazolium acetate (EmimOAc) were tested. Cellulose saccharification ratio was about 20% for kenaf powders pretreated in BmimCl, AmimCl, EmimCl, and EmimDep by conventional heating at 110 °C for 120 min. Conversely, 60-95% of cellulose was hydrolyzed to glucose, subsequent to ultrasonic pretreatment in the same ILs for 120 min at 25 °C. The cellulose saccharification ratio of kenaf powder in EmimOAc was 86% after only 15 min of the ultrasonic pretreatment at 25 °C, compared to only 47% in that case of thermal pretreatment in the IL.

Effects of abscisic acid on ethylene biosynthesis and perception in Hibiscus rosa-sinensis L. flower development.

The effect of the complex relationship between ethylene and abscisic acid (ABA) on flower development and senescence in Hibiscus rosa-sinensis L. was investigated. Ethylene biosynthetic (HrsACS and HrsACO) and receptor (HrsETR and HrsERS) genes were isolated and their expression evaluated in three different floral tissues (petals, style-stigma plus stamens, and ovaries) of detached buds and open flowers. This was achieved through treatment with 0.1 mM 1-aminocyclopropane-1-carboxylic acid (ACC) solution, 500 nl l(-1) methylcyclopropene (1-MCP), and 0.1 mM ABA solution. Treatment with ACC and 1-MCP confirmed that flower senescence in hibiscus is ethylene dependent, and treatment with exogenous ABA suggested that ABA may play a role in this process. The 1-MCP impeded petal in-rolling and decreased ABA content in detached open flowers after 9 h. This was preceded by an earlier and sequential increase in ABA content in 1-MCP-treated petals and style-stigma plus stamens between 1 h and 6 h. ACC treatment markedly accelerated flower senescence and increased ethylene production after 6 h and 9 h, particularly in style-stigma plus stamens. Ethylene evolution was positively correlated in these floral tissues with the induction of the gene expression of ethylene biosynthetic and receptor genes. Finally, ABA negatively affected the ethylene biosynthetic pathway and tissue sensitivity in all flower tissues. Transcript abundance of HrsACS, HrsACO, HrsETR, and HrsERS was reduced by exogenous ABA treatment. This research underlines the regulatory effect of ABA on the ethylene biosynthetic and perception machinery at a physiological and molecular level when inhibitors or promoters of senescence are exogenously applied.

Deciduous shrubs for ozone bioindication: Hibiscus syriacus as an example.

Ozone-like visible injury was detected on Hibiscus syriacus plants used as ornamental hedges. Weekly spray of the antiozonant ethylenediurea (EDU, 300ppm) confirmed that the injury was induced by ambient ozone. EDU induced a 75% reduction in visible injury. Injury was more severe on the western than on the eastern exposure of the hedge. This factor of variability should be considered in ozone biomonitoring programmes. Seeds were collected and seedlings were artificially exposed to ozone in filtered vs. not-filtered (+30ppb) Open-Top Chambers. The level of exposure inducing visible injury in the OTC seedlings was lower than that in the ambient-grown hedge. The occurrence of visible injury in the OTC confirmed that the ozone sensitivity was heritable and suggested that symptomatic plants of this deciduous shrub population can be successfully used as ozone bioindicators. EDU is recommended as a simple tool for diagnosing ambient ozone visible injury on field vegetation.

Nitrogen dioxide at an ambient level improves the capability of kenaf (Hibiscus cannabinus) to decontaminate cadmium.

As reported previously, atmospheric nitrogen dioxide (NO2) at an ambient level increased plant size and the contents of cell constituents. We investigated this effect of atmospheric NO2 on decontamination of cadmium (Cd) by kenaf (Hibiscus cannabinus). Seventeen-day-old seedlings of kenaf were grown in air either with NO2 or without NO2. (Plants were exposed to 100 +/- 50 ppb NO2 for 10 d under irrigation of 0.1% Hyponex supplemented with 20 microM CdCl2.) Plants were then harvested and the biomass of stems, leaves, and roots, as well as the content of Cd in the organs, was determined. The stem and root biomass per plant were 1.25-1.27-fold greater in +NO2 plants than in -NO2 plants. The Cd content per stem was more than 30% greater in +NO2 plants than in -NO2 plants.

Inhibition of nitric oxide synthase (NOS) underlies aluminum-induced inhibition of root elongation in Hibiscus moscheutos.

Aluminum (Al) is toxic to plants when solubilized into Al(3+) in acidic soils, and becomes a major factor limiting plant growth. However, the primary cause for Al toxicity remains unknown. Nitric oxide (NO) is an important signaling molecule modulating numerous physiological processes in plants. Here, we investigated the role of NO in Al toxicity to Hibiscus moscheutos. Exposure of H. moscheutos to Al(3+) led to a rapid inhibition of root elongation, and the inhibitory effect was alleviated by NO donor sodium nitroprusside (SNP). NO scavenger and inhibitors of NO synthase (NOS) and nitrate reductase had a similar inhibitory effect on root elongation. The inhibition of root elongation by these treatments was ameliorated by SNP. Aluminum inhibited activity of NOS and reduced endogenous NO concentrations. The alleviation of inhibition of root elongation induced by Al, NO scavenger and NOS inhibitor was correlated with endogenous NO concentrations in root apical cells, suggesting that reduction of endogenous NO concentrations resulting from inhibition of NOS activity could underpin Al-induced arrest of root elongation in H. moscheutos.

Phytotoxicity data safeguard the performance of the recipient plants in leachate irrigation.

Leachates from an operating and a closed landfill were examined for their phytotoxicity by seed germination/root elongation tests using seeds of Brassica chinensis and Lolium perenne. Their EC50s ranged from 3% to 46% v/v, which varied remarkably with the operating status of the landfills. Seedlings of twelve tree species were grown in pots, which were irrigated with landfill leachate at the EC50 levels, with tap water as control. No tree mortality or growth inhibition was observed after 90 days of leachate application. Chlorophyll fluorescence measurement also showed that plants receiving leachate did not suffer from a decline in photosynthetic efficiency. Litsea glutinosa and Hibiscus tiliaceus had remarkable growth, and other non-N-fixers were not inferior to the N-fixing Acacia auriculiformis. Leachate irrigation improved soil N content, though P deficiency is still a problem. The seed bioassay provided a conservative estimate of the phytotoxicity of landfill leachate. Plants irrigated can be protected from growth inhibition when the leachate irrigation plan is designed with reference to phytotoxicity data.