Menthol tolerant clones of Mentha arvensis: approach for in vitro selection of menthol rich genotypes

In vitro raised shoots of Mentha arvensis L. were screened for menthol tolerance level by growing them in media containing 0–100 g ml^–1 menthol. A total of 2850 regenerated shoots were step wise screened for menthol tolerance at the concentrations of 50 g ml^–1 followed by 60 and 70 g ml^–1. In this screening, only 30 individual regenerated shoots were able to survive. The clones from the primary screen were inoculated into rooting medium and, after rooting, transferred to pots in the greenhouse. Ultimately, these 30 menthol tolerant clones were multiplied and grown in the field in replicated plots of 2.5×2.5 m sizes. Twigs of 30 clones from the replicated trials were rechecked for tolerant phenotypes at a concentration of 70 g ml^–1 menthol wherein, these survived even after 7 days (secondary screening). These clones were checked for oil and menthol content and were found to be better than the control plants. Out of these 30 plants, five tolerated 80 g ml^–1 menthol (tertiary level screening) and were found to contain the highest amount of menthol per g leaf biomass. Molecular analysis through RAPD showed distinct variation in the profiles of these five plants, in comparison to the control. Using this method the relationship between the primer OPT 04, menthol tolerance and high menthol content character of the genotype was established. Further, a cultivar `Saksham' was released from the selections by CIMAP for superior performance.     

Structural and spectroscopic studies of tripodal [MgL]2+ chelates containing only nitrogen donor atoms: alkaline earth metal complexes as potential drug delivery agents

Several tripodal diimine ligands, tris(2-(2-thiazolyl)methyliminoethyl)amine, 2-Tatren, tris(2-(4-(5-methyl)imidazolyl)methyliminoethyl)amine, 5-Me-4-Imtren, tris(2-(4-imidazolyl)methyliminoethyl)amine, 4-Imtren, tris(2-(2-imidazolyl)methyliminoethyl)amine, 2-Imtren, and their Mg(2+) complexes were prepared and characterized. X-ray diffraction studies show that the Mg(2+) ions are six-coordinate, with three acyclic imine N atoms and three imidazolyl or thiazolyl N atoms coordinated with the general formula [Mg(L)](ClO(4))(2) (L=4-Imtren (1), 2-Imtren (2), 2-Tatren (3), and 5-Me-4-Imtren (4)). These complexes are chiral with both Delta and Lambda isomers present in the unit cell. (1)H NMR titrations reveal that complexes also form in solution and that the chirality is maintained. Variable temperature (1)H NMR reveals that the Delta and Lambda isomers interconvert in the intermediate to slow time scale. The interconversion rate slows with increasing pK(a) of the ligand heterocycle, suggesting that interconversion proceeds through a partially dissociated state. These complexes undergo trans-metallation by Zn(2+), indicating that their ligands can be released in a kinetically facile manner to form more stable metal ion complexes.     

Production and secretion of interleukin-1alpha proteins by rat testis

The present study characterizes constitutively expressed rat testicular interleukin-1α (IL-1α) proteins. IL-1 bioactivity of crude testis protein was completely neutralized by IL-1α antiserum, IL-1 receptor antagonist, and soluble type I IL-1 receptor. Upon non-denaturating gel permeation chromatography, bioactive IL-1 eluted at molecular sizes of 45, 31, and 17 kDa and at charges of pH 5.7 and 6.0 after chromatofocusing. SDS-PAGE/Western blot analysis of proteins extracted from whole testis, seminiferous tubules, interstitial, and seminiferous tubule fluids all demonstrated IL-1α immunoreactivity at 45, 24, and 19 kDa. Activated macrophages and tissue proteins from endotoxin treated rats showed immunoreactive 31 and 19 kDa IL-1α. The results indicate that the testis produces three isoforms of IL-1α proteins that are secreted into the interstitial compartment and tubular lumen where they may exert paracrine functions. The testicular IL-1α isoforms may represent posttranslationally modified precursor, mature IL-1α, and a 24-kDa alternate splice form.     

Proton efflux from oat coleoptile cells and exchange with wall calcium after IAA or fusicoccin treatment

Elongation growth of plant cells occurs by stretching of cell walls under turgor pressure when intermolecular bonds in the walls are temporarily loosened. The acid-growth theory predicts that wall loosening is the result of wall acidification because treatments (including IAA and fusicoccin) that cause lowered wall pH cause elongation. However, conclusive evidence that IAA primarily reduces wall pH has been lacking. Calcium has been reported to stiffen the cell walls. We have used a microelectrode ion-flux measuring technique to observe directly, and non-invasively, the net fluxes of protons and calcium from split coleoptiles of oats (Avena sativa L.) in unbuffered solution. Normal net fluxes are 10 nmol · m⁻² · s⁻¹ proton efflux and zero calcium flux. The toxin fusicoccin (1 μM) causes immediate efflux from tissue not only of protons, but also of calcium, about 110 nmol · m⁻² · s⁻¹ in each case. The data fit the “weak acid Donnan Manning” model for ion exchange in the cell wall. Thus we associate the known “acid-growth” effect of fusicoccin with the displacement of calcium from the wall by exchange for protons extruded from the cytoplasm. Application of 10 μM IAA causes proton efflux to increase transiently by about 15 nmol · m⁻² · s⁻¹ with a lag of about 10 min. The calcium influx decreases immediately to an efflux of about 20 nmol · m⁻² · s⁻¹. It appears that auxin too causes an “acid-growth” effect, with extruded protons exchanging for calcium in the cell walls. I. Arif is currently recieving an AIDAB scholarship. This work was supported by an Australian Research Council grant to I.A. Newman.     

Kinetics of reduction of ferrioxamine B by chromium(II), vanadium(II), and dithionite

Kinetic studies of the reduction of ferrioxamine B (Fe(Hdesf)⁺) by Cr(H₂O)₆²⁺, V(H₂O)₆²⁺, and dithionite have been performed. For Cr(H₂O)₆²⁺ and V(H₂O)₆²⁺, the rate is ?d[Fe(Hdesf)⁺]/dt = k[Fe(Hdesf)⁺][M²⁺]. For Cr(H₂O)₆²⁺, k = 1.19 × 10⁴ M^?1 sec^?1 at 25°C and μ = 0.4 M, and k is independent of pH from 2.6 to 3.5. For V(H₂O)₆²⁺, k = 6.30 × 10² M^?1 sec^?1 at 25°C, μ = 1.0 M, and pH = 2.2. The rate is nearly independent of pH from 2.2 to 4.0. For Cr(H₂O)₆²⁺ and V(H₂O)₆²⁺, the activation parameters are ΔH^≠ = 8.2 kcal mol^?1, ΔS^≠ ?12 eu and ΔH^≠ = 1.7 kcal mol^?1, ΔS^≠ = ?40 eu (at pH 2.2) respectively. Reduction by Cr(H₂O)₆²⁺ is inner-sphere, while reduction by V(H₂O)₆²⁺ is outer-sphere. Reduction by dithionite follows the rate law ?d[Fe(Hdesf)⁺]/dt =kK^{$\text{1}\text{2}$}[Fe(Hdesf)⁺][S₂O₄^2?]^{$\text{1}\text{2}$} where K is the equilibrium constant for dissociation of S₂O₄^2? into SO₂^? radicals. The value of k at 25°C and μ = 0.5 is 2.7 × 10³ M^?1 sec^?1 at pH 5.8, 3.5 × 10³ M^?1 sec^?1 at pH 6.8, and 4.6 × 10³ M^?1 sec^?1 at pH 7.8, and ΔH^≠ = 6.8 kcal mol^?1 and ΔS^≠ = ?19 eu at pH 7.8.     

An ecdysone-inducible putative "DEAD box" RNA helicase in the spruce budworm (Choristoneura fumiferana)

RNA helicases are a family of enzymes that unwind nucleic acid duplexes, such as RNA/RNA and RNA/DNA, in a 3' to 5' direction into single-stranded polynucleotides. A putative RNA helicase cDNA (CfrHlc64) was isolated from the spruce budworm, Choristoneura fumiferana. CfrHlc64 was 1998 nucleotides in length, and the deduced protein had 565 amino acids with a predicted molecular mass of 64 kDa. It contained eight functional motifs conserved in the "DEAD box" family of RNA helicases. The deduced amino acid sequence showed 10-50% identities to homologues of other species from bacteria to human. In vitro expression of the cDNA resulted in recombinant proteins of 64 kDa as expected from the deduced amino acid sequence. Northern blotting and RT-PCR analyses revealed the presence of CfrHlc64 mRNA in all developmental stages from embryo to adult. Higher levels of CfrHlc64 mRNA were detected in the fat body and midgut than in the epidermis of sixth instar larvae. The CfrHlc64 protein was distributed mainly in the fat body. Female adults expressed CfrHlc64 mRNA at higher levels than male adults. The nonsteroidal ecdysone agonist, tebufenozide, enhanced the expression of CfrHlc64 in a dose-dependent manner.     

Mental health and coping in a war situation: the case of Bosnia and Herzegovina

The latter part of the twentieth century has seen an increased concern for the implications of war for civilian populations, and more attention has been given to psychosocial impacts of uprooting and displacement. 'Loss of place', acute and chronic trauma, family disruption and problems of family reunification have become issues of concern. The war in Bosnia was characterized by massive displacement, disruption and loss of life, relatives and property. Health and psychosocial well-being were affected in a number of ways. There was an overwhelming loss of perceived power and self-esteem. Over 25%, of displaced people, for example, said they no longer felt they were able to play a useful role; even in non-displaced populations approximately 11% of those interviewed said that they had lost a sense of worth. Widespread depression and feelings of fatigue and listlessness were common and may have prevented people from taking steps to improve their situation. Almost a quarter of internally displaced people had a high startle capacity and said they were constantly nervous. Most adverse psychosocial responses increased with age and in a population that includes many elderly people this poses serious problems. The findings point to major challenges with respect to repatriation and reconstruction. They highlight the importance of family reunification and the facilitating of decision-making by affected people themselves. The findings also shed light on potential problems associated with over-dependence on external assistance and hence the need for people to be given the means of using their skills and knowledge to control their day-to-day lives.     

Modeling carbon monoxide dehydrogenase/acetyl-CoA synthase (CODH/ACS): a trinuclear nickel complex employing deprotonated amides and bridging thiolates

Carbon monoxide dehydrogenase/acetyl-CoA synthase (CODH/ACS) utilizes a unique Ni-M bimetallic site in the biosynthesis of acetyl-CoA, where a square-planar Ni ion is coordinated to two thiolates and two deprotonated amides in a Cys-Gly-Cys motif. The identity of M is currently a matter of debate, although both Cu and Ni have been proposed. In an effort to model ACSs unusual active site and to provide insight into the mechanism of acetyl-CoA formation and the role of each of the metals ions, we have prepared and structurally characterized a number of Ni(II)–peptide mimic complexes. The mononuclear complexes Ni(II) N,N-bis(2-mercaptoethyl)oxamide (1), Ni(II) N,N-ethylenebis(2-mercaptoacetamide) (2), and Ni(II) N,N-ethylenebis(2-mercaptopropionamide) (3) model the Ni(Cys-Gly-Cys) site and can be used as synthons for additional multinuclear complexes. Reaction of 2 with MeI resulted in the alkylation of the sulfur atoms and the formation of Ni(II) N,N-ethylenebis(2-methylmercaptoacetamide) (4), demonstrating the nucleophilicity of the terminal alkyl thiolates. Addition of Ni(OAc)₂·4H₂O to 3 resulted in the formation of a trinuclear species 5, while 2 crystallizes as an unusual paddlewheel complex (6) in the presence of nickel acetate. The difference in reactivity between the similar complexes 2 and 3 highlights the importance of ligand design when synthesizing models of ACS. Significantly, 5 maintains the key features observed in the active site of ACS, namely a square-planar Ni coordinated to two deprotonated amides and two thiolates, where the thiolates bridge to a second metal, suggesting that 5 is a reasonable structural model for this unique enzyme.Ø. Hatlevik and M.C. Blanksma contributed equally to this work