Modeling ATP protonation and activity coefficients in NaClaq and KClaq by SIT and Pitzer equations

The acid-base properties of Adenosine 5'-triphosphate (ATP) in NaCl and KCl aqueous solutions at different ionic strengths (0相似文献   

Distribution of saccharides and salts on amphoteric ion-exchange resin

An amphoteric ion-exchange resin hardly shrank in 550 and 300 g/L glucose and sodium chloride solutions, respectively; however, the bed packed with a cation-exchange resin shrank considerably. From the distribution coefficients of some saccharides, the swelling pressure of the amphoteric ion-exchange resin was estimated to be 2.0 MPa at 25 °C. The distribution coefficients of glucose, galactose, fructose, and mannose were independent of their concentration and were about 0.621. On the other hand, the apparent distribution coefficients of NaF, NaCl, NaBr, NaI, LiCl, KCl, and CsCl largely depended on concentration. A model for the distribution of salts on the amphoteric resin was proposed, assuming an interaction between the anion of the salt and the positively charged fixed ions with binding constant B. The B values of the chloride salts were nearly the same (1.69–2.94 L/mol), while the values of the sodium salts were largely different depending on the anion.     

Conformational change of poly-N epsilon-glutaryl-L-lysine and poly-N epsilon-succinyl-L-lysine in aqueous salt solutions

The conformational changes of poly-N^ε-glutaryl-L -lysine (PGL) and poly-N^ε-succinyl-L -lysine (PSL) in various salt solutions were studied by use of ORD and potentiometric titration measurements. The addition of alkali metal salts to the fully ionized PGL or PSL solution caused helix formation. The helical content of the polymers increases in the following sequences: at salt concentration 0–2 M, CsCl < KCl < LiCl < NaCl; and at 2–3 M, LiCl < CsCl < KCl ～ NaCl. The preferential binding of the solvent components with various alkali metal salts of PGL or PSL was measured in LiCl, NaCl, and KCl solutions by means of equilibrium dialysis and differential refractometry. It was found that with increasing salt concentration, the polymers were preferentially hydrated in NaCl and KCl soultions; however the salt was preferentially bound to the polymers in LiCl solution. Such preferential binding was suggested to be closely related to conformational change. The addition of CaCl₂ to polymer solutions led to the stabilization of the helical structure of PGL or PSL.     

Temperature and salts effects on the peptidase activities of the recombinant metallooligopeptidases neurolysin and thimet oligopeptidase.

We report the recombinant neurolysin and thimet oligopeptidase (TOP) hydrolytic activities towards internally quenched fluorescent peptides derived from the peptide Abz-GGFLRRXQ-EDDnp (Abz, ortho-aminobenzoicacid; EDDnp, N-(2,4-dinitrophenyl) ethylenediamine), in which X was substituted by 11 different natural amino acids. Neurolysin hydrolyzed these peptides at R-R or at R-X bonds, and TOP hydrolyzed at R-R or L-R bonds, showing a preference to cleave at three or four amino acids from the C-terminal end. The kinetic parameters of hydrolysis and the variations of the cleavage sites were evaluated under different conditions of temperature and salt concentration. The relative amount of cleavage varied with the nature of the substitution at the X position as well as with temperature and NaCl concentration. TOP was activated by all assayed salts in the range 0.05-0.2 m for NaCl, KCl, NH4Cl and NaI, and 0.025-0.1 m for Na2SO4. Concentration higher than 0.2 N NH4Cl and NaI reduced TOP activity, while 0.5 N or higher concentration of NaCl, KCl and Na2SO4 increased TOP activity. Neurolysin was strongly activated by NaCl, KCl and Na2SO4, while NH4Cl and NaI have very modest effect. High positive values of enthalpy (DeltaH*) and entropy (DeltaS*) of activation were found together with an unusual temperature dependence upon the hydrolysis of the substrates. The effects of low temperature and high NaCl concentration on the hydrolytic activities of neurolysin and TOP do not seem to be a consequence of large secondary structure variation of the proteins, as indicated by the far-UV CD spectra. However, the modulation of the activities of the two oligopeptidases could be related to variations of conformation, in limited regions of the peptidases, enough to modify their activities.     

The effect of electrolyte on the morphology of vesicles composed of the dialkyl polyoxyethylene ether surfactant 2C18E12

Small-angle neutron-scattering (SANS) studies were performed on vesicles composed of 1,2-di-O-octadecyl-rac-glyceryl-3-(omega-methoxydodecaethylene glycol), in deuterium oxide (D2O) solutions with various ionic strengths of LiCl, NaCl and NaI. Gross vesicle morphologies, examined using freeze-fracture electron microscopy, showed that NaCl promoted the formation of multilamellar vesicles. Model fitting of the SANS data showed changes in bilayer parameters such as thickness and repeat spacings, in response to the presence of ions in the bulk solution. 2C18E12 vesicles in D2O are shown to exist as predominantly unilamellar structures with a bilayer thickness of approximately 51 A. Vesicles in increasing concentrations of LiCl and NaCl exhibit decreased layer thickness and increased lamelarity. Little change was observed for vesicles formed in NaI solutions. We suggest that these changes result from intrusion of E12 headgroups into the alkyl chain region of the vesicle bilayers, in response to the increase in concentration of ions present and their charge density.     

Modulation of thrombin-fibrinogen interaction by specific ion effects.

Steady-state measurements of synthetic substrate hydrolysis by human alpha-thrombin in the presence of human fibrinogen, under experimental conditions where light scattering due to the formation of fibrin aggregates is negligible, have allowed for a quantitative evaluation of Km for fibrinogen. Measurements of Km for fibrinogen carried out at pH 7.5 and 37 degrees C as a function of NaCl, NaBr, KCl, and KBr concentration, from 50 to 500 mM, show that the derivative d ln Km/d ln a +/-, where a +/- is the mean ion activity, is constant over the entire range of salt concentrations and is strictly dependent on the particular salt present in solution. The values of d ln Km/d ln a +/- are found to be equal to 0.75 +/- 0.03 (NaCl), 0.90 +/- 0.01 (NaBr), 0.62 +/- 0.07 (KCl), and 0.60 +/- 0.03 (KBr). Measurements of Km for two synthetic amide substrates, under identical solution conditions, reveal practically no change in Km with salt concentration, while they show a significant decrease in kcat when Na+ salts are replaced by K+ salts. The drastic difference in the salt dependence of Km between fibrinogen and the synthetic amide substrate points out that a significant role may be played by the fibrinogen recognition site in the energetics of thrombin-fibrinogen interaction. The sensitivity of Km for fibrinogen to different salts unequivocally demonstrates that specific ion effects, rather than nonspecific ionic strength effects, modulate thrombin-fibrinogen interaction under experimental conditions of physiological relevance. Analysis of ion effects on clotting curves obtained at pH 7.5 and 37 degrees C also shows a drastic differential effect of cations and anions.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of high concentration of salts on the esterase activity and structure of a kiwifruit peptidase, actinidain

Effects of salts on the activity and stability of actinidain were examined. With increasing salt concentration up to 0.5 M, the activity (kcat/Km) for N-alpha-Cbz-L-lysine p-nitrophenyl ester decreased to 40% of that in the absence of salt. The inhibitor constant Ki of LiCl, NaCl, and KCl was 0.16-0.43 M. With 3 M KCl and NaCl, the specificity constant kcat/Km recovered to 110 and 75%, respectively. No re-activation was observed with LiCl. The inhibition and re-activation were dependent on the changes in both Km and kcat, whereas no CD change was observed. The tryptophan fluorescence of actinidain was not affected by 0-0.5 M salt, but a considerable decrease in its intensity was observed with increasing salt concentration from 0.5 to 3.0 M. These results suggest that the inhibition observed with the lower salt concentration (<0.5 M) is due to attenuation of the electrostatic interaction between the enzyme and substrate, and the higher concentration (0.5-3.0 M) induces structural change in the states of tryptophan residues, which is associated with the re-activation. Actinidain keeps considerably high activity and stability even in the presence of 3 M salts.     

Adsorption of biopolymers at hydrophilic cellulose-water interface

The extent of adsorption (Gamma2(1)) of bovine serum albumin (BSA), beta-lactoglobulin, lysozyme, gelatin, and DNA from aqueous solution onto the hydrophilic surface of cellulose has been measured as function of biopolymer concentration at different temperatures, pHs, and ionic strengths, and in the presence of a high concentration of inorganic salts and denaturants. In all cases, the value of Gamma2(1) increases with the increase of biopolymer concentration (X2) in bulk and it attains a maximum value at a critical mole fraction concentration X2m. The value of Gamma2m depends upon the nature of protein, temperature, pH, and ionic strength, as well as the nature of neutral salts present in excess. Gamma2m for proteins at a fixed physicochemical condition stands in the following order: Gelatin>betalactoglobulin>lysozyme>BSA. The isotherms for adsorption of DNA nucleotides on cellulose surface at pH 4.0 have been compared at different temperatures and ionic strengths, and in the presence of high concentration of inorganic salts LiCl, NaCl, KCl, and Na2SO4. Values of Gamma2m for different systems have been evaluated and critically compared. At pH 6.0 and 8.0, Gamma2(1) values of DNA nucleotides on cellulose are all negative due to the excess positive hydration of cellulose. At pH 4.0, adsorption of nucleotides of acid, alkali, and heat-denatured DNA widely differ from each other and in the presence of excess concentration of urea becomes negative. The probable mechanisms of biopolymer-cellulose adsorption in terms of polymer hydration, steric interaction, London-van der Waals, hydrophobic, and other types of interactions have been discussed qualitatively. The standard free energy change for the adsorption of protein and DNA nucleotides on the cellulose surface at the state of adsorption saturation has been calculated in kJ per kg of cellulose using an integrated form of the Gibbs adsorption equation. The relation between DeltaG degrees and maximum affinities between biopolymers and the polysaccharide interface have been discussed for various systems.     

Texture formation in DNA films with alkali metal chlorides

The formation of textures in DNA films with LiCl, NaCl, KCl, RbCl, and CsCl salts has been studied. The films are prepared by evaporation of water solution with highly polymerized calf thymus DNA and excess salt of specific type. For DNA solution with 10 mM concentration of NaCl, KCl, and RbCl the films with dendritic textures have been obtained, whereas in case of CsCl the textures in the films appear only at 30 mM concentration of excess salt in the initial solution. In the solution with LiCl, the textures in DNA films have not been observed within the whole range of concentration of excess salt under consideration. The analysis of parameters of DNA films with different salts has showed that evaporation of solution leads to crystallization of salt ions on DNA macromolecule and formation of DNA‐salt complexes. Electrostatic energy of the system of crystalline ordered ions and charges of DNA chains has been estimated to study the stability of DNA‐salt complexes. The results obtained for different salts have been showed that the presence of DNA macromolecule enhances crystallization as compared with solution without DNA. The property of excess salt to form the crystalline structures has been found to decrease in the following order: KCl > NaCl > RbCl > CsCl > LiCl. The results of estimation are in good agreement with the experimentally observed dependence of texture formation on excess salt type. © 2013 Wiley Periodicals, Inc. Biopolymers 99: 508–516, 2013.     

Use of a D-optimal design with constrains to quantify the effects of the mixture of sodium,potassium, calcium and magnesium chloride salts on the growth parameters of <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis>

The combined effect of NaCl, KCl, CaCl(2), and MgCl(2) on the water activity (a (w)) and the growth parameters of Saccharomyces cerevisiae was studied by means of a D-optimal mixture design with constrains (total salt concentrations < or = 9.0%, w/v). The a (w) was linearly related to the concentrations of the diverse salts; its decrease, by similar concentrations of salts, followed the order NaCl > CaCl(2) > KCl > MgCl(2), regardless of the reference concentrations used (total absence of salts or 5% NaCl). The equations that expressed the maximum specific growth (mu (max)), lag phase duration (lambda), and maximum population reached (N (max)) showed that the values of these parameters depended on linear effects and two-way interactions of the studied chloride salts. The mu (max) decreased as NaCl and CaCl(2) increased (regardless of the presence or not of previous NaCl); however, in the presence of a 5% NaCl, a further addition of KCl and MgCl(2) markedly increased mu (max). The lambda was mainly affected by MgCl(2) and the interactions NaCl x CaCl(2) and CaCl(2) x MgCl(2). The further addition of NaCl and CaCl(2) to a 5% NaCl medium increased the lag phase while KCl and MgCl(2) had negligible or slightly negative effect, respectively. N (max) was mainly affected by MgCl(2) and its interactions with NaCl, KCl, and CaCl(2); MgCl(2) stimulated N (max) in the presence of 5% NaCl while KCl, NaCl, and CaCl(2) had a progressive decreasing effect. These results can be of interest for the fermentation and preservation of vegetable products, and foods in general, in which this yeast could be present.     

Modulation of thrombin-hirudin interaction by specific ion effects.

Kinetic studies of the inhibition of thrombin amidase activity by recombinant hirudin have been conducted as a function of salt concentration in the range 0.05 to 1 M, using NaCl, KCl, NaBr and KBr. At the same ionic strength, the value of KI for thrombin-hirudin interaction is found to be different with different salts. The slope d ln KI/d ln a+/-, where a+/- is the mean ion activity, is constant in the range 0.05 to 0.5 M, is sensitive to the particular salt present in solution and is equal to 1.07 +/- 0.09 (NaCl), 0.92 +/- 0.10 (KCl), 1.37 +/- 0.10 (NaBr) and 0.56 +/- 0.10 (KBr). These results indicate that specific ion effects are involved in the modulation of thrombin-hirudin interaction in the form of ion release, as recently found in the case of thrombin interaction with its natural substrate fibrinogen. The linkage hierarchy for ion release found in the case of thrombin-fibrinogen interaction also applies in the case of thrombin-hirudin interaction, with the number of released ions decreasing in the order NaBr greater than NaCl greater than KCl greater than KBr. It is proposed that the process of bridge-binding to the fibrinogen recognition site and the catalytic pocket of the enzyme, as seen in the case of fibrinogen and hirudin, is linked to ion release and controlled by modulation of the association rate constant.     

Growth characteristics and salt requirement ofDeleya halophila in a defined medium

Growth characteristics ofDeleya halophila (CCM 3662^T), were determined using a defined medium.Deleya halophila presented its optimal growth at 7.5% (wt/vol) total salts when it was grwon at incubation temperatures of 32° and 42°C; when the temperature was lowered to 22°C, it had optimal growth at 5% (wt/vol) total salts. This bacterium had an absolute requirement for the Na⁺ cation; it could not be replaced by other cations. NaBr, Na₂SO₄, or Na₂S₂O₃ could be substituted for NaCl in the growth medium, but, when MgCl₂, KCl, LiCl, NaI, NaF, or NaNO₃ was substituted for NaCl, the medium did not support growth. Growth rates of the strain were diverse when NaCl was partially replaced by other sodium salts. Finally,D. halophila suffered loss of viability when the culture was diluted into different low NaCl concentrations (0, 0.5%, and 1%, wt/vol) at various incubation temperatures.     

Differentiation and characterization of the cytoplasmic and nuclear deoxyribonucleic acid polymerases from baby hamster kidney cells.

Distinct DNA polymerase activities have been found in the cytoplasmic and nuclear fractions of a baby hamster kidney cell line. They were separated by chromatography on DEAE-cellulose and partially purified by ammonium sulfate fractionation, DNA - cellulose and linear sucrose gradients. The cytoplasmic DNA polymerase exhibited an S-coefficient of 6.95 S in 0.15 M NaCl and its activity was highly sensitive to inhibition by N-ethylmaleimide and elevated temperatures, regardless of the presence of DNA template or other cofactors. It was stimulated by monovalent salts in the order of NH4 Cl greater than KCl greater than NaCl greater than CsCl greater than LiCl (inhibitory). The DNA polymerase extracted from nuclei sedimented with an S-value of 3.47 S, was resistant to inactivation by N-ethylmaleimide, and maximally stimulated by NaCl, while also being inhibited by LiCl. For optimal activity, both DNA polymerase activities required a divalent cation, with MgCl2 being more effective than MnCl2. Although the optimal pH values for the two enzyme activities differed slightly, glycine - NaOH buffer induced an alkaline shift of 1.5 pH units in the optimum of both enzymes. This was accompanied by an increase in the effectiveness of MnCl2 relative to MgCl2 for the cytoplasmic DNA polymerase.     

Cation radius effects on cell-free translation in rabbit reticulocyte lysate

The effect of monovalent cation concentrations on the translation was examined in the rabbit reticulocyte cell-free system. The translation of standard reporter gene luciferase was studied using different concentrations of LiCl, NaCl, KCl, RbCl, CsCl, NH(4)Cl, and (CH(3))(4)NCl and the acetates of Na(+), K(+), and NH4(+). Only the salts of K(+), Rb(+), and NH4(+) and to some minor extent of Cs(+) significantly supported translation. Optimum concentrations were dependent on the cation used. Optimum concentrations ranged between 40 mM (NH(4)Ac), 80 mM (KCl, NH(4)Cl), and 100 mM (RbCl, KAc). The maximum efficiency of translation depends on the ionic radius of the cation used. KCl and RbCl were superior to all other salts tested in stimulating in vitro translation. The results were confirmed, using a second reporter system, M-hirudin. Here, however, broad optima were observed with RbCl being slightly superior to KCl in supporting translation.     

Modulation of brain S2 serotonin receptors by lithium, sodium and potassium chloride

The effect of LiCl, NaCl, and KCl on serotonin competition for 3H-ketanserin binding to S2 serotonin receptors in homogenates of rat prefrontal cortex were investigated. LiCl was the most potent of the ionic modulators in lowering the apparent affinity of serotonin for the S2 serotonin receptor. A threshold effect was noted at 12 mM LiCl (a 60% change in IC50); at 120 mM LiCl a nine-fold shift in the serotonin IC50 was noted. 120 mM NaCl or KCl demonstrated similar effects as 12 mM LiCl in reducing serotonin's apparent affinity. These results indicated that monovalent cations modulate S2 serotonin receptor affinity for serotonin and that lithium ion is more potent than sodium or potassium.     

Modeling the acid–base properties of glutathione in different ionic media, with particular reference to natural waters and biological fluids

The acid-base properties of γ-L-glutamyl-L-cysteinyl-glycine (glutathione, GSH) were determined by potentiometry (ISE-H(+), glass electrode) in pure NaI((aq)) and in NaCl((aq))/MgCl(2(aq)), and NaCl((aq))/CaCl(2(aq)) mixtures, at T = 298.15 K and different ionic strengths (up to I(c) ~ 5.0 mol L(-1)). In addition, the activity coefficients of glutathione were also determined by the distribution method at the same temperature in various ionic media (LiCl((aq)), NaCl((aq)), KCl((aq)), CsCl((aq)), MgCl(2(aq)), CaCl(2(aq)), NaI((aq))). The results obtained were also used to calculate the Specific ion Interaction Theory (SIT) and Pitzer coefficients for the dependence on medium and ionic strength of glutathione species, as well as the formation constants of weak Mg(j)H( i )(GSH)((i+2j-3)) and Ca(j)H(i)(GSH)((i+2j-3)) complexes. Direct calorimetric titrations were also carried out in pure NaCl((aq)) and in NaCl((aq))/CaCl(2(aq)) mixtures at different ionic strengths (0.25 ≤ I (c )/mol L(-1) ≤ 5.0) in order to determine the enthalpy changes for the protonation and complex formation equilibria in these media at T = 298.15 K. Results obtained are useful for the definition of glutathione speciation in any aqueous media containing the main cations of natural waters and biological fluids, such as Na(+), K(+), Mg(2+), and Ca(2+). Finally, this kind of systematic studies, where a series of ionic media (e.g., all alkali metal chlorides) is taken into account in the determination of various thermodynamic parameters, is useful for the definition of some trends in the thermodynamic behavior of glutathione in aqueous solution.     

Structural transitions of deoxyribonucleic acid in aqueous electrolyte solutions. I. Reference spectra of conformational limits.

The circular dichroism properties of calf thymus DNA have been examined at 27 degrees over the wavelength range of 215-300 nm in aqueous solutions of NaCl, KCl, LiCl, CsCl, and NH4Cl at pH 7. The concentrations of these electrolytes were varied from 0.01 to ca. 5-10 m. The spectral changes induced by changes in concentration of NaCl and KCl and all but the highest concentrations of NH4Cl as well as lower concentrations of Cstcl and LiCl could be represented by a common two-state transition involving the conversion of the typical conservative spectrum commonly seen in dilute solutions of these salts to a nonconservative spectrum similar to that obtained by Tunis-Schneider and Maestre ((1970), J. Mol. Biol. 52, 521) for the C form of DNA. At higher concentrations of CsCl, LiCl, and NH4Cl, an additional component, resembling an A type spectrum, was required to account for the observed CD changes with changing concentration of electrolyte. Relying on the published spectra of the B, the C, and the A forms of DNA by Tunis-Schneider and Maestre for identification and approximate values of the molecular ellipticities of these forms, we have analyzed these spectral transitions by two least mean squares methods in order to obtain accurate reference spectra of aqueous "B", C, and "A" conformations of calf thymus DNA. The results obtained suggest that although the C form in solution is identical with that obtained in film, the aqueous B conformational limit is not identical with the crystallographic Watson-Crick structure. In addition, the A form generated in solution under our experimental conditions appears to be more similar to that assumed by low molecular weight Escherichia coli DNA at 75% relative humidity rather than calf thymus DNA at the same relative humidity.     

Effect of Salts on Growth and Pectinase Production by Halotolerant Yeast, <Emphasis Type="Italic">Debaryomyces nepalensis</Emphasis> NCYC 3413

In this study, Debaryomyces nepalensis NCYC 3413 isolated from rotten apple was studied for its halotolerance and its growth was compared with that of Saccharomyces cerevisiae in high salt medium. The specific growth rate of D. nepalensis was not affected by KCl even up to a concentration of 1 M, whereas NaCl and LiCl affected the growth of D. nepalensis. Among all tested salts, LiCl showed maximum inhibition on growth. At all conditions, halotolerance of D. nepalensis was much higher than that of S. cerevisiae. D. nepalensis showed maximum viability (80–100%) when grown in KCl, which was higher than with NaCl and LiCl. Pectinase production by D. nepalensis was noted at all high salt concentrations, namely, 2 M NaCl, 2 M KCl, and 0.5 M LiCl, and the maximum specific activity was observed when the strain was grown in 2 M NaCl.     

MICRURGICAL STUDIES IN CELL PHYSIOLOGY : V. THE ANTAGONISM OF CATIONS IN THEIR ACTIONS ON THE PROTOPLASM OF AM?BA DUBIA.

I. The Plasmalemma. 1. On the plasmalemma of amebæ CaCl₂ antagonizes the toxic action of LiCl better than it does NaCl, and still better than it does KCl. MgCl₂ antagonizes the toxic action of NaCl better than it does LiCl and still better than it does KCl. 2. CaCl₂ antagonizes the toxic action of LiCl and of KCl better than does MgCl₂: MgCl₂ antagonizes NaCl better than does CaCl₂. II. The Internal Protoplasm. 3. The antagonizing efficiency of CaCl₂ and of MgCl₂ are highest against the toxic action of KCl on the internal protoplasm, less against that of NaCl, and least against that of LiCl. 4. CaCl₂ antagonizes the toxic action of LiCl better than does MgCl₂: MgCl₂ antagonizes the toxic action of NaCl and of KCl better than does CaCl₂. 5. LiCl antagonizes the toxic action of MgCl₂ on the internal protoplasm more effectively than do NaCl or KCl, which have an equal antagonizing effect on the MgCl₂ action. III. The Nature of Antagonism. 6. When the concentration of an antagonizing salt is increased to a toxic value, it acts synergistically with a toxic salt. 7. No case was found in which a potentially antagonistic salt abolishes the toxic action of a salt unless it is present at the site (surface or interior) of toxic action. 8. Antagonistic actions of the salts used in these experiments are of differing effectiveness on the internal protoplasm and on the surface membrane.     

A novel method for increasing the frequency of somatic embryogenesis in wheat tissue culture by NaCl and KCl supplementation

The effect of NaCl, KCl and LiCl on the growth and morphogeneis of tissue cultures originating from immature embryos of four wheat (Triticum aestivum L.) and one triticale (Triticosecale)varieties was investigated. The morphogenetic pathway to plant regeneration in Chinese Spring wheat was determined as incomplete somatic embryogenesis because the differentiation and subsequent germination of the shoot apices happened in the early phase of embryo development. Culture medium supplemented by NaCl suppressed the differentiation of shoot apices resulting in the development of more typical somatic embryoids. Forty mM concentrations of both NaCl or KCl increased the formation of somatic embryos in Chinese Spring. Arthur and GK Kincso wheat varieties while Lasko triticale regenerated well without the addition. The salts inhibited plantlet formation from somatic embryoids so the salts supplement should be omitted. Forty mM LiCl inhibited growth while 10mM LiCl had no effect on growth or embryogenesis.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - MS Murashige and Skoog (1962) medium