Formation of calcium-deficient hydroxyapatite from alpha-tricalcium phosphate

This study investigated the factors influencing the kinetics of Ca9HPO4(PO4)5OH (calcium deficient hydroxyapatite or CDHAp) formation from alpha-Ca3(PO4)2 (alpha-TCP). The kinetics of CDHAp formation were investigated by isothermal calorimetry at constant temperatures ranging between 30 and 75 degrees C and by changes in pH at 37.4 and 70 degrees C. The calorimetric curves were characterized by two reaction peaks. Activation energies were calculated for the events resulting in these peaks. Values obtained were 48.4 and 67.7 kJ mol(-1), respectively, indicating nucleation and growth mechanisms for both events. Temperature had a significant effect on the growth rate as indicated by a decrease in surface area (26.5-15.0 m2 g(-1)) of the CDHAp with increasing temperature (30-75 degrees C). A linear relationship between hydrolysis temperature and CDHAp surface area was observed. The morphology of the CDHAp was plate-like and the crystallites became more regular as the reaction temperature was increased. A rapid elevation in pH upon mixing with water indicated the synthesis method initially used did not entirely eliminate slight compositional variations within the alpha-Ca3(PO4)2. Rapid elevation in pH retarded subsequent reaction. This effect was eliminated by increasing the duration of high-temperature firing during alpha-TCP synthesizing.     

Selective Precipitation of Phosphate from Semiconductor Wastewater

Selective precipitation of phosphate from fluoride-containing wastewater was studied using magnesium salts. Wastewater sampled from a semiconductor manufacture was found to contain 936 mg/L of fluoride (F?), 640 mg/L of sulfate (SO42?), 118 mg/L of phosphate (PO43?), and 26.72 mg/L ammonium (NH4+). Magnesium chloride (MgCl2) was more effective than magnesium oxide (MgO) in inducing precipitation reactions between magnesium and phosphate. Effects of both molar ratio ([Mg2+]:[PO43?]) and pH were examined and experimental results were compared with those from equilibrium modeling by PHREEQC. A total of 41.72% of phosphate was removed and recovered when at molar ratio of 3:2 and pH of 10. Precipitation of phosphate increased with increasing molar ratio and 66.19% of phosphate was recovered when molar ratio was 3:1. Precipitation of phosphate increased as pH changed from 8 to 10. However, it decreased as pH increased from 10 to 12, probably because of competition between phosphate and hydroxyl (OH?) ions. Solid precipitates were characterized by field emission scanning electron microscope with energy dispersive spectrometer and X-ray diffraction. In accordance with theoretical modeling, it was found that the precipitate in pH range of 8–10 was predominantly bobierrite [Mg3(PO4)2.8H2O] and some amorphous precipitates. However, brucite [Mg(OH)2] precipitate would start to form when pH became higher than 10. Results showed that MgCl2 can be a selective precipitation reagent for phosphate removal from semiconductor wastewater since it does not form precipitate with fluoride, sulfate, and ammonium.     

NaCl-KCl-CaCl_2熔盐体系中Mg~(2+)在Mo电极上的阴极过程

采用钼电极,在973K的温度下,研究了MgCl2在三元NaCl-KCl-CaCl2熔融氯化物混合物中的电化学行为。采用方波伏安法(SWV)、循环伏安法(CV)、计时电流法(CA)和计时电位法(CP)研究Mg~(2+)在钼电极上的反应过程。结果表明,Mg~(2+)在钼电极上发生一步还原反应(Mg~(2+)+2e=Mg),是由扩散控制的可逆反应,镁在钼电极上的成核过程是瞬时成核,这与半球核的形成和生长机制是一致的。通过CV、CP和CA在973K的不同方法计算的Mg~(2+)扩散系数分别为1.09×10-5、1.24×10-5和3.23×10-5 cm2/s。在给定电位下阴极沉积产物为金属镁。     

Chemical ligation of oligodeoxyribonucleotides on circular DNA templates

We report the use of small circular DNA as a triplex-directing template for the highly efficient chemical ligation of oligodeoxyribonucleotides (ODNs) using cyanogen bromide (BrCN). These investigations compared the use of a linear homopyrimidine DNA template (17mer) and a circular pyrimidine-rich DNA template (44mer) for directing the chemical ligation of two homopurine ODNs (6mer + 11mer). The effects of substrate/template ratio, buffer, salt, ionic strength, pH and temperature have been examined in the BrCN activated ligation reactions. The optimal yield of 51% for ligation on the linear template was at pH 6.0, 200 mM MgCl2, 4 degreesC. In contrast, near quantitative ligation on the circular template occurred at higher pH, higher temperature, and showed less dependence on Mg2+concentration (97% yield, pH 7.5, 200 mM MgCl2, 25 degreesC). The relative observed rate of the ligation reaction was a minimum of 35 times faster on the circular DNA template relative to the linear template at pH 7.5, 200 mM MgCl2, 4 degreesC. These investigations reveal that chemical ligation of short ODNs on circularized DNA templates through triplex formation is a highly efficient process over a broad range of conditions.     

Removal of magnesium from nickel laterite leach liquors by chemical precipitation using calcium hydroxide and the potential use of the precipitate as a filler material

The chemical precipitation of magnesium from sulphate solution, resulting from heap leaching of nickeliferous laterites with sulphuric acid, was studied. Magnesium was removed as hydroxide using calcium hydroxide (Ca(OH)₂) and the precipitate produced was a mixture of magnesium hydroxide (Mg(OH)₂) and gypsum (CaSO₄·2H₂O). The variables studied were the temperature and the stoichiometric quantity of Ca(OH)₂. The responses measured were magnesium removal and the specific surface of the precipitate. Design of the experiments and statistical analysis of the data were used in order to determine the main effects and interactions of the factors. Scanning electron microscopy (SEM) was also used to investigate the effect of precipitation conditions on the morphological characteristics of the Mg(OH)₂–CaSO₄·2H₂O mixture. Kinetic analysis with the aid of Nielsen theory allowed the determination of the Mg(OH)₂ formation mechanism. The use of a magnesium hydroxide–gypsum mixture as a filler material was also examined. The suitability of the precipitate was evaluated by measuring a set of properties that can characterize a material as a filler and by measuring mechanical properties of polymers filled with the precipitate at various addition levels. The magnesium hydroxide–gypsum precipitate proved to be promising for this application, as it was found to have similar properties with other commercial products.     

Identification of members of the HSP30 small heat shock protein family and characterization of their developmental regulation in heat-shocked Xenopus laevis embryos

Uptakes of (28)Mg at 10 s were measured at 0.1 and 1mM MgCl(2), to mainly represent one or other of the two uptake mechanisms earlier shown to be present in rat jejunal brush border membrane vesicles, one with an apparent KT of 0.2 mM, the other in the millimolar range. Both mechanisms had an optimal temperature close to 28 degrees C, inactivation at 37 degrees C being more acute for the low affinity mechanism (55 percent, P < 0.01). Both mechanisms were equally stimulated by an electrical potential difference (negative inside the vesicles) imposed by a potassium gradient and not affected by the nature of the anion accompanying magnesium. At 0.1 mM MgCl(2), the uptake was increased by an outwardly directed proton gradient, pH 8.2 outside and 7.4 inside (38 percent, P < 0.05), but not depressed when the gradient was in the opposite direction, pH 6.6 outside and 7.4 inside. It was trans-stimulated by magnesium, strongly inhibited by amiloride and to a smaller extent by furosemide, but uninfluenced by 0.1 mM NaCl or by 100 mM NaCl, NaSCN or KCl. A slight but significant inhibition (20-30 per cent) was recorded in the presence of 0.1 mM CoCl(2), NlCl(2) or BaCl(2). At 1 mM MgCl(2), the uptake was not influenced by pH gradient, was not trans-stimulated by Mg and was not affected by furosemide. A 40 percent inhibition by amiloride was, however, recorded. Also 100 mM NaCl or KCl doubled the uptake, while 1 mM NaCl or 100 mM NaSCN did not affect it. In contrast, all the divalent cations tested produced an inhibition (from 60 to 12 percent) in the following order: Co > or = Mn > Ca > or = Ni> Ba > Sr, when used at the same concentration as magnesium. The results showed that cobalt and calcium were not true competitors. In conclusion, two distinct mechanisms would operate magnesium entry at the brush border: (1) an electrogenic high affinity Mg/Mg,H exchange, sensitive to amiloride and furosemide, and (2) an electrogenic low affinity mechanism, inhibited by the presence of several divalent cations and dependent on the presence or activity of alkaline phosphatase.     

Structural, electrical and surface morphological studies of Cd2SnO4 and Mg doped Cd2SnO4 thin films

The technologically important conductive Cadmium stannate (Cd₂SnO₄) and Magnesium doped cadmium stannate (Cd₂SnO₄: Mg) thin films were prepared using Cadmium acetate and stannous chloride by spray pyrolysis technique. Films prepared are crystalline and have cubic CdO phase with SnO₂. The X-ray diffraction patterns of both thin films shows perfect crystalline structure and from the data, the particle diameter of both films were calculated. Atomic Force Microscopy (AFM) of both films reveals the uniform thickness of the films and the presence of uniform grain growth in Cd₂SnO₄ and Mg doped Cd₂SnO₄ thin films. Thickness of Cadmium stannate film is 725nm and that of Magnesium doped Cadmium stannate film is 285nm. The indirect band gap energy of Cd₂SnO₄ film is 2.71eV and for magnesium doped Cd₂SnO₄ is 2.97eV were observed from the UV-Visible absorption spectrum studies. Presence of uniform grain growth is found in both thin films. The doping of Magnesium in Cadmium stannate film improves the electrical properties without affecting its structural properties.     

Assembly mechanism of Dictyostelium myosin II: regulation by K+, Mg2+, and actin filaments

Regulated assembly of myosin II in Dictyostelium discoideum amoebae partially controls the orderly formation of contractile structures during cytokinesis and cell migration. Kinetic and structural analyses show that Dictyostelium myosin II assembles by a sequential process of slow nucleation and controlled growth that differs in rate and mechanism from other conventional myosins. Nuclei form by an ordered progression from myosin monomers to parallel dimers to 0.43 microns long antiparallel tetramers. Lateral addition of dimers to bipolar tetramers completes the assembly of short (0.45 microns) blunt-ended thick filaments. Myosin heads are not staggered along the length of tapered thick filaments as in skeletal muscle, nor are bipolar minifilaments formed as in Acanthamoeba. The overall assembly reaction incorporating both nucleation and growth could be kinetically characterized by a second-order rate constant (kobs,N+G) of 1.85 x 10(4) M-1 s-1. Individual rate constants obtained for nucleation, kobs,N = 4.5 x 10(3) M-1 s-1, and growth, kobs,G = 2.5 x 10(4) M-1 s-1, showed Dictyostelium myosin II to be the slowest assembling myosin analyzed to date. Nucleation and growth stages were independently regulated by Mg2+, K+, and actin filaments. Increasing concentrations of K+ from 50 to 150 mM specifically inhibited lateral growth of dimers off nuclei. Intracellular concentrations of Mg2+ (1 mM) accelerated nucleation but maintained distinct nucleation and growth phase kinetics. Networks of actin filaments also accelerated the nucleation stage of assembly, mechanistically accounting for spontaneous formation of actomyosin contractile fibers via myosin assembly (Mahajan et al., 1989). The distinct assembly mechanism and regulation utilized by Dictyostelium myosin II demonstrates that myosins from smooth muscle, striated muscle, and two types of amoebae form unique thick filaments by different pathways.     

Lime-Soda Softening Process Modifications for Enhanced NOM Removal

In this research, a number of process modifications to the lime-soda softening process were examined, including utilization of high Mg-content lime, addition of MgCl2, and the recycling of softening sludge, in order to improve the removal of natural organic matter (NOM) and reduce the formation of disinfection byproducts (DBPs). Jar test results showed that dissolved organic carbon (DOC) removal increased and trihalomethane (THM) formation was reduced as the magnesium in hydrated lime increased, and was directly correlated with the amount of magnesium removed from the system. However, a dolomitic quick lime hydrated under atmospheric conditions resulted in less effective DOC removal due to a lack of available Mg, and subsequently, less co-precipitation of Mg(OH)2-NOM complexes. The addition of MgCl2 to the raw water also increased DOC removal and reduced THM formation in both the presence and absence of softening sludge, with DOC removal increasing as softening sludge and magnesium dosages increased. As high as 43% removal of DOC was achieved at the stoichoimetric lime-soda ash dose in the presence of 457 mg/L sludge and 7.5 mg/L MgCl2, as compared to only 13% removal in the absence of sludge and MgCl2. The recycling of softening sludge had little or no effect on the hardness and the level of inorganic elements in treated water. The results presented here provide new approaches for improving DBP precursor removal during lime-soda softening without significantly increasing lime and soda ash dosage or the generation of waste sludge.     

水氯镁石复盐法脱水工艺

用烧瓶蒸馏装置模拟固定床脱水设备,研究用苯胺盐酸盐作脱水剂的复盐法将水氯镁石(MgCl2.6H2O)脱水制备无水氯化镁(MgCl2)的过程,考察复盐配料比例、保护气含水浓度、热解温度、热解时间等因素对所制备无水氯化镁产品中氧化镁和水分含量的影响,优化用固定床脱水工艺条件。结果表明,完全脱水和脱苯胺盐酸盐的最佳温度和时间分别是300℃与25～30 min和320～350℃与10～20min。最佳配料摩尔比C6H5NH2.HCl∶MgCl2.6H2O为1.1∶1.0。在最佳条件下,可得到合格的无水氯化镁产品MgCl298%～98.6%,MgO 0.03%～0.38%,苯胺盐酸盐0.10%～0.12%,水分0.60%～0.81%。     

Supramolecular self-assembly of d(TGG)4, synergistic effects of K+ and Mg2+

Spectral evidence indicates that molar concentrations of K+ can induce aggregate formation in d(TGG)4. The 320-nm turbidity monitoring indicates that more than 1 M KCl is needed for the onset of aggregation to occur at 20 degrees C within the time span of 24 h. The kinetic profile is reminiscent of autocatalytic reactions that consist of a lag period followed by accelerative and levelling phases. Progressive shortening of lag periods and more rapid accelerative phases accompany further increases in [K+]. Interestingly, the presence of Mg2+ greatly facilitates the aggregate formation and results in the prominent appearance of an intense psi-type CD. For example, whereas 1 M K+ fails to induce aggregate formation of d(TGG)4 within 24 h, the addition of 1 mM Mg2+ to a 1 M K+ solution is sufficient to induce the onset of aggregation in approximately 12 h. Furthermore, adjustment of the buffer to 16 mM Mg2+/1 M KCl reduces the lag time to less than 10 min and aggregation is nearly complete in 2 h. The requirement of [K+] for aggregation is reduced to 2 mM in the presence of 16 mM Mg2+, a reduction of nearly three orders of magnitude when compared to solutions without Mg2+. The effects of K+ and Mg2+ ions are synergistic, because the presence of 16 mM Mg2+ alone does not induce aggregate formation in this oligomer. Thermal stabilities of the aggregates are strongly dependent on the concentrations of these two ions. Although aggregates formed in the presence of 2 M KCl alone melt around 55 degrees C, those formed with added 16 mM Mg2+ melt at approximately 90 degrees C, with some aggregates remaining unmelted even at 95 degrees C. The slow kinetics of aggregate formation led to the appearance of gross hystereses in the cooling profiles. The interplay of these two ions appears to be specific, because the replacement of K+ by Na+ or the replacement of Mg2+ by other divalent cations does not lead to the observed self-assembly phenomenon, although Sr2+ can substitute for K+. A possible mechanism for the formation of self-assembled structures is suggested.     

Physicochemical properties of calcific deposits isolated from porcine bioprosthetic heart valves removed from patients following 2-13 years function

The purpose of this study was to characterize the physicochemical properties of calcific deposits that cause the failure of tissue-derived heart valve bioprostheses. This was done in an effort to understand the mechanism of pathologic biomineralization in the cardiovascular system and potentially prevent deterioration of bioprostheses. Calcific deposits taken from 10 failed bioprosthetic valves that had been implanted in patients for 2-13 years were characterized by chemical analysis, x-ray diffraction, FTIR spectroscopy, scanning electron microscopy, polarized light microscopy, and solubility measurements. The combined results identified the biomineral as an apatitic calcium phosphate salt with substantial incorporation of sodium, magnesium and carbonate. The average Ca/PO4 ratio for this "young" pathologic biomineral was approximately 1.3, considerably lower than approximately 1.7 found in mature atherosclerotic plaque biomineral and mature skeletal biomineral, both of which approximate hydroxyapatite in composition. Deproteinated calcific deposits from bioprostheses had thermodynamic solubilities comparable to those of both atherosclerotic plaque, typical pathologic biomineral and hydrolyzed octacalcium phosphate (OCP, Ca4H(PO4)3 x 2.5 H2O), a proposed precursor phase to biomineral apatite. This later finding, together with chemical composition and structural details of the bioprosthetic deposits themselves, supports a mechanism of cardiovascular calcification in which OCP plays a crucial role in the formation of the final apatitic phase. This suggests an approach toward prevention of bioprosthetic tissue calcification through control of the formation of the kinetically favored OCP precursor and/or its transformation into bioapatite.     

Teratogenicity study of magnesium chloride hexahydrate in rats

Teratogenicity of magnesium chloride hexahydrate (MgCl2.6H2O) was examined in rats. Magnesium chloride hexahydrate dissolved in distilled water was given to pregnant Wistar rats by gavage once a day from day 6 through 15 of pregnancy at doses of 0, 200, 400 and 800 mg/kg/day. The pregnant rats were sacrificed on day 20 of pregnancy and their fetuses were examined for malformation. Magnesium chloride hexahydrate caused no increased incidences of fetal malformation, and no toxic signs in the pregnant rats and the fetuses. It was concluded that magnesium chloride hexahydrate has no teratogenicity in rats when given by gavage. The no observed adverse effect level was estimated to be over 800 mg/kg/day for both pregnant rats and rat fetuses.     

Kinetics Effects on Preferential Struvite Accumulation in Wastewater

The kinetics of struvite (MgNH4PO4?6H2O) formation was studied to determine the cause of preferential accumulation at certain locations within the anaerobic digestion and postdigestion processes in wastewater treatment. Mechanisms controlling nucleation were also studied in laboratory induction time experiments. A field study of struvite accumulation on postdigestion process appurtenances was conducted to evaluate mechanisms affecting crystal growth rates. Struvite nucleation was found to be reaction-controlled, with nucleation strongly dependent on the struvite supersaturation level. Increased mixing intensity at constant supersaturation had a minor effect on induction times. The crystal growth rate was found to be transport-limited, with the growth rate a function of mixing energy input. Evaluation of the effect of pH change resulting from CO2 evolution in high turbulence areas showed that it has a minor influence on the crystal growth rate. Preferential struvite accumulation was found to be primarily controlled by growth kinetics, which was strongly dependent on mixing energy. Preferential accumulation of struvite on selected construction materials was also studied, and experimental evidence suggested that surface roughness exerted the greatest control over preferential struvite accumulation on various materials under similar mixing and supersaturation conditions.     

NaCl-KCl-MgCl_2熔盐中的CaF_2溶解行为及其对液镁汇集的影响

采用XRD、气冷阴极等手段对CaF_2在NaCl-KCl-MgCl_2电解质体系中的溶解行为及其对液镁汇集状况进行研究。研究表明,CaF_2在NaCl-KCl-MgCl_2熔盐溶解行为表现为CaF_2与MgCl_2反应生成CaCl_2和MgF_2,且随着CaF_2含量的添加,体系氟离子和钙离子浓度逐渐升高,但氟离子增加量相当较慢;CaF_2对液镁汇集的影响表现为与电解质中的MgO结合生成Mg_2OFCl和Mg_2OF_2,降低MgO对液镁汇集的影响,镁在阴极呈现大颗粒的片状结构;随着CaF_2添加量的增加,NaCl-KCl-MgCl_2熔盐体系表面张力逐渐降低,主要受反应产物CaCl_2生成的影响。     

Nucleation of hydroxyapatite by bone sialoprotein

Bone sialoprotein (BSP) and osteopontin, the major phosphorylated proteins of mammalian bone, have been proposed to function in the initiation of mineralization. To test this hypothesis, the effects of BSP and osteopontin on hydroxyapatite crystal formation were determined by using a steady-state agarose gel system. At low calcium phosphate concentrations, no accumulation of calcium and phosphate occurred in control gels or gels containing osteopontin. Gels containing BSP at 1-5 micrograms/ml, however, exhibited a visible precipitation band and significantly elevated Ca + PO4 contents. By powder x-ray diffraction, the precipitate formed in the presence of BSP was shown to be hydroxyapatite. These findings suggest that bone sialoprotein may be involved in the nucleation of hydroxyapatite at the mineralization front of bone.     

电解液浓度对CaTi4(PO4)6微弧氧化陶瓷膜生长过程的影响

通过微弧氧化技术一步合成了CaTi4(PO4)6生物陶瓷膜,并研究了电解液浓度对微弧氧化膜生长过程的影响.结果表明,在同一电解液浓度下,微弧氧化膜层质量的增加与时间呈线性关系.随着电解液浓度的增加,膜层的增重速度加快,形成时间缩短,击穿电压降低,且陶瓷膜表面Ca元素含量增加,P元素含量基本不变;但当电解液浓度增大到一定程度后再继续增加时,陶瓷膜表面的Ca、P元素含量逐渐降低.     

Growth of a turbidostat culture of yeasts in the presence of a high concentration of compounds in a stationary mode and under osmotic shock conditions

The growth of a turbidostat culture of the yeast strain Saccharomyces cerevisiae 14 was studied under steady-state conditions in the presence of high (1.2-3 M) concentrations of various soluble compounds and under transition conditions upon rapid concentration changes. The concentration was varied from the optimal one to that decreasing the growth rate two times and back to the optimal one. The effects of the following compounds on yeast growth in the steady-state regime were studied: glucose, sucrose, (NH4)2SO4, MgSO4, NaH2PO4, Na2SO4, KCl, NaCl, NH4Cl, MgCl2, and LiCl. To study transition processes, the following compounds were used: NH4Cl, NaH2PO4, MgCl2, MgSO4, (NH4)2SO4, KCl, and NaCl. When the NaCl concentration increased rapidly, the transition process was completed in the first generation. For the other compounds, this process lasted for several generations. The transition process was completed in the first generation in cultures previously inhibited by (NH4)2SO4 or NaH2PO4. Cell volume varied insignificantly among steady-state cultures grown under different conditions, except for those inhibited by NH4Cl or MgSO4. The level of glucose utilization increased with an increase in the concentrations of the compounds studied. The ratio of the duration of the juvenile developmental phases (prophase, metaphase) to the duration of the postjuvenile phases (anaphase, telophase) was constant over the concentration ranges tested. The inhibitory effect of a compound depended on its chemical nature rather than on its osmotic effect.     

Fluorescent monitoring of Jurkatt cell intracellular magnesium during metabolic poisoning

Divalent cation movement characterizes the final common pathway of cellular death from ischemic or metabolic injury. The influx of calcium is an essential step in cellular death. We hypothesized that intracellular magnesium levels may change during the progression to cellular death. Verapamil-sensitive changes in free ionized intracellular Mg2+ ([Mg2+[i) and Ca2+ ([Ca2+]i) levels were estimated in transformed T-lymphocytes exposed to metabolic inhibitors. Separate experiments used a Mg(2+)-sensitive fluoroprobe, fura-2 (Ex 1,344, Ex 2,376, Em 500), and a Ca(2+)-sensitive fluoroprobe, fura-2 (Ex 1,340, Ex 2,380, Em 510). Chemical anoxia (sodium cyanide 1 mM, iodoacetic acid 10 mM) caused a gradual increase in [Ca2+]i (control 126 +/- 13 nM) to > 1 mM by 10 min. This increase in [Ca2+]i was not affected by verapamil treatment. In separate experiments, [Mg2+]i levels were monitored during chemical anoxia. The specificity of mag-fura for Mg2+ over Ca2+ was reflected in the absence of a response to the lymphocyte Ca2+ mobilizer OKT-3. Uncorrected control [Mg2+]i levels (.4 +/- .1 mM) were not affected by the combined cyanide-iodoacetate treatment. A small increase in mag-fura-2 fluorescence was noted, probably due to binding of Ca2+ to the fluoroprobe when [Ca2]i exceeded 1 mM. Elimination of Ca2+ from the extracellular buffer increased the resting estimate of intracellular [Mg2+] to 1.6 + .1 mM. These results indicate that 1) extracellular Ca2+ can interfere with the fluorescent determination of intracellular magnesium concentration, and 2) intracellular free Mg2+ concentrations do not change in this cell line during chemical anoxia.     

Ionic requirements for RNA binding, cleavage, and ligation by the hairpin ribozyme

Metal ion requirements for RNA binding, cleavage, and ligation by the hairpin ribozyme have been analyzed. RNA cleavage is observed when Mg2+, Sr2+, or Ca2+ are added to a 40 mM Tris-HCl buffer, indicating that these divalent cations were capable of supporting the reaction. No reaction was observed when other ions (Mn2+, Co2+, Cd2+, Ni2+, Ba2+, Na+, K+, Li+, NH4+, Rb+, and Cs+) were tested. In the absence of added metal ions, spermidine can induce a very slow ribozyme-catalyzed cleavage reaction that is not quenched by chelating agents (EDTA and EGTA) that are capable of quenching the metal-dependent reaction. Addition of Mn2+ to a reaction containing 2 mM spermidine increases the rate of the catalytic step by at least 100-fold. Spermidine also reduces the magnesium requirement for the reaction and strongly stimulates activity at limiting Mg2+ concentrations. There are no special ionic requirements for formation of the initial ribozyme-substrate complex--analysis of complex formation using native gels and kinetic assays shows that the ribozyme can bind substrate in 40 mM Tris-HCl buffer. Complex formation is inhibited by both Mn2+ and Co2+. Ionic requirements for the ribozyme-catalyzed ligation reaction are very similar to those for the cleavage reaction. We propose a model for catalysis by the hairpin ribozyme that is consistent with these findings. Formation of an initial ribozyme-substrate complex occurs without the obligatory involvement of divalent cations. Ions (e.g., Mg2+) can then bind to form a catalytically proficient complex, which reacts and dissociates.(ABSTRACT TRUNCATED AT 250 WORDS)