Antihepatotoxic and antioxidant activities of methanol extract and isolated compounds from Ficus chlamydocarpa

Free radicals, in particular radical oxygen species (ROS), play an important role in the aetiology and pathogenesis of various diseases. Current research in many countries focuses on the use of local medicinal plants as a promising source of liver protective agents. This paper describes the hepatoprotective effects of the methanol extract and four isolated compounds from Ficus chlamydocarpa on CCl4-induced liver damage, as well as the possible antioxidant mechanisms involved in this protection. The DPPH test, along with the beta-Carotene-Linoleic Acid Model System and Ferric-Reducing Antioxidant Power assays, as well as the inhibition of microsomal lipid peroxidation were used to measure radical-scavenging and antioxidant activities. Pretreatment of rats with the methanol extract of F. chlamydocarpa before CCl4 administration, significantly prevented serum increase of hepatic enzyme markers, glutamate oxaloacetate transaminase (GOT) and glutamate pyruvate transaminase (GPT), in a dose-dependent manner. The hepatoprotection was also associated with a significant enhancement in hepatic reduced glutathione (GSH) and a marked decrease of liver malondialdehyde (MDA). Among the four compounds 1-4, isolated from the methanol extract, alpha-amyrin acetate (1) and luteolin (4) showed a significant hepatoprotective activity, as indicated by their ability to prevent liver cell death and lactate dehydrogenase (LDH) leakage during CCl4 intoxication.     

Influence of various diet regimens on deterioration of hepatic function and hematological parameters following carbon tetrachloride: a potential protective role of natural honey

The objective was to assess the effects of commercial regular diet as control, total food restriction with honey, commercial regular diet with dextrose, or total food restriction with dextrose, on blood variables after carbon tetrachloride (CCl4) administration. Sprague Dawley albino rats were divided into four groups, 10 rats each; Group 1 rats were on commercial regular diet, Group 2 rats were on commercial regular diet with 50% dextrose, Group 3 rats were on total food restriction with 50% dextrose, and Group 4 rats were on total food restriction with 50% honey. Rats in all the groups were i.m. administered CCL4 (2.4 mL kg b. wt.-1). Blood tests including ALT, AST, serum albumin, serum protein, BUN, blood glucose (BG), hemoglobin (Hb), and white blood cell (WBC) were performed before CCl4 administration and repeated after 48 and 96 h of post-injection. In Group 1, CCl4 caused significant elevation in AST and ALT, and decrease in BS, WBC, and BUN; lower elevation in AST and ALT at 48 h and decreased AST and ALT at 96 h were obtained when dextrose was added to commercial regular diet (Group 2). Using dextrose alone (Group 3), though there was significant elevation of AST and ALT and decrease in BUN and WBC as compared to baseline values, significant decrease in ALT, AST, and BUN as compared to control was obtained. During absolute honey feeding (Group 4), elevation in AST and ALT obtained, following CCl4 administration was significantly less than the values obtained in all other groups; with lower elevation in AST and ALT as compared to baseline values. Honey increased serum albumin, serum protein, BG, and caused lower reduction in Hb. Conclusively, exclusive honey feeding (50% concentration) significantly modifies and ameliorates biochemical and hematological changes obtained after CCl4 injection.     

Pharmacokinetics of [6]-gingerol after intravenous administration in rats with acute renal or hepatic failure.

The pharmacokinetics of [6]-gingerol were investigated in rats with acute renal failure induced by bilateral nephrectomy, or those with acute hepatic failure induced by a single oral administration of carbon tetrachloride (CCl4), to clarify the contribution of the kidney and liver to the elimination process of [6]-gingerol. After bolus intravenous administration, a plasma concentration-time curve of [6]-gingerol was illustrated by a two-compartment open model. There was no significant difference in either the plasma concentration-time curve or any pharmacokinetic parameters between the control and nephrectomized rats. It is suggested, therefore, that renal excretion does not contribute at all to the disappearance of [6]-gingerol from plasma in rats. In contrast, hepatic intoxication with CCl4 elevated the plasma concentration of [6]-gingerol at the terminal phase. Its elimination half-life increased significantly, from 8.5 to 11.0 min, in CCl4-intoxicated rats. The extent of [6]-gingerol bound to serum protein was more than 90% and was affected very slightly by the CCl4-intoxication. These aspects indicate that [6]-gingerol is eliminated partly by the liver.     

Protective effects of various Chinese traditional medicines against experimental cholestasis

In a previous paper, we reported that methanol extracts obtained from 13 Chinese traditional medicines showed remarkable choleretic effects in normal rats. This paper examines the protective effects against experimental cholestasis induced by carbon tetrachloride (CCl4) or alpha-naphthylisothiocyanate (ANIT) in rats. No medicines, including sodium dehydrocholate and 1-phenylpropanol which are used clinically as choleretic drugs, inhibited the decrease of bile flow induced by CCl4. On the other hand, Intinko-to, Saiko-seikan-to and Bohu-tusyo-san revealed marked improvement of the dysfunction in bile secretion induced by ANIT. These three medicines inhibited the decrease of excretion of bile acid or bilirubin in the bile. They also exerted a protective effect against the alterations of serum components induced by ANIT, i.e., of glutamic oxaloacetic transaminase, glutamic pyruvic transaminase, alkaline phosphatase and the concentration of serum bilirubin. These results indicate that methanol extracts of Intinko-to, Saiko-seikan-to and Bohu-tusyo-san demonstrate not only choleretic effects but also improvement of cholestasis and liver parenchymal injury in rats.     

Increased transport of theophylline into gastrointestinal lumen and gastrointestinal dialysis by activated charcoal in rats with hepatic cirrhosis

The characteristics of exsorption and/or excretion of theophylline into the small intestinal lumen in rats with hepatic cirrhosis (HC rats) induced by carbon tetrachloride were investigated by an in situ single-pass perfusion technique. The serum concentrations of theophylline after i.v. administration of aminophylline (10 mg/kg) in the HC rats were significantly higher than those in normal rats during the experimental period. Moreover, the exsorption of theophylline from blood into the intestinal lumen was significantly increased in the HC rats compared with the normal rats. Treatments with oral activated charcoal reduced the serum theophylline levels in the HC rats. Consequently, gastrointestinal dialysis by oral administration of activated charcoal may be a useful method to remove poisonous drugs from the blood in patients with hepatic failure (including cirrhosis), which decreases the systemic clearance.     

Macromolecule-macromolecule interaction in drug distribution: effect of alpha-globulin concentration on the hepatic uptake of fractionated 3H-heparin by perfused rat liver.

The effect of alpha-globulin, the dominant binding protein for fractionated 3H-heparin, on the hepatic uptake of 3H-heparin was studied by liver perfusion experiments in rats. Fractionated 3H-heparin concentration in the recirculated perfusate decline one-exponentially with time for each of six initial concentration levels of alpha-globulin. The hepatic uptake clearance of fractionated 3H-heparin was 0.154 ml/min/g liver in the absence of alpha-globulin, and it decreased with increasing alpha-globulin concentrations. This result indicates that the hepatic uptake rate of alpha-globulin-bound fractionated 3H-heparin is lower than that of unbound fractionated 3H-heparin. On the other hand, it was indicated that almost all fractionated 3H-heparin binds to alpha-globulin at 8 mg/ml of alpha-globulin in in vitro study. However, the hepatic uptake clearance of the heparin at the concentration was of a certain value that could not be ignored. It was suggested that alpha-globulin-bound fractionated 3H-heparin also contributed to the hepatic uptake of fractionated 3H-heparin. Therefore, a protein-mediated transport system, which has been reported for some low molecular weight drugs, may also exist in the hepatic uptake of such a high molecular weight compound as fractionated 3H-heparin.     

Comparative evaluation of four different99mTc-sulfur colloid kits used for liver scanning

The distribution of four different commercially available A, B, C, D kits (^99mTc-sulfur colloid) for hepatoimaging was compared in mice by organ radioassay and in rabbits for blood clearance. The distribution of kits A and C (single step kits) was assessed in the human by blood clearance, external liver, spleen measurement, and scintillation camera imaging. Kit A reaches a high concentration in liver within 15–20 minutes with relatively high surrounding tissue background, and superior spleen scintiphotos. However, when kit C was used, a high activity concentration in the liver was reached within 10–15 minutes with low tissue background and faint visualization of the radiotracer in the spleen. Blood clearance of the four^99mTc-sulfur colloids was determined in rabbits. The data obtained indicated that the four hepatoagents exhibit rapid blood clearance but the initial decrease of blood activity curve of kit D was relatively faster than the other three hepatic agents. The biodistribution is similar for the four^99mTc-S-colloids but the blood retained higher activity residue using kit A compared with others. The formation of^99mTc-sulfur colloid using kits B, D (multistep kits) involves many steps after the addition of^99mTcO ₄ ^– to the reagent. These procedures are time consuming, required facilities at the medical institutions and give rise to the radiation exposure. While single step kits A and C have the same diagnostic value, the use of kit C allows a reduction of absorbed radiation, which may be useful in the liver exploration in children.     

Technetium-99m-diethyl-IDA instant kit: Labelling kinetics and biological properties

A formulation of stannous-diethyl-IDA freeze-dried kit, containing 50 mg diethyl-IDA and 0.4 mg hydrated stannous chloride, to be labelled with technetium has been developed for hepatobiliary scintigraphy. Gel chromatography column scanning technique has been applied for determination of technetium fractions in the preapration. The optimal pH value of the preparation with a high hepatobiliary specificity was found between 5.5 to 6.0. Effect of ligand to metal ratio on the stability of the prepared Sn-diethyl-IDA solution prior mixing with technetium has been investigated. The reaction conditions (initial pH) between the ligand and stannous ion affects the percent hydrolysis of the labelled compund, i.e. the formation rate of the complex. The organ distribution data of^99mTc-diethyl-IDA in mice for 60 minutes post injection were satisfactory. The radiopharmaceutical exhibits rapid blood clearance, great hepatic clearance and very short hepatocyte transit time. Uptake of the radiopharmaceutical in various organs of mouse at 5 minute post injection showed that the greater part of the injected radioactivity has distributed between liver and intestine. However, about 12% and 60% of the injected dose has been found in liver and intestine respectively. The renal uptake of the HB agent in mice is relatively low and decreases with time to become about 0.3% at 60 minutes post injection. Blood clearance data of the radiopharmaceutical kit in rabbits showed that the HB agent is-rapidly cleared, since the initial decrease was very fast with a half-time of 1.5 minutes.     

Protective effects of a neutral polysaccharide isolated from the mycelium of Antrodia cinnamomea on Propionibacterium acnes and lipopolysaccharide induced hepatic injury in mice

Mycelia of Antrodia cinnamomea were extracted with chloroform and hot water. A neutral polysaccharide named ACN2a separated from the water extract was purified using 10% CCl3COOH, and repeated column chromatography on HW-65 and DE-52 cellulose. Its structure was determined by chemical and spectroscopic analyses. ACN2a was composed of Gal, Glc, Fuc, Man and GalN (in the ratio 1:0.24:0.07:0.026:faint), in which an alpha-D-(1-->6)-Gal linkage accounted for 73% of all linkages. The ratio of branch points was about 16% of the total residual numbers, and branches were attached to C-2 of galactosyl residues of the main chain. ACN2a had an average molecular weight of 12.9x10(5) Daltons, [alpha]D25=+115 degrees (c=0.44, H2O); [eta]=0.0417dl.g-1, Cp=0.2663 cal/(g. degrees C). The hepatoprotective effect of ACN2a was evaluated using a mouse model of hepatic injury that was induced by Propionibacterium acnes (P. acnes) and lipopolysaccharide (LPS). The administration of ACN2a (0.4, 0.8 g/kg/d, p.o.), significantly prevented increases in serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) enzyme activities in mice treated with P. acnes-LPS, indicating hepatoprotective activity in vivo.     

Increase in the plasma protein binding of weakly basic drugs in carbon tetrachloride-intoxicated rats.

Plasma protein binding of weakly basic drugs such as propranolol and quinidine was determined in rats with carbon tetrachloride (CCl4)-induced hepatic disease. Free fractions of propranolol and quinidine in the plasma of rats at 24 h after CCl4-intoxication were decreased by 41 and 30%, respectively, compared to those of control rats. An addition of Tris (butoxyethyl) phosphate (TBEP), a specific displacer for basic drugs from alpha 1-acid glycoprotein (AGP), to the plasma increased the free fractions of the basic drugs, resulting in no difference in the extent of the plasma free fraction of each drug between control and CCl4-intoxicated rats. Plasma concentration of AGP in CCl4-intoxicated rats was elevated 2.7-fold of that in control rats at 24 h after the CCl4 intoxication and reached a peak of 4.8-fold elevation at 48 h. A regression analysis revealed a high degree of positive correlation between ratios of bound to free fraction of propranolol and plasma concentrations of AGP. These results suggest that the plasma protein binding of the basic drugs was increased mainly due to the rise in the plasma AGP concentration in CCl4-intoxicated rats.     

Electron-impact ionization of CCl4 and CCl2F2

Absolute partial and total cross sections for electron-impact ionization of CCl4 and CCl2F2 are reported for electron energies from threshold to 1000 eV. The product ions are mass analyzed using a time-of-flight mass spectrometer and detected with a position-sensitive detector whose output demonstrates that all product ion species are collected with equal efficiency irrespective of their initial kinetic energies. Data are presented for production of CCl3(+), CCl2(+), CCl+, C+, Cl2(+), and CCl3(2+) from CCl4; and for production of CCl(2)F+, CClF2(+), CClF(+), (CCl+ + CF2(+)), Cl+, CF+, F+, and C+ from CCl2F2. Data are also reported for formation of (CCl2(+),Cl+) and (CCl+, Cl+) ion pairs from CCl4. The total cross section for each target is obtained as the sum of the partial cross sections. The overall uncertainty in the absolute cross sections for most of the singly charged ions is +/- 5-7 %. The present partial cross sections for lighter fragment ions are found to be considerably greater than had been previously reported but the most recent total cross section measurements agree well with those reported here. Neither the binary-encounter-Bethe theory nor the Deutsch-Mark theory reproduces the experimental cross sections correctly for both targets.     

固本活血壮骨颗粒对实验性肝纤维化大鼠肝细胞的保护作用

为探讨固本活血壮骨颗粒对实验性肝纤维化大鼠肝细胞的保护作用,采用CCl4致大鼠肝纤维化模型,观察固本活血壮骨颗粒对肝功能及肝细胞损伤超微结构的影响。结果表明,固本活血壮骨颗粒明显改善肝细胞变性及炎性细胞浸润等情况,同时明显降低血清转氨酶活性、升高血清白蛋白,降低白蛋白与球蛋白比例并使之接近正常;电镜观察表明固本活血壮骨颗粒对实验性肝纤维化大鼠肝细胞线粒体等细胞器的损伤有保护作用。提示固本活血壮骨颗粒明显减轻四氯化碳所致的肝细胞损伤,保护肝细胞,改善肝功能。     

An evaluation of predicting postoperative residual liver function using 99mTc tin colloid

The rate of clearance (K value) of 99mTc tin colloid in the liver differentiates normal subjects from liver cirrhosis patients; so 99mTc tin colloid is as useful as 198Au colloid as a marker of liver function. There are several reports concerning volume estimation using liver scintigraphy. Our original method was devised to measure the effective liver volume by scintigraphy. By combining the K value with effective liver volume, a predictive index was obtained in order to predict the residual liver function before hepatic resection. The index in 24 patients with liver diseases was investigated before hepatic resection. Three of them died due to hepatic failure after hepatic resection. The indices were between 0.40 and 0.45 in two of these three patients and 0.338 in one. Among the patients without hepatic failure, the indices showed more than 0.45 in 19 patients and between 0.40 and 0.45 in two. These results indicate that the limitation of hepatic resection is between 0.40 and 0.45 of the predictive index.     

Antioxidant activity of new aramide nanoparticles containingredox-active N-phthaloyl valine moieties in the hepatic cytochrome P₄₅₀ system in male rats

We report the synthesis of aramide nanoparticles containing a chiral N-phthaloyl valine moiety and their antioxidant activities on hepatic contents of cytochrome P???, amidopyrene N-demethylase, aniline-4-hyroxylase and induced the hepatic content of cytochrome b5 and nicotinamide adenine dinucleotide phosphate (NADPH) cytochrome C-reductase. Polymers were obtained as well-separated spherical nanoparticles while highly aggregated particles via H-bonding organization of the aramide-containing pyridine led to a thin layer formation. The effects of the nanoparticles and CCl? on enzyme activities and thiobarbituric acid reactive substances (TBARS) levels of male rat liver were studied. Pretreatments of rats with the polyamides prior to the administration of CCl? decreased the hepatic content of the tested enzymes. Doses reduced the toxic effects exerted by (?CCl?) upon the liver through inhibition of the cytochrome P??? system. Inhibition of such metabolizing enzymes could reduce the carcinogenic effects of chemical carcinogens.     

Fully automated determination of a new anthracycline N-l-leucyldoxorubicin and six metabolites in plasma by high-performance liquid chromatography with on-line sample handling.

N-l-Leucyldoxorubicin (Leu-Dox) was developed as a prodrug of doxorubicin (Dox) in order to diminish the cardiotoxic side-effect associated with repeated anthracycline treatment. To study the pharmacokinetics of Leu-Dox, Dox and other metabolites a sensitive and selective assay was needed. Leu-Dox and six of its known metabolites were extracted from plasma using an in-line reversed-phase precolumn (40-50 microns C8 particles). The trapped analytes were subsequently flushed to the analytical column (3 microns C18) using 0.5 ml of phosphate buffer (pH 3.5)-acetonitrile (2:1, v/v), which also served as the isocratic mobile phase. Within 12 min, a clean baseline-resolved chromatogram is obtained by fluorescence detection. Recoveries were almost quantitative and highly reproducible, with standard deviations less than or equal to 5.4% and less than or equal to 2.7% at spiked concentrations of 10 and 100 nM. Using 300 microliters of plasma, detection limits ranged from 0.3 to 0.8 nM at a signal-to-noise ratio of 3. The calibration curves were linear from 1 to 300 nM (r2 greater than or equal to 0.999) for each of the seven compounds. The between-day accuracy was in the range 91-99% and 99-105% at 10 and 100 nM, respectively, with standard deviations of 1-4%. Application of the assay to the analysis of plasma from patients after administration of Leu-Dox proved successful.     

A thermodynamic approach, using speciation studies, towards the evaluation and design of bone-seeking radiopharmaceuticals as illustrated for 117mSn(II)-PEI-MP

Blood plasma modeling has proved effective in the evaluation of clinical observations recorded for baboon and rat tests with ¹⁵³Sm- ethylenediaminetetraphosphonic acid (EDTMP) as well as for ¹⁶⁶Ho-EDTMP. In the search for a cure for metastatic bone cancer, ^117mSn with its conversion electrons of discrete energies shows low bone marrow toxicity, providing the opportunity to increase the administered dose. Selective accumulation in lesions would capitalize on this advantage. The 10-30 kDa fraction of the water-soluble polymer polyethyleneimine, functionalized with methylene phosphonate groups (PEI-MP) and labeled with ^99mTc, has shown selective uptake into bone tumours. This paper relates the speciation of Sn(II)-PEI-MP and other known ^117mSn(II) containing bone-seeking radiopharmaceuticals in blood plasma. Apparent formation constants for the complexation of Sn^II with PEI-MP, DTPA, HEDP and other important blood plasma ligands were measured potentiometrically or estimated by linear free energy relationships (LFER). These data were added to the ECCLES database in order to construct a blood plasma model for Sn^II. From this model it is predicted that Sn^II will remain bound to the polymer (PEI-MP) in blood plasma and therefore, have only slight reticuloendothelial uptake. Preliminary primate studies indeed proved that the complex between Sn^II and PEI-MP remains intact in blood plasma, which is consistent with the observation for PEI-MP labeled with ^99mTc. From these data, it was also possible to explain in retrospect the lower bone uptake, the slow blood clearance and the liver uptake of the agents ^117mSn(II) DTPA and ^117mSn(II) HEDP agents as reported in the literature.     

Solvent dependence of absorption intensities and wavenumbers of the fundamental and first overtone of NH stretching vibration of pyrrole studied by near-infrared/infrared spectroscopy and DFT calculations

Near-infrared (NIR) and IR spectra were measured for pyrrole in CCl(4), CHCl(3), and CH(2)Cl(2) to study solvent dependence of absorption intensities and wavenumbers of the fundamental and first overtone of NH stretching vibration. It was found that the wavenumbers of the NH fundamental and its first overtone decrease in the order of CCl(4), CHCl(3), and CH(2)Cl(2), which is the increasing order for of the dielectric constant of the solvents. Their absorption intensities increase in the same order, and the intensity increase is more significant for the fundamental than the overtone. These results for the solvent dependence of the wavenumbers and absorption intensities of NH stretching bands of pyrrole are quite different from those due to the formation of hydrogen bonds. Quantum chemical calculations of the wavenumbers and absorption intensities of NH stretching bands by using the 1D Schr?dinger equation based on the self-consistent reaction field (SCRF)/isodensity surface polarized continuum model (IPCM) suggest that the decreases in the wavenumbers of both the fundamental and the overtone of the NH stretching mode with the increase in the dielectric constant of the solvents arise from the anharmonicity of vibrational potential and their intensity increases come from the gradual increase in the slope of the dipole moment function.     

Synthesis of (S)-N-[methyl-11C]nicotine and its regional distribution in the mouse brain: a potential tracer for visualization of brain nicotinic receptors by positron emission tomography.

A nicotine agonist, 11C-labeled (S)-nicotine, was synthesized by N-methylation of (S)-nornicotine with [11C]-methyl iodide in dimethylformamide-dimethylsulfoxide in order to study nicotinic receptors in the human brain by positron emission tomography. The radiochemical yield of this N-methylation reaction was more than 90% within 5 min. After purification by high performance liquid chromatography the radiochemical purity of the product was more than 99% and the specific radioactivity was 7.4-11.1 GBq/mumol. The regional distribution of (S)-[11C]nicotine in the mouse brain after intravenous injection was compared with that of (R)-[11C]nicotine. After injection of (S)-[11C]nicotine, the regional uptake of radioactivity was in the following order: cortex greater than thalamu approximately hippocampus greater than striatum greater than hypothalamus greater than cerebellum. Moreover, (S)-[11C]nicotine was displaced from the brain by unlabeled (S)-nicotine, but unlabeled (R)-nicotine caused no change in uptake. In contrast, (R)-[11C]nicotine showed a lower brain uptake and lesser regional differences in radioactivity.     

Technetium-99m dextran kit: formulation and biological behaviour

A new formulation of stannous-dextran (Sn-Dx) freeze dried kit, containing 60 mg dextran (Dx-70) and 0.08 mg SnCl₂·2H₂O, to be labelled with^99mTc, has been developed. At pH 6.5–7.0. the labelling efficiency was greater than 95%. Gel chromatography column scanning technique was applied for radiochemical purity determination of^99mTc-Dx preparation and the degree of in vivo plasma protein binding. Not less than 70% of the administered activity was bound to plasma and remained constant over a 1h period. The biological behaviour after intravenous injection of^99mTc-Dx kit was characterized by high and efficient yield of the radiopharmaceutical. The preliminarly clinical results on normal subjects showed that the radiopharmaceutical could be a useful agent for scintigraphy of leg lymph vesel, pelvic and inguinal lymph nodes. The activity uptake in liver and kidney (60 min) was relatively very low, whereas the urinal bladder activity (30 min) represents the drainage of the activity entering the blood stream after interdigital injection of^99mTc-Dx.     

New BATO complexes for brain and myocardial imaging

Two new BATO complexes^99mTcC1 (dmg)₃BC₆H₄CH₃ (Cholor[bis {2,3-butanedionedioxime(1-)-O} {2,3-butanedionedioximato (2-)-N,N',N",N'",N",N"'} (m-tolueneborato) technetium]) and^99m TcCl (4-MCDO)₃MeB, ([bis{4-methyl-1,2-cyclohexanedioximato (1-)-O} {4-methyl-cyclohexane-1, 2-dione-dioximato (2-)-O} {methyl-borato (2-)-N,N', N", N'", N", N"'} chlorotechnetium]), generally called BATO (Boronic Acid Adducts of Technetium Dioximes, had been synthesized and evaluated for potential use in brain and myocardial perfusion imaging. Their labeling conditions were also investigated. In their biodistribution studyies they showed higher radiochemical stability and rapid brain uptake and myocardial uptake in mice. After i.v administration, the first complex had 0.87%ID in the brain and 1.02% ID in the heart at 2 min and it had a longer retention in brain (0.62%ID was maintained at 15 min postinection) but rapidly cleared from heart (0.33%ID postinjection). For the second complex, it showed very rapid blood clearance. The uptake of heart, lung and blood in mice at 2 min respectively were: 1.32%ID, 2.48%ID and 6.66%ID. These two complexes formation were 2rapid, simple and of high yield(91%). The processes were easy to kit formulation.