• Journal of Pharmaceutical Investigation
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• v.25 no.2
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• pp.101-108
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• 1995

The surface of albumin microspheres could be modified with methotrexate (MTX) by using 1,3-dicyclohexylcarbodiimide (DCC). Surface-modified albumin microspheres entrapping no MTX (SAMS), free MTX (SAMSF) and MTX-bovine serum albumin (BSA) conjugates (SAMSC) were prepared. respectively, and their release characteristics were investigated in the presence of trypsin using a dissolution tester. The mean diameters of all the microspheres were $5{\sim}8\;{\mu}m$, and their shapes was small and uniform. MTX bound tn their surfaces was released slower than the entrapped free MTX, and laster than the entrapped MTX-BSA conjugates. Also, surface-modified MTX was scarcely released in the absence of a proteolytic enzyme. Therefore, the surface-modified MTX may be released rapidly from SAMSC at the target site, and thereafter MTX may be released slowly from the encapsulated MTX-BSA conjugates in SAMSC for a long period.

• Archives of Pharmacal Research
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• v.27 no.8
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• pp.878-883
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• 2004

Tacrolimus (FK506), which is isolated from Streptomyces tsukubaensis, is a new potent immu-nosuppressant. Because of poor solubility in water, the conventional intravenous dosage forms of tacrolimus contain surfactants such as cremophor EL (BASF Wyandotte Co.) or hydroge-nated polyoxy 60 castor oil (HCO-60) which may cause adverse effects. This study relates to a polymer-tacrolimus conjugate, which can be dissolved in water, formed by chemically binding the sparingly soluble drug, tacrolimus, with the water soluble polymer, methoxypoly(ethylene glycol) (mPEG). Water soluble tacrolimus-mPEG conjugates have been synthesized and shown to be function in vitro as prodrugs. These conjugates are in the form of an ester wherein the 24-, 32- or 24,32-positions are esterified. The desired 24-, 32- or 24,32-esterified com-pounds were obtained by initially acylating of tacrolimus with iodoacetic acid at the 24-,32-, or 24,32-positions and then reacting the resulting acylated tacrolimus with a mPEG in the pres-ence of a base such as sodium bicarbonate. These conjugates were converted again into tac-rolimus by the action of enzymes in human liver homogenate, and the half-lives of the conjugates are approximately 10 min in the homogenate, indicating that the esterified tacroli-mus derivatives may be practically applicable as a prod rug for the immunosuppressant.

• Archives of Pharmacal Research
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• v.12 no.3
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• pp.186-190
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• 1989

The release rates of methotrexate (MTX) from MTX-human serum albumin (HSA) conjugate, and 5-fluorouracil (5-FU) from 5-FU acetic acid (AA)-HSA conjugate were determined after incubation of the conjugates in various conditions. The concentrations of 5-FU released from the conjugate increased monoexponentially, however those of MTX increased biexponentially in all studies. It indicated that there are two distinct types of MTX-HSA linkage, weakly and tightly bound linkages. The release rates of 5-FU were lower than those of MTX in all studies indicating that the bond of 5-FU-AA-HSA conjugate is very stable, which is supported by the higher value of activation energy (39. 9 vs 10. 7 Kcal/mole) using Arrhenius equation. The release rates of MTX and 5 -FU from the conjugates increased with incubation temperatures. Proteolytic enzyme and liver homogenates accelerated significantly the release rates of MTX and 5-FU. Approximately 1.30 and 22.0% of MTX were released after 12 hours of incubation in the absence and presence of protease, respectively. The corresponding values for 5-FU were released after 12 hours of incubation with rat liver homogenates which were diluted 6 times with phosphate buffer of pH 6.0. The MTX-HSA and 5-FU-AA-HSA conjugates were very stable in rat plasma.

• Archives of Pharmacal Research
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• v.27 no.5
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• pp.562-569
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• 2004

Self-assembling nanospheres of hydrophobized pullulan have been developed. Pullulan acetate (PA), as hydrophobized pullulan, was synthesized by acetylation. Carboxymethylated poly(ethylene-glycol) (CMPEG) was introduced into pullulan acetate (PA) through a coupling reaction using N, N'-dicyclohexyl carbodiimide (DCC). A synthesized PA-PEG-PA (abbreviated as PEP) conjugate was confirmed by Fourier transform-infrared (FT-IR) spectroscopy. Since PEP conjugates have amphiphilic characteristics in aqueous solution, polymeric nanoparticles of PEP conjugates were prepared using a simple dialysis method in water. From the analysis of fluorescence excitation spectra primarily, the critical association concentration (CAC) of this conjugate was found to be 0.0063 g/L. Observations by scanning electron microscopy (SEM) showed the spherical morphologies of the PEP nanoparticles. The particle size distribution of the PEP conjugates was determined using photon correlation spectroscopy (PCS) and the intensity-average particle size was 193.3 ${\pm}$ 13.53 nm with a unimodal distribution. Clonazepam (CNZ), as a model drug, was easy to entrap into polymeric nanoparticles of the PEP conjugates. The drug release behavior was mainly diffusion controlled from the core portion.

• Macromolecular Research
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• v.12 no.5
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• pp.534-539
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• 2004

Two biopolymers, silk fibroin (SF) and chitosan, were conjugated by tyrosinase (EC 1.14.18.1), a polyphenolic oxidase, to improve their physicochemical properties, such as their thermal properties and morphological stabilities in organic solvents. The crosslinking between SF and chitosan took place mainly through Michael addition reactions. A main reaction between the amino groups in chitosan and o-quinone, the oxidation product of the tyrosyl residue in SF, was confirmed by UV spectroscopy. Measurements of viscosity and light scattering indicated that the crosslinked SF/chitosan conjugate was compact: it had a smaller particle size because of tight bonding forces between the SF and chitosan molecular chains. Thermal decomposition of SF/chitosan conjugates crosslinked by tyrosinase occurred at higher temperatures. The adhesiveness of the SF/chitosan conjugates decreased steadily as the crosslinking reaction progressed. We propose that this new crosslinking method be used for the preparation of silk fibroin/chitosan conjugates using tyrosinase. We expect that SF/chitosan conjugates crosslinked by tyrosinase can be used preferentially in biomedical applications because of its unique properties and non-toxicity.

• Food Science of Animal Resources
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• v.42 no.5
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• pp.889-902
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• 2022

With increasing consumer demand for "clean label" products, the use of natural ingredients is required in the food industry. Protein/polysaccharide complexes are considered good alternatives to synthetic emulsifiers and stabilizers for formulating stable emulsion-based foods. Milk protein and carrageenan are widely used to improve the physical properties and stability of dairy food products. In a previous study, milk protein isolate (MPI) was conjugated with 𝛋-carrageenan (𝛋-Car) in a wet-heating system through the Maillard reaction, and the Maillard conjugates (MC) derived from MPI and 𝛋-Car effectively improved the stability of oil-in-water emulsions. Therefore, MPI/𝛋-Car conjugates were used in whipping cream as natural emulsifiers in this study, and the physical and rheological properties of whipping creams stabilized using MPI/𝛋-Car MC and MPI/𝛋-Car electrostatic complexes (EC) were investigated. The whipping creams stabilized with MPI/𝛋-Car MC have lower rheological parameters (η_a,50, K, G', and G'') than those of whipping creams stabilized with MPI/𝛋-Car EC. Although the overrun value was slightly reduced owing to the addition of MPI/𝛋-Car MC, the stability of the whipped creams with MC was effectively improved due to enhanced water-holding ability by conjugation.

• Archives of Pharmacal Research
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• v.23 no.1
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• pp.22-25
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• 2000

Formation of glutathione (GSH) conjugates with 2- or 6-(1-hydroxymethyl)- and 2-(1-hydroxyethyl)-DMNQ derivatives (DMNQ, 5,8-dimethoxy-1,4-naphthoquone was carried out in phosphate buffer (pH 7.4), in the presence of glutathione-S-transferase (GST), in rat liver S-9 fraction and by perfusion, and the rates of conjugates formation were compared and correlated to cytotoxicity. The GSH conjugates of 6-(1-hydroxyalkyl)-DMNQ derivatives were formed faster than 2-(1-hydroxyalkyl)-DMNQ derivatives under all of the media, implying that steric hindrance was the cause of lowering the rate of conjugate formation of 2-substituted derivatives. For both isomers, addition of GST did not improve the reaction rate, compared with that in buffer, while the reaction in the S-9 fraction and the perfusate was accelerated to a great extent. The catalytic effect of the S-9 fraction and the perfusate contain an effective system relaxing the steric hindrance of 2-(1-hydroxyalkyl)-DMNQ derivatives. Furthermore, a good correlation between the formation of the GSH conjugates and the cytotoxic activity of both naphthazarin isomers suggests that the steric hindrance is a cause of lowering the cytotoxicity of 2-isomers.

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.10
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• pp.5797-5804
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• 2013

In order to enhance the bioavailability of curcumin its conjugates with piperic acid and glycine were synthesized by esterifying the 4 and 4' phenolic hydroxyls, the sites of metabolic conjugation. Antiproliferative and apoptotic efficacy of synthesized conjugates was investigated in MCF-7 and MDA-MB-231 cell lines. $IC_{50}$ values of di-O-glycinoyl (CDG) and di-O-piperoyl (CDP) esters of curcumin were found to be comparable with that of curcumin. Both conjugates induced chromatin condensation fragmentation and apoptotic body formation. CDP exposure to MCF-7 cells induced apoptosis initiating loss of mitochondrial membrane potential (${\Delta}{\Psi}m$) followed by inhibition of translocation of transcription factor NF-${\kappa}B$ and release of Cytochrome-C. Reactive oxygen species (ROS) production was evaluated by fluorescent activated cell sorter. Change in ratio of Bcl2/Bclxl was observed, suggesting permeablization of mitochondrial membrane leading to the release of AIF, Smac and other apoptogenic molecules. DNA fragmentation as a hallmark for apoptosis was monitored by TUNEL as well as agrose gel electrophoresis. Thus, it was proven that conjugation does not affect the therapeutic potential of parent molecule in vitro, while these could work in vivo as prodrugs with enhanced pharmacokinetic profile. Pharmacokinetics of these molecules under in vivo conditions is a further scope of this study.

• The Korean Journal of Nuclear Medicine
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• v.14 no.1
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• pp.45-56
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• 1980

$T_3-BSA\;and\;T_4-BSA$ conjugates were prepared and identified spectrophotometrically. The ${\lambda}max$ of the conjugates was just coincided with that of BSA, but the molar extinction coefficients of the conjugates were generally larger than that of BSA itself. The molar ratios of $T_3:\;BSA\;and\;T_4:\;BSA$ in the prepared conjugates were found to be 9:1 and 5:1, respectively. The titers of the $T_3$ antisera were generally higher (max. $1.5{\times}10^4:1$) than those of $T_3$ (max. $2{\times}10^3:1$), and the average cross reactivity of the $T_3$ antibody with $T_3$ was lower (0.45%) than that of $T_4$ antibody with $T_3(3{\sim}4%)$. The results of the study indicate that the predominant cause of the lower titers and the lower specificity of the $T_4$ antisera comparing with those of $T_3$ is mainly due to the unstability of the $T_4-BSA$ and consequent degradation of the conjugate to $T_3-BSA$ during preparation, purification, and even during immunization. The lower molar ratio of $T_4$ to BSA in the preparation stage is also considered to be a minor factor. By measuring $T_3,\;T_4$ levels in the reference control serum, it has been confirmed that the prepared antisera can sufficiently be utilized, respectively, in the established radioimmunoassay systems.

• Archives of Pharmacal Research
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• v.15 no.2
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• pp.162-168
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• 1992

Release characteristics of five different types of hydrophilic albumin microspheres (HAM) containing different ratios of methotrexate-albumin (MTX-HSA) conjugates to free MTX: 1 : 0 (HAMC), 3 :1 (HAMC 3F), 1 :1 (HAMCF), 1:3 (HAMCF3) and 0 : 1 (HAMF) were investigated in the absence or presence of protease using dissolution tester. In all the HAMs studied except HAMC, the MTX was released bi-exponentially in the absence of protease; an initial fast release period up to approximately 6h, and thereafter the release rate was very much slower. The fast release of MTX from the HAMs (such as HAMC3F, HAMCF, HAMCF3 and HAMF) at the initial phase in probably due to the release of "physically associated" MTX from the core of the HAMs. The initial rate constants were 7.2, 8.7, 8.5 and 5.9 times greater than the second rate constants for HAMF, HAMCF3, HAMCF and HAMC3F, respectively. MTX release from HAMC was very slow and mono-phasic. It was at most 2.2% of the total entrapped amount by 24 h. The protease accelerated the release of MTX from the HAMs. The percentages of MTX released from HAMs up to 24 h were 100, 89.0, 75.0, 66.0 and 61.0% for HAMF, HAMCF3, HAMCF, HAMC3F and HAMC, respectively in the presence of protease and the corresponding values in the absence of protease were 30.2 19.0, 10.0, 6.5 and 2.2%, respectively. In vitro release of MTX in the presence of protease varied according to the ratios of MTX-HSA conjugates to MTX; the data set from HAMF, HAMCF3 and HAMCF fits better to monophasic first-order profile more adequately than to zero-order profile, that of HAMC3 monophasic first-order, and that of HAMC to bi-phasic zero-order. Above results suggested that zero-order release rate can be achieved by adjusting the ratio of MTX-HSA conjugates to MTX in the preparation of HAMs such as HAMC3F.as HAMC3F.