Naja melanoleuca cobra venom contains two forms of complement-depleting factor (CVF)

Two forms of complement-depleting cobra venom factor (CVFm1 and CVFm2), possessing molecular masses of 142.6 kDa (CVFm1) and 143.1 kDa (CVFm2), according to MALDI mass-spectrometry, were isolated from the Naja melanoleuca cobra venom. As shown by polyacrylamide gel electrophoresis in the presence of SDS, both forms similarly to factor from the Naja kaouthia cobra venom (CVFk) consist of three polypeptide chains with molecular masses of about 70, 50, and 30 kDa, the two large subunits being glycosylated. As determined by MALDI mass-spectrometry, 30 kDa subunits of CVFm1 and CVFm2 have considerably different finger-prints of tryptic digests that suggests differences in their amino acid sequences. A study of activity in vivo has shown no significant differences in C3 consumption by CVFm1, CVFm2 and CVFk in mouse blood. However, as shown by an immunoassay method, they differ in their ability to activate the complement system via C3 conversion, the ratio of these activities for CVFm1:CVFm2:CVFk being 2.5:1.6:1. Kinetic studies using a hemolytic test showed that complement depletion by CVFm1 is faster than that by CVFm2. Thus, for the first time the presence in a single venom of two forms of CVF differing by both amino acid sequence and biological activity has been shown.     

A lethal cardiotoxic-cytotoxic protein from the Indian monocellate cobra (Naja kaouthia) venom

A lethal cardiotoxic-cytotoxic protein (mol. wt. 6.76 kDa) has been purified from the Indian monocellate cobra (Naja kaouthia) venom by ion-exchange chromatography and HPLC. CD spectra indicated the presence of 23% α helix, 19% β sheets and 35% coil. Complete amino acid sequence was determined by MALDI, which showed similar homology with cardiotoxins/cytotoxins isolated from venom of other Naja species. Intraperitoneal LD₅₀ was 2.5 mg kg⁻¹ in BalbC male mice. In vitro cardiotoxicity studies on isolated guinea pig auricle showed that the molecule produced auricular blockade that was abolished after trypsin treatment. Cytotoxicity studies on human leukemic U937 and K562 cells showed that it significantly inhibited cell proliferation in a dose and time dependent manner, as observed by trypan blue exclusion method and tetrazolium bromide reduction assay. IC₅₀ on U937 and K562 cells were 3.5 μg/ml and 1.1 μg/ml respectively. Morphometry and cell sorting studies indicated apoptosis induction in toxin treated leukemic cells. Apoptosis was caspase 3 and 9 dependent and the treated leukemic cells were arrested in sub-G1 stage. There was an increase in Bax-Bcl2 ratio, decrease in HSP (Heat shock protein) 70 and HSP90 and induction of PARP cleavage after NK-CT1 treatment. The toxin showed low cytotoxic effect on normal human leukocytes as compared with imatinib mesylate. Further detailed cytotoxic and cardiotoxic effects at the molecular level are in progress.     

Isolation and characterization of two toxins from the venom of the Malayan cobra (Naja naja sputatrix)

Two principal toxins of the Malayan cobra (Naja naja sputatrix) venom have been purified to electrophoretic homogeneity by successive SP-Sephadex C-25 ion exchange chromatography and Sephadex G-50 gel filtration chromatography. They are designated as sputa-neurotoxin 1 (SN1) and sputa-neurotoxin 2 (SN2), respectively. Both toxins belong to the group of short neurotoxins which are composed of approximately 60 amino acid residues. The ld₅₀ values following i.p. injection of the purified toxins are 91 (SN1) and 71 (SN2) μg/kg mouse. Amino acid compositions of the two toxins show close similarities to other postsynaptically acting curaremimetic cobra neurotoxins.     

Peripheral versus central action of a toxin from indian cobra (Naja naja naja) venom

A homogeneous toxin (NT fraction) purified from Indian cobra venom produced (at 0·25–1·0 mg/kg) predominant respiratory paralysis prior to any effect on the contractions of tibialis anterior muscle (cats) and gastrocnemius muscle (rabbits). NT fraction produced no direct effects on cardiovascular system or nerve action potential. The preferential action of the toxin on the respiratory process, over the tibialis anterior muscle twitches, was well differentiated at lower doses of NT fraction. Recovery from the respiratory and peripheral muscle paralytic effects was observed in cats assisted with artificial ventilation, suggesting that the toxin could be removed from the neuromuscular receptor sites, possibly by metabolic degradation. Phrenic nerve discharges and diaphragm contractions were unaffected at a time when animals stopped spontaneous breathing. In contrast, the electrical activity of the intercostal muscles disappeared at a faster rate, concurrent with the respiratory impairment. It is concluded that the effect of NT fraction on respiration is of peripheral origin and is possibly related to paralysis of the muscles of respiration among which the intercostal muscles appear to be the most susceptible. The neurotoxin does not seem to exert any direct effect on central respiratory mechanisms.     

Purification of two phospholipase A isoenzymes with anticoagulant activity from the venom of the cobra Naja naja siamensis

Two non-lethal phospholipases A with anticoagulant activity were purified from the venom of Naja naja siamensis by ion-exchange chromatography on Bio-Rex 70, gel filtration on Sephadex G-75 and ion-exchange chromatography on sulphopropyl-Sephadex C-25. They have 119 amino acid residues, seven disulfides and a very similar composition. Their sequences might differ only at two positions. The phospholipases can exist in three forms, as a monomer and two different aggregates, which behave differently in ion-exchange chromatography and short time analytical electrofocusing. A single phospholipase can therefore appear to be inhomogeneous. Upon preparative electrofocusing run for a long time each enzyme focused into a single zone with isolelectric points of 4·76 and 5·41, respectively.Basic phospholipases A are in general much more lethal than neutral or acidic ones. Some basic phospholipases are myotoxins and presynaptic neurotoxins, and it is proposed that the basic regions of the molecules facilitate the binding of these toxins to the negatively charged muscle and nerve membranes. A similar correlation does not exist with respect to the anticoagulant activity, since it was shown that a basic phospholipase A can be a powerful anticoagulant while another equally basic enzyme is a very poor one.Modification with p-bromophenacyl bromide suppresses both the phospholipase A and anti-coagulant activity. That does not necessarily mean that the anticoagulant activity is a consequence of the enzymic action, i.e. caused by the hydrolysis products or by destruction of essential phospholipids. Loss of anticoagulant activity may rather indicate that the modified protein is no longer capable of binding phospholipids. According to present hypothesis, phospholipases A inhibit coagulation by binding to phospholipids, which normally complex with many coagulation factors and thereby accelerate their activation.     

Unifying the mathematical modeling of in vivo and in vitro microdialysis

A unifying approach is presented for developing mathematical models of microdialysis that are applicable to both in vitro and in vivo situations. Previous models for cylindrical probes have been limited by accommodating analyte diffusion through the surrounding medium in the radial direction only, i.e., perpendicular to the probe axis, or by incomplete incorporation of diffusion in the axial direction. Both radial and axial diffusion are included in the present work by employing two-dimensional finite element analysis. As in previous models, the nondimensional clearance modulus (Θ) represents the degree to which analyte clearance from the external medium influences diffusion through the medium for systems exhibiting analyte concentration linearity. Incorporating axial diffusion introduces a second dimensionless group, which is the length-to-radius aspect ratio of the membrane. These two parameter groups uniquely determine the external medium permeability, which is time dependent under transient conditions. At steady-state, the dependence of this permeability on the two groups can be approximated by an algebraic formula for much of the parameter ranges. Explicit steady-state expressions derived for the membrane and fluid permeabilities provide good approximations under transient conditions (quasi-steady-state assumption). The predictive ability of the unifying approach is illustrated for microdialysis of sucrose in vivo (brain) and inert media in vitro, under both well-stirred and quiescent conditions.     

Synthesis,characterization, in vitro and in vivo evaluation of PEGylated oridonin conjugates

Oridonin (ORI), a diterpenoid compound with promising antitumor activity, was proved to possess potent antileukemia efficacies in vitro and in vivo recently. However, the development and application of ORI was limited by its poor solubility and rapid plasma clearance. The purpose of this study was to solve these problems. PEGylated oridonin linked with succinic acid (SA) as spacer moiety (PEG-SA-ORI conjugate) was synthesized. mPEG amines with four specifications of molecular weight (MW) were utilized. All polymeric conjugates showed satisfactory aqueous solubility and in vitro studies implied that the drug solubility and release features of conjugates were relevant to PEGs. The drug solubility increased more when the MW of PEG was lower, while more significant sustained-release effect was shown with higher PEG MW. Moreover, the release behaviors of conjugates showed a pH-sensitive property. In vivo pharmacokinetic studies demonstrated that the elimination half-life was prolonged in comparison with ORI solution. PEGylation could be a promising method to obtain better efficacy in the field of drug delivery system.     

Peroxidase-mediated biodegradation of carbon nanotubes in vitro and in vivo

As a result of their unique electronic, optical, and mechanical properties, carbon nanotubes (CNTs) have been implemented in therapeutic and imaging applications. In an idealized situation, CNTs would be disposed of after they transport their theranostic payloads. Biodegradation represents an attractive pathway for the elimination of CNT carriers post-delivery and may be integral in catalyzing the release of the cargo from the delivery vehicle. Accordingly, recent research efforts have focused on peroxidase-driven biodegradation of CNTs. In this review, we not only summarize recent efforts to biodegrade CNTs in the test tube, in vitro, and in vivo, but also attempt to explore the fundamental parameters underlying degradation. Encouraged by the in vivo results obtained to date, we envision a future, where carbon-based nano-containers, which are specifically designed to target organs/cells, deliver their cargo, and biodegrade via peroxidase-driven mechanism, will represent an attractive therapeutic delivery option in nanomedicine.     

Improvement of Malayan cobra (Naja naja sputatrix) antivenin

A low molecular weight toxic fraction was isolated from venom of the Malayan cobra (Naja naja sputatrix) by Sephadex G-50 gel filtration chromatography. The fraction accounted for almost 100% of the venom lethality. Antisera were prepared by immunizing rabbits with the low molecular weight toxic fraction, the glutaraldehyde-treated low molecular weight toxic fraction and the glutaraldehyde-treated, sea snake neurotoxin-enriched low molecular weight toxic fraction, respectively. Only the serum of rabbits immunized with the glutaraldehyde-treated, neurotoxin-enriched fraction gave effective protection against high doses of Malayan cobra venom. This antiserum is thus a potent Malayan cobra antivenin. One milliliter of this antiserum was able to neutralize 1.5 mg of Malayan cobra venom. It is thus 4 – 8 times more potent than the commercially available Malayan cobra antivenins produced by immunizing horses with whole Malayan cobra venom.     

Chitosan-graft-spermine as a gene carrier in vitro and in vivo

Chitosan has been proposed as a non-viral gene carrier because of its biodegradable and biocompatible cationic polymeric properties. However, the transfection efficiency of chitosan-DNA complexes is still too low for clinical trials. To improve transfection efficiency, we prepared a chitosan-graft-spermine (CHI-g-SPE) copolymer by an imine reaction between periodate-oxidized chitosan and spermine. The CHI-g-SPE copolymer was complexed with plasmid DNA in various copolymer-DNA weight ratios, and the complexes were characterized. The CHI-g-SPE copolymer showed good DNA binding ability and high protection of DNA from nuclease attack. The CHI-g-SPE/DNA complexes had well-formed spherical shapes and a nanoscale size with homogenous size distribution. The CHI-g-SPE copolymer had low cytotoxicity and CHI-g-SPE/DNA complexes showed transfection efficiency that was enhanced over that of chitosan-DNA. Furthermore, aerosol delivery of CHI-g-SPE/GFP complexes showed higher GFP expression compared with chitosan/GFP complexes, without toxicity. Our results indicate that the CHI-g-SPE copolymer has potential as a gene carrier.     

Assessing the anti-tumour properties of Iraqi propolis in vitro and in vivo

The study was designed to evaluate anti-tumour properties of Iraqi propolis collected from Mosul region (M) on HL-60 and HCT-116 cell lines and on HCT-116 in vivo. M induced an inhibitory effect against the proliferation of HL-60 and colony potential of HCT-116 cells. The apoptosis in HL-60 cells was associated with down-regulation of Bcl-2 and activation of Bax, while in HCT-116 cells, necrotic features were observed; size of cells was dramatically increased by swelling of cytoplasm and loss of membrane integrity, cell rupture and release of cellular contents. Analysis of BrdU/DNA cell cycle in both cell lines showed that M induced cell cycle perturbations in both BrdU positive and BrdU negative cells. The exposure of HL-60 to M caused γ-H2AX in a dose dependent manner and was associated with induction of apoptosis. The experiments in HCT-116 tumor-bearing mice showed that oral administration of propolis at doses that caused no detectable toxicity was associated with a decrease in mitotic cells and an increase in endoreduplications, increased p53 and decreased Ki-67 expression of cells in tumor sections. This study provides the rationale to investigate the potential beneficial effect of propolis in the diet of patients receiving anti-cancer therapies.     

Acrolein induces Alzheimer's disease-like pathologies in vitro and in vivo

The pathologic mechanisms of Alzheimer's disease (AD) have not been fully uncovered. Acrolein, a ubiquitous dietary pollutant and by-product of oxidative stress, can induce cytotoxicity in neurons, which might play an important role in the etiology of AD. Here, we examined the effects of Acrolein on the AD pathologies in vitro and in vivo. We found Acrolein induced HT22 cells death in concentration- and time-dependent manners. Interestingly, Acrolein increased proteins’ levels of amyloid precursor protein (APP), β-secretase (BACE-1) and the amyloid β-peptide transporter receptor for advanced glycation end products, and decreased A-disintegrin and metalloprotease (ADAM) 10 levels. In vivo, chronic oral exposure to Acrolein (2.5 mg/kg/day by intragastric gavage for 8 weeks) induced mild cognitive declination and pyknosis/atrophy of hippocampal neurons. The activity of superoxide dismutase was down-regulated while the level of malondialdehyde was up-regulated in rat brain. Moreover, Acrolein resulted in activation of astrocytes, up-regulation of BACE-1 in cortex and down-regulation of ADAM-10 in hippocampus and cortex. Taken together, our findings suggest that exposure to Acrolein induces AD-like pathology in vitro and in vivo. Scavenging Acrolein might be beneficial for the therapy of AD.     

In vitro and in vivo studies on some toxic effects of the venom from Hemiscorpious lepturus scorpion

The aim of this laboratory-based study was to investigate some of the toxic effects induced by the venom from Hemiscorpious lepturus (H. lepturus). For this aim, pharmacological, histological, biochemical methods as well as complete blood cell count were used to assess these toxic actions. In addition, in vitro haemolysis studies on human washed blood suspension and cytotoxicity on cultured fibroblasts were also undertaken. In vitro pharmacological test was made on rat isolated ileal segment. To this end, the effects of the venom on the contractile responsiveness to acetylcholine were recorded using F30 transducer and Darco chart recorder. For assessment of the haemolytic potency, varying concentrations (2, 10, 20 and 40 μg/ml) of the venom were added to 0.5 ml of 5% washed human blood and after 30 min, 2, 4, 8, 12 and 24 h of exposure, the degree of lysis (extent of redness developed in the supernatant solution after centrifugation) were measured by ELISA method. Cytotoxicity potential of the venom was assessed by trypan blue exclusion test. The venom (0.1, 1 and 10 μg/ml) was mixed with confluent fibroblast cell culture and the extent cytotoxicity was assessed microscopically. In vivo studies were conducted by a subcutaneous administration of sub-lethal dose (10 μg) of the venom and after 7 days the skin, at the site of injection, and kidney samples were stained by H & E method and examined microscopically. In addition, biochemical assessments including measurement of serum aspartate aminotransferase (AST), alanine transaminase (ALT), alkaline phosphatase (ALP) and amylase levels and urine analysis were made. The results showed that the venom prevented the relaxation phase of the acetylcholine-induced contractions on the isolated ileal segments and finally produced sustained spasmodic contractions. This spasmodic action was abolished by 1 μM atropine. The venom produced haemolysis of red blood cells in a concentration-dependent and duration-of-exposure manner, with 100% of haemolysis produced after 24 h following exposure to 40 μg/ml of venom. While cultured fibroblasts cells were more sensitive and disintegrated after 15 min of exposure to 1 μg/ml of the venom. Histological findings showed evidences of excessive inflammatory responses accompanied with signs of necrosis in the skin at the site of injection as well as structural damage in the nephrones. There was a significant rise in the serum enzymes. In addition, the number of the RBCs were reduced. The urine showed positive readings for proteinuria, blood and intact RBCs. The overall results suggest that the venom from H. lepturus primarily is a cytotoxic agent and has haemolytic, nephrotoxic and to some extent hepatotoxic activity.     

Disparate in vivo and in vitro immunomodulatory activities of rhodamine B

At nontoxic concentrations (1–10 μg/ml) rhodamine B (N-[9-(2-carboxyphenyl)-6-(diethylamino)-3 H-xanthen-3-ylidene]-N-ethylethanaminium chloride; D & C Red No. 19; CAS no. 81-88-9), a low-molecular-weight dye, suppressed the primary in vitro plaque-forming-cell response of BDF₁ mouse spleen cells both to the thymus-dependent antigen, sheep red blood cells and to the thymus-independent antigen, E. coli 0127:B8 lipopolysaccharide. Suppression was effective when the D & C Red No. 19 was added at any time up to 48 hr of culture. Rhodamine B at concentrations from 1 to 10 μg/ml significantly suppressed mitogen-induced proliferation of both B- and T-lymphocytes in vitro. BDF₁ mice exposed to 50 and 100 ppm dye in their drinking water for 14 days prior to splenectomy gave a depressed in vitro plaque-forming-cell response to sheep red blood cells. In contrast, a significant enhancement of antibody response resulted when mice were exposed to the dye and sheep-red-blood-cell antigen was administered in vivo. Mitogen-induced B- and T-lymphocyte proliferation was also enhanced in mice exposed to the dye for 14 days.     

Genotoxic evaluation of titanium dioxide nanoparticles in vivo and in vitro

With the extensive application of titanium dioxide (TiO₂) nanoparticles (NPs) in food industry, there is a rising debate concerning the possible risk associated with exposure to TiO₂ NPs. The purpose of this study is to evaluate the genotoxicity of TiO₂ NPs using in vivo and in vitro test systems. In vivo study, the adult male Sprague-Dawley rats were exposed to anatase TiO₂ NPs (75 ± 15 nm) through intragastric administration at 0, 10, 50 and 200 mg/kg body weight every day for 30 days. The γ-H₂AX assay showed TiO₂ NPs could induce DNA double strand breaks in bone marrow cells after oral administration. However, the micronucleus test revealed that the oral-exposed TiO₂ NPs did not cause damage to chromosomes or mitotic apparatus observably in rat bone marrow cells. In vitro study, Chinese hamster lung fibroblasts (V79 cells) were exposed to TiO₂ NPs at the dose of 0, 5, 10, 20, 50 and 100 μg/mL. Significant decreases in cell viability were detected in all the treated groups after 24 h and 48 h exposure. Significant DNA damage was only observed at the concentration of 100 μg/mL after 24 h treatment using the comet assay. The obvious gene mutation was observed at the concentration of 20 and 100 μg/mL after 2 h treatment using hypoxanthine-guanine phosphoribosyl transferase (HPRT) gene mutation assay. This study presented a comprehensive genotoxic evaluation of TiO₂ NPs, and TiO₂ NPs were shown to be genotoxic both in vivo and in vitro tests. The gene mutation and DNA strand breaks seem to be more sensitive genetic endpoints for the detection of TiO₂ NPs induced genotoxic effects.     

Effects of anticonvulsants on the in vivo and in vitro release of gaba

The actions of various anticonvulsant compounds on GABA release in vivo and in vitro were studied. An in vivo, superfusion of sensorimotor cerebral cortex was employed and drugs were administered either by intraperitoneal injection, or in superfusion fluid and release of endogenous amino acids was measured. The in vitro method involved superfusion of synaptosomes, with drugs dissolved in superfusate, with monitoring of the release of pre-loaded [U¹⁴C]-GABA. Two alkyl-GABA analogues, γ-acetylenic GABA and γ-vinyl GABA caused enhanced release of GABA to superfusate both in vivo and in vitro. However, phenobarbitone, diphenyl hydantoin, sodium n-dipropyl acetate and carbamazepine were without effect on GABA release in either test system. Taurine caused no detectable GABA release in vivo, or from purified synaptosomes in vitro, but did stimulate release in vitro, from crude synaptosome preparations containing mitochondria in large quantities, though histidine and leucine were equally effective.     

Extracts of Halenia elliptica exhibit antioxidant properties in vitro and in vivo

The antioxidant properties of different extracts of Halenia elliptica was investigated by employing various established in vitro systems. The results showed that various extracts possessed strong antioxidant activity in vitro, and the 70% methanol extract (ME) had the strongest antioxidant activity. Based on our in vitro results, ME was used for investigating the antioxidant properties of H. ellipticain vivo. The liver and kidney of CCl₄-intoxicated animals exhibited a significant decrease in superoxide dismutase (SOD), catalase (CAT) and glutathione (GSH) levels. Additionally, these organs exhibited a significant increase in the level of malondialdehyde (MDA). These changes were significantly reversed, in a dose-dependent manner, after treatment with ME and the standard treatment Vitamin E. Thus, it may be concluded that the ME possesses potent antioxidant properties, and might be valuable natural source of antioxidants that could be applicable to both the medical and food industries.     

Stability of cadmium resistance and metallothionein levels in vitro and in vivo

Cultured cells can be adapted to large concentrations of the toxic cadmium ion, apparently by induction of synthesis of the Cd-binding protein metallothionein (MT). One human epithelial line derived from normal skin (HE₁₀₀) and one murine fibroblast line, derived from L cells (C1 1D₁₀₀), were used to study the stability of Cd resistance, the MT levels following omission of Cd, and the inducibility of MT synthesis in cells on reexposure to Cd. In the murine cells there was no significant loss of resistance during a 4-week period either after cultivation in vitro or after growing the cells in nude mice. In the human cells a decrease (50%) in resistance was noted the first week after Cd omission. After removing Cd from the cells, a rapid decrease in MT content was demonstrated. After 3 weeks of cultivation only trace amounts were left in both cell lines. However, approximately 60% (HE₁₀₀) and 80% (C1 1D₁₀₀) of the previous levels were demonstrated after reexposure to maximum tolerable doses for 24 hr. The data indicate that the degree of stability of Cd resistance is dependent on the capacity in cells for an immediate de novo synthesis of large amounts of MT on reexposure to Cd. The animal experiments demonstrate that Cd resistance is maintained even after growing the cells in vivo.     

Hexavalent chromium inhibited the expression of RKIP of heart in vivo and in vitro

Background

Chromium (Cr) is considered to be a risk factor to the cardiovascular effects of fine particulate matter components to PM2.5 from traffic in highway patrol officers. RKIP (raf kinase inhibitor protein) is a physiological inhibitor of GRK-2 (G-protein-coupled receptor kinase 2) and affects β-adrenergic signaling and contractile activity in cardiomyocytes.

Objectives

In this study, we explored the change of RKIP in heart of chromium (VI)-exposed rats and cultured myocardial cells with chromium (VI) treatment.

Method

Wistar rats were divided into six groups which were chronically fed with 250, 500, 750, 1000, and 1250 ppm Na₂Cr₂O₇ and water for 60 days, respectively. Na₂Cr₂O₇ dose of 0.25, 0.5, 1.5, 3, 4.5, and 0 ppm (control group) was applied in cultured myocardial cells. The level of heart Cr (VI) was determined by electrothermal atomic absorption spectrometry. The expression of RKIP was measured by Western blot method. The MTT assay was used to measure the toxicity of myocardial cells with Cr (VI) treatment. The apoptosis test of myocardial cells was determined by caspase-3 colorimetric assay kit.

Result

The result showed that the expression of RKIP in heart (in vivo) and myocardial cells (in vitro) was decreased following Cr (VI) dose-dependent treatment.

Conclusion

We suggested that the decrement of RKIP of heart and myocardial cells with Cr (VI) treatment resulted in the function of cardiovascular system decreased.     

Formulation and in vitro/in vivo evaluation of levodopa transdermal delivery systems

This study aims to investigate the feasibility of Levodopa transdermal delivery systems (TDSs). Levodopa TDSs were formulated using various vehicles and permeation enhancers, and in vitro permeation and in vivo pharmacokinetic studies were carried out. In the in vitro study, ester-type vehicles showed relatively high enhancing effects; propylene glycol monocaprylate and propylene glycol monolaurate showed the highest permeation fluxes from both solution and pressure sensitive adhesive (PSA) TDS formulations. Lag time was dramatically shortened with PSA TDS formulations as compared with solution formulations. In the in vivo study, the addition of fatty acids increased blood drug concentrations regardless of the kind or concentration of fatty acid; the AUC_inf increased up to 8.7 times as compared with propylene glycol (PG) alone. PSA TDS containing 10% linoleic acid exhibited prolonged T_max as compared with oral form. Total clearance of L-dopa from PSA TDSs was significantly lower than from oral form (up to 86.8 times). Especially, PSA TDS containing 10% linoleic acid (LOA) revealed 76.2 fold higher AUC_inf than oral administration. Based on our results, the L-dopa PSA TDS containing PG with 10% LOA could be used as a good adjuvant therapy for Parkinson's disease patients who experience symptom fluctuation by L-dopa oral administration.