Levodopa Concentrations Research Articles

LPS and hypoxia alters the production/release of catecholamines in Immortalized Human Mesangial Cell

Catecholamines (CA)‐dopamine (DA), L‐Dopa, epinephrine (EP) and norepinephrine‐are involved in filtration and renal vasoconstriction processes, and can be produced by mesangial cells (MC). MC expresses Toll Like Receptor 4, that detects lipopolysaccharide (LPS). LPS and hypoxia, both present in sepsis, can lead to acute renal failure. The aim was to asses the effect of LPS and hypoxia on the production/release of CA in immortalized human mesangial cell (IHMC). IHMC were cultured in DMEM and incubated with LPS (0.1, 1.0, 5.0 and 100 μg/mL) for 2 hours, or exposed to hypoxia for 30 min. CA were extracted from the medium and cell lysates was quantified by HPLC‐ED. Tyrosine hydroxylase (TH) expression ‐ step‐limiting enzyme ‐ was evaluated by Western blotting. LPS at 1 μg/mL induced a decline in intracellular L‐Dopa level as compared with control (5.3±1.6 vs.10.8±1.0 pg/mg protein). The concentration of 5 μg/mL decreased the intracellular level of L‐Dopa (4.9±1.3 vs. 10.3±1.0 pg/mg protein) and increased EP concentration (8.8±1.9 vs. 2.2±0.5 pg/mg protein). LPS at 100 μg/mL increased the levels of intracellular EP (102.7±8.4 vs.65.8±10.1 pg/mg), L‐Dopa (48.5±2.4 vs.22.2±4.1 pg/mg protein) and DA (24.9±2.9 vs.15.9±1.4 pg/mg protein) and decreased the concentration of EP (125.2±13.7 vs.205.4±10.4 pg/mg protein) and L‐Dopa (34.6±5.7 vs.57.4±8.6 pg/mg protein) in extracellular. TH expression was significantly affected by LPS (100 μg/ml), however it did not occurr under hypoxia condition. These results suggest that LPS can influence the production/release of CA in IHMC.Support by CNPq.

New drug delivery strategies for improved Parkinson's disease therapy

Increasing interest has been addressed toward the introduction of new therapeutic approaches to obtaining continuous dopaminergic stimulation (CDS). The goal of this therapeutic strategy is to reduce the occurrence and severity of L-DOPA (LD)-associated motor fluctuations and dyskinesia, and provide good long-term safety and tolerability. CDS can be achieved by the administration of oral dopamine (DA) agonists with a long half-life, transdermal or subcutaneous delivery of DA agonists, or intestinal LD infusion. To allow higher concentrations of LD to reach the brain and to reduce peripheral side effects, the therapeutic approach provides the concomitant administration of LD, carbidopa and entacapone that have been developed in tablet form, standard LD/carbidopa, LD/benserazide, LD/entacapone, LD/tolcapone associations or long-acting controlled release formulations, LD/carbidopa and LD/benserazide. Alternatively to solid formulations, LD/carbidopa liquid forms have been developed. Furthermore, the authors examine a series of new LD codrugs and non-dopaminergic drugs for Parkinson's disease treatment, together with a variety of experimental delivery strategies including transdermal therapeutic systems, liposomes, solid lipid nanoparticles and biocompatible microparticles. This review provides an overview of progress in anti-Parkinson therapy, mainly focused on delivery strategies and codrug approach for treatment of this neurological disorder.

Comparison of pharmacokinetic profile of levodopa throughout the day between levodopa/carbidopa/entacapone and levodopa/carbidopa when administered four or five times daily

To compare plasma levodopa concentrations after repeated doses of levodopa/carbidopa/entacapone (LCE) and levodopa/carbidopa (LC). Open-label, randomized, two-period, active-controlled, cross-over study with four dosing regimens: groups I and II (healthy volunteers and Parkinson's disease patients) received levodopa 100 mg or 150 mg four times daily, respectively, and groups III and IV (healthy volunteers) received the same strengths of levodopa five times daily. Pharmacokinetic (PK) parameters determined for levodopa included Cmin, Cmax, Cmax - Cmin, AUC, t(1/2), and tmax. In healthy volunteers and PD patients, mean trough levels (Cmin), Cmax, and AUC of levodopa were, in general, significantly higher during LCE compared to LC administration. Compared to Cmin, Cmax, and AUC, differences between the treatments in variability of levodopa concentrations (Cmax - Cmin) were less consistent. The present results on the differences in levodopa PK between LCE and LC provide a basis to evaluate the relationship of levodopa PK and the induction of motor complications in an on-going study in early Parkinson's disease using similar dosing regimens.

Effect of a slow release preparation of levodopa on Parkinson's disease in combination with a peripheral decarboxylase inhibitor

Plasma concentrations of levodopa were determined after therapeutic oral levodopa-carbidopa doses. The wide fluctuations observed in plasma levodopa levels could be considerably reduced by the addition of a slow release levodopa preparation. This kind of combination medication was given to 15 parkinsonian patients, whose earlier therapy had proved inadequate. With the combination medication, levodopa-carbidopa, on an average 420 mg/42 mg combined with 950 mg of levodopa in slow release form, a statistically significant improvement in parkinsonian signs could be achieved without any worsening of the side effects. The results suggest that parkinsonian patients may tolerate much higher daily levodopa doses if the fluctuations in plasma levels of the drug can be diminished.

Genetic polymorphism of catechol O-methyltransferase and pharmacokinetics of levodopa in healthy Chinese subjects

We studied the distribution of catechol O-methyltransferase (COMT) genotypes in the Chinese Fujian Han population and explored the potential effect of COMT genetic polymorphism on the pharmacokinetics of levodopa. Polymerase chain reaction (PCR) was employed in the COMT genotype analysis of 166 volunteers. After a single oral dose of levodopa/benserazide, the plasma concentration of levodopa was determined by high-performance liquid chromatography (HPLC) with electrochemical detection (ECD). In the 166 subjects, the frequencies of G/G, G/A and A/A COMT genotypes were 58.4%, 36.7% and 4.9%, respectively. The frequency of the homozygous A/A genotype was much lower than in caucasians and Southwest Asians. COMT activity of erythrocytes in the G/A genotype group was significantly lower than in the G/G genotype group but significantly higher than in the A/A genotype group. There was no significant difference in pharmacokinetic parameters, including t(1/2alpha), t(1/2beta), AUC(0-infinity), CL/F, C(max), t(max) and V/F. The frequency of COMT genotypes in the Chinese Fujian Han population, which is related to COMT enzyme activity, is significantly different from that in caucasians and Southwest Asians. The pharmacokinetic characteristics of levodopa in healthy Chinese subjects may not be dependent on their COMT genotype status. From the 166 volunteers, 5 G/G, 5 G/A and 4 A/A genotype male subjects were recruited for the study of levodopa pharmacokinetics.

The Effect of Entacapone on Levodopa Rate of Absorption and Latency to Motor Response in Patients With Parkinson Disease

The aim of the study was to explore the potential effect of the catechol-O-methyltransferase inhibitor entacapone coadministration on the rate of absorption and matched latency to motor response of an oral test dose of levodopa/benserazide in the treatment of Parkinson disease (PD). Fourteen PD patients (Hoehn and Yahr stages 2-2.5) entered the study protocol. They underwent an oral levodopa instrumental kinetic-dynamic test on 2 occasions, 4 weeks apart, according to an intrasubject open comparative design: at the first examination, while receiving their usual levodopa/benserazide therapy, and at the second one after a 4-week entacapone (200 mg) dosing concomitantly with usual first morning levodopa/benserazide intake. Entacapone coadministration was associated with a median 15-minute lengthening (P < 0.05) of time to peak levodopa plasma concentration compared with levodopa/benserazide alone but failed to have a statistically significant effect on matched latency to drug motor effect. Levodopa peak plasma concentration was nearly identical in the 2 treatments. Overall, the delayed latency to single-dose peak plasma levodopa concentration on entacapone cotreatment was modest and of negligible clinical significance in a cohort of moderately affected PD patients.

Influences of levodopa on adipose tissue and skeletal muscle metabolism in patients with idiopathic Parkinson’s disease

The substantial weight loss in Parkinson's patients may be related to direct influences of levodopa treatment on fat mobilization/oxidation. We assessed systemic and local metabolic responses to levodopa/benserazide in patients with idiopathic Parkinson's disease. We studied 10 Parkinson's disease patients and examined adipose tissue and skeletal muscle metabolism directly with microdialysis. We monitored dialysate concentrations of ethanol, glucose, lactate, pyruvate, and glycerol to assess tissue blood flow and metabolism before and after levodopa/benserazide intake. We also conducted in vitro studies on adipocytes from healthy women. Levodopa/benserazide increased serum levodopa, 3,4-dihydroxyphenylacetic acid (DOPAC), and norepinephrine (P < 0.01). Serum adipose tissue and skeletal muscle glycerol did not change or decreased. Adipose tissue glycerol was inversely correlated with serum levodopa concentrations (P < 0.05). In isolated adipocytes, levodopa attenuated isoproterenol-induced glycerol release (P < 0.05). Levodopa/benserazide elicits pronounced metabolic changes in both adipose tissue and skeletal muscle with a switch from lipid to carbohydrate metabolism. In adipose tissue, levodopa/benserazide failed to activate lipolysis. Therefore, we suggest that levodopa/benserazide does not induce fat wasting through direct and acute influences on adipose tissue metabolism.

New L‐Dopa Codrugs as Potential Antiparkinson Agents

This paper reports the synthesis and preliminary evaluation of new L-dopa (LD) conjugates (1 and 2) obtained by joining LD with two different natural antioxidants, caffeic acid and carnosine, respectively. The antioxidant efficacy of compounds 1 and 2 was assessed by evaluating plasmatic activities of superoxide dismutase (SOD) and glutathione peroxidase (GPx) in the rat. Rat striatal concentration of LD and dopamine (DA), and central nervous effects were evaluated after oral administration of the codrugs 1 and 2. The results suggest that, though our codrugs are devoid of significant antioxidant activity, they are able to induce sustained delivery of DA in rat striatum and can improve LD and DA release in the brain.

Determination of levodopa in pharmaceutical preparations by irreversible biamperometry

Based on the electrocatalytic oxidation of levodopa at gold electrode and the reduction of permanganate at platinum electrode, a novel flow injection irreversible biamperometric method is developed for the determination of levodopa under the potential difference of 0 V imposed between two electrodes. In H 2SO 4 solution, the linear relationships between currents and the concentrations of levodopa are obtained in the range from 0.04 mg/L to 20 mg/L with the detection limit of 0.012 mg/L. The proposed method is applied to the determination of levodopa in pharmaceutical preparations.

Development of a sensitive flow‐injection–chemiluminescence detection method for the determination of laevodopa

A simple chemiluminometric method using flow injection has been developed for the determination of levodopa, based on its sensitizing effect on the weak chemiluminescence (CL) reaction between Na(2)SO(3) and acidic KMnO4. Under optimum experimental conditions, the CL intensity was linearly related to the concentration of levodopa from 3.4 x 10(-8) to 2.4 x 10(-5) mol/L and the detection limit was 1.1 x 10(-8) mol/L (s:n = 3). The relative standard deviation (RSD) of the proposed method calculated from 20 replicate injection of 3 x 10(-7) mol/L levodopa was 3.3%. The correlation coefficient was 0.997. The method was successfully applied to the determination of levodopa in commercial pharmaceutical formulations and spiked urine samples.

Nutritional characterization of Mucuna pruriens: 1. Effect of maturity on the nutritional quality of botanical fractions and the whole plant

This study was conducted to determine the stage of maturity at which the dry matter (DM) yield and nutritive value of velvet bean ( Mucuna pruriens) is optimized. Mucuna was harvested at 77, 110 and 123 days after planting (DAP) from quadruplicate 5 m × 1 m plots within each of 6 blocks. At each DAP, DM yield, chemical composition, botanical composition, in vitro rumen fluid-pepsin DM digestibility (IVDMD) and concentrations of total polyphenols, l-dopa and tannins were determined on the whole plant and botanical fractions. Whole-plant Mucuna DM yield increased ( P<0.01) linearly with maturity; proportions of leaves and stems decreased linearly ( P<0.01), whereas proportion of pods increased ( P<0.01). Concentrations of neutral-detergent fiber (aNDF) in whole plant, leaf, and stem increased ( P<0.05), or tended ( P<0.10) to increase linearly with maturity, as did the acid-detergent fiber concentration of leaves and stems. Maturity decreased ( P<0.05) ether extract concentrations of leaves linearly, and stems quadratically, but increased ( P<0.05) whole-plant and pod starch concentrations. Pods contained relatively high concentrations of lysine, histidine, phenylalanine, aspartate, glutamate, leucine, isoleucine, and valine, but low concentrations of methionine and cystine. The essential amino acid index did not vary with maturity. Most minerals in Mucuna are concentrated in the leaves and the whole plant contains sufficient Ca, P, K, Mg, Fe, Cu, Na, Mo, Mn, and Zn for growing sheep, although their bioavailability of these minerals is unknown. Total polyphenol concentration quadratically ( P<0.01) increased with maturity in the whole plant, tended to increase ( P<0.10) in pods, linearly ( P<0.01) decreased in stems and fluctuated in leaves. Maturity quadratically increased l-dopa concentration of the whole plant ( P<0.05) and stems ( P<0.01), but did not affect those of leaves and pods. Maturity quadratically increased ( P<0.05) total tannin concentration in the whole plant, but decreased ( P<0.10) that of pods. The l-dopa was concentrated in the seeds and pods of mature (110–123 DAP) plants, but tannins were concentrated in leaves and stems. Whole-plant IVDMD was not affected by maturity, but digestible DM yield linearly ( P<0.01) increased with increasing DM yield. There was a 2-week harvest window (110–123 DAP) during which whole-plant crude protein and IVDMD remained unchanged. Nevertheless, harvesting at 123 DAP gave the best combination of biomass yield and nutritive value.

Nutritional characterization of Mucuna pruriens: 4. Does replacing soybean meal with Mucuna pruriens in lamb diets affect ruminal, blood and tissue l-dopa concentrations?

The harmful effects of the 3,4-dihydroxy- l-phenylalanine ( l-dopa) in Mucuna pruriens (Velvet bean) seeds have limited its use as a protein supplement for monogastrics and humans. Little is known about the extent of metabolism of Mucuna l-dopa in ruminants or its accumulation in ruminant tissues consumed as food by humans. This study aimed to determine if replacing soybean meal (SB) with Mucuna increases concentrations of l-dopa in rumen fluid, blood and muscle tissue of lambs. Twenty-seven RM lambs (RM; initial body weight, BW = 33.8 ± 5.44 kg) and 12 Florida Native (FN; initial BW = 24.9 ± 8.63 kg) lambs were assigned to four treatments and fed a basal diet of coastal bermudagrass hay, corn grain, and liquid molasses for 42 (FN) or 49 days (RM) and then slaughtered. Dietary supplements were formulated by substituting 0 (SB), 330 (Lo), 670 (Med) or at least 1000 (Hi) g/kg of SB with rolled Mucuna seeds ( l-dopa, 24 g/kg dry matter, DM). Body weight was measured weekly and carcass characteristics and concentrations of l-dopa in rumen fluid, blood and the sterno-mandibularis muscle were measured at slaughter when concentrations of blood urea N, glucose, haptoglobin and cerruloplasmin were also measured. Lambs fed SB had higher (P<0.05) average daily gain than those fed Mucuna (0.20 versus 0.15 kg/day for RM; 0.21 versus 0.14 kg/day for FN). However, concentrate protein source did not affect dressing and concentrations of blood urea N, or blood glucose. Feeding the Hi diet versus the SB diet did not increase concentrations of blood cerruloplasmin, or l-dopa or concentrations of l-dopa metabolites in blood. No l-dopa was found in the ruminal fluid of the lambs and l-dopa concentrations in the sterno-mandibularis muscle were low ( i.e., <5 ng l-dopa/g), and unaffected (P>0.05) by diet. Ingested Mucuna l-dopa was extensively metabolized in lambs, and did not accumulate to toxic levels in muscle tissues.

Nutritional characterization of Mucuna pruiriens: 2. In vitro ruminal fluid fermentability of Mucuna pruriens, Mucunal-dopa and soybean meal incubated with or without l-dopa

Three in vitro experiments were conducted to determine the rumen fermentability of Mucuna (M) pruriens (24 g 3,4-dihydroxy- l-phenylalanine ( l-dopa)/kg dry matter (DM) and soybean meal treated with (SBD) or without (SB) 138 g l-dopa/kg DM). Additional objectives were to determine if l-dopa inhibits rumen fermentation, and if ruminal microbes can adapt to l-dopa or M. In Experiment 1, ground (1 mm) substrates were incubated in triplicate at 38 °C in 9 ml nutrient media and 1 ml rumen fluid in a series of six, 48 h, consecutive batch cultures. The first culture was inoculated with rumen fluid from two donor cows. Subsequent cultures were inoculated with fluid (1 ml) from the previous culture. The DM digestibility (DMD, 616 g/kg vs. 540 g/kg; P<0.01) and gas production (51.7 ml/g vs. 44.2 ml/g DM; P<0.05) were higher from fermentation of M versus SB but similar for SB and SBD (540 g/kg vs. 554 g/kg and 44.2 ml/g DM vs. 43.5 ml/g DM, respectively). The slopes of the relationships between DMD (g/kg) or gas production (ml/g DM) and fermentation period were not reduced by fermenting M (−0.014 DMD slope; 2.28 gas production slope) or SBD (−0.014 DMD slope; 0.459 gas production slope), instead of SB (−0.002 DMD slope; 1.039 gas production slope), indicating microbial adaptation to M and SBD. Total volatile fatty acid concentration (VFA; 53.7, 54.9 and 54.9 mmol/l) and molar proportions of VFA were similar among substrates. Gas production kinetics of M versus SB (Experiment 2), and SB versus SBD (Experiment 3) were also measured after substrates were incubated in triplicate in buffered rumen fluid for 24 h using a non-linear exponential model to fit the data. Residual l-dopa was measured after separate fermentation of substrates in triplicate for 0, 4, 8, 16 and 24 h. Fermentation of M versus SB produced more ( P<0.05) gas (250 ml/g vs. 100 ml/g DM) and total VFA (203 mmol/l vs. 180 mmol/l) and a lower ( P<0.05) acetate:propionate ratio (1.35 vs. 1.87; P<0.05). Adding l-dopa to SB increased ( P<0.01) gas production (92 ml/g DM vs. 200 ml/g DM), and total VFA concentration (132 mmol/l vs. 188 mmol/l), but reduced ( P<0.05) gas production rate (0.08 ml/h vs. 0.05 ml/h). The concentration of l-dopa in fermented M and SBD decreased by 53 and 47%, respectively during fermentation. In conclusion, M was more fermented than SB and degradation of l-dopa during ruminal fermentation and microbial adaptation to l-dopa were confirmed. Adding l-dopa to SB did not impair ruminal fermentation.

Enteral Levodopa/Carbidopa Infusion in Advanced Parkinson Disease

In patients with advanced Parkinson disease, levodopa/carbidopa formulated as a gel suspension (Duodopa) permits continuous delivery into the small intestine using a portable pump, resulting in less variability in levodopa concentrations and fewer motor fluctuations and dyskinesias than with oral levodopa administration. This is a retrospective analysis of the long-term clinical experience with this agent. All but 1 of the patients who had received enteral levodopa infusion treatment between January 1, 1991, and June 30, 2002, consented to a review of their hospital charts. Of the 65 patients with initial testing of the treatment, 86% opted for continued treatment via percutaneous endoscopic gastrostomy or gastrojejunostomy. Total exposure to levodopa infusion was 216 patient-years (mean, 3.7 years). Maximum treatment duration was 10.7 years. Fifty-two patients were treated for 1 year or longer. The adverse effect profile of levodopa/carbidopa infusion was similar to that observed with oral administration of levodopa. Seven deaths occurred, all considered unrelated to the treatment. Intestinal tube problems, including dislocation of the intestinal tube to the stomach, were the most common technical problem, occurring in 69% of the patients during the first year. The optimal daily dose of levodopa decreased by an average of 5% during follow-up. The safety of enteral infusion of levodopa/carbidopa formulated as a gel suspension was found acceptable. For most patients, the technical challenges posed by the enteral infusion system were offset by the improvement in motor fluctuations and dyskinesias offered by this technique.

Inhibition of glycogen synthase kinase-3 reduces l-DOPA-induced neurotoxicity

The neurotoxicity of l-3,4-dihydroxyphenylalanine ( l-DOPA), used for the treatment of Parkinson's disease, remains controversial. Although there are many reports suggesting that long-term treatment of l-DOPA causes neuronal death, an increasing body of recent evidence has proposed that l-DOPA might be neuroprotective rather than neurotoxic. We investigated the effect of l-DOPA on neuronally differentiated PC12 (nPC12) cells by treating cells with various concentrations of l-DOPA for 24 h. We also studied whether glycogen synthase kinase (GSK)-3 activation is related to l-DOPA-induced neurotoxicity by simultaneously treating cells with several concentrations of l-DOPA and a GSK-3 inhibitor for 24 h. MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) assay, trypan blue staining, cell counting kit-8, and DAPI staining all showed that l-DOPA decreased nPC12 cell viability at high concentrations. In addition, 100 μM l-DOPA treatment significantly increased the activity of GSK-3 and death signals including cytochrome c, activated caspase-3 and cleaved PARP, and decreased survival signals including heat shock transcription factor-1 in a concentration-dependent manner. Treatment with GSK-3 inhibitor VIII or lithium chloride prevented l-DOPA-induced cell death. Together, these results suggest that l-DOPA induces neuronal cell death at high concentrations and that the neurotoxic effect of l-DOPA might be mediated in part by GSK-3 activation.

이온토포레시스를 이용한 levodopa의 경피전달: electroosmosis 및 electrorepulsion의 역할

The objective of this work is to study transdermal delivery of levodopa using iontophoresis and evaluate various factors which affect the transdermal transport. Levodopa is unstable in aqueous solution, and, in order to establish a stable condition for levodopa for the duration of experiment, we investigated the stability of levodopa in aqueous solutions of different pHs with/without the addition of dextrose or the application of current. Using stable aqueous solution, we have studied the effect of pH, polarity and penetration enhancer (ethanol) on transdermal flux and compared the results. We also investigated the iontophoretic flux from hydroxypropyl cellulose (HPC) hydrogel. In vitro flux study was performed at , using side-by-side diffusion cell. Full thickness hairless mouse skin and rat skin were used for this work. Current densities applied were 0.4 or and current was off after 6 hour application. Stability study showed that levodopa solution with a pH 2.5 or 4.5 maintained the initial concentration of levodopa for 24 hours with the addition of 5% dextrose. However, at pH 9.5, levodopa was unstable and 30 to 40% of levodopa degraded within 24 hours, even with the addition of 5% dextrose. Hydrogel swollen with dextrose added levodopa solution maintained about 97% of the initial concentration of levodopa for 13 days, when stored in . The application of current did not affect the stability of levodopa in hydrogel. Flux study from levodopa solution with pH 2.5 showed that cathodal delivery of levodopa was higher than passive or anodal delivery. When the pH of the donor solution was 4.5, anodal delivery of levodopa was higher than passive or cathodal delivery. These results seem to indicate that electroosmosis plays more dominant role than electrorepulsion in the flux of levodopa at pH 2.5, and the reverse situation applies for pH 4.5. The passive flux was unexpectedly high for the ionized levodopa. Similar to the results from aqueous solution, cumulative amount of levodopa transported trom HPC hydrogel by cathodal delivery was significantly higher than passive or anodal delivery. The treatment of 70% ethanol cotton ball by scrubbing increased passive, anodal and cathodal flux, with the largest increase for anodal flux. These results indicate that iontophoretic delivery of zwitterion such as levodopa is much complicated than that can be expected from small ionic molecules with single charge. The results also indicate that the balance between electroosmosis and electrorepulsion plays a very important role in the transport through skin.

Simultaneous determination of levodopa and carbidopa by synchronous fluorescence spectrometry using double scans

A synchronous fluorescence spectrometric method is described for the simultaneous determination of binary mixtures of levodopa and carbidopa in pharmaceutical formulation and urine sample, without prior separation steps, using two scans. At Δ λ = 30 nm, only carbidopa yields a detectable signal that is independent of the presence of levodopa. Similarly, at Δ λ = 65 nm the signal of levodopa is not influenced by the presence of carbidopa. Signals at two wavelengths, 288 nm (Δ λ = 30 nm) and 281 nm (Δ λ = 65 nm), vary linearly with carbidopa and levodopa concentrations over the range 0.019–1.971 μg mL −1 (for levodopa) and 0.022–2.262 μg mL −1 (for carbidopa), respectively. The correlation coefficients for the standard calibration graphs were 0.9962 and 0.9951 ( n = 10) for carbidopa and levodopa, respectively. The limits of detection (LOD estimated as per IUPAC recommendations) were 0.01 and 0.006 μg mL −1 for carbidopa and levodopa, respectively. The method was successfully applied to the determination of levodopa and carbidopa in pharmaceutical formulation and urine sample. The recovery results were satisfactory.

Estimation of L-Dopa from Mucuna pruriens LINN and Formulations Containing M. pruriens by HPTLC Method

A selective, precise, and accurate high-performance thin-layer chromatographic (HPTLC) method has been developed for the analysis of L-dopa in Mucuna pruriens seed extract and its formulations. The method involves densitometric evaluation of L-dopa after resolving it by HPTLC on silica gel plates with n-butanol-acetic acid-water (4.0+1.0+1.0, v/v) as the mobile phase. Densitometric analysis of L-dopa was carried out in the absorbance mode at 280 nm. The relationship between the concentration of L-dopa and corresponding peak areas was found to be linear in the range of 100 to 1200 ng/spot. The method was validated for precision (inter and intraday), repeatability, and accuracy. Mean recovery was 100.30%. The relative standard deviation (RSD) values of the precision were found to be in the range 0.64-1.52%. In conclusion, the proposed TLC method was found to be precise, specific and accurate and can be used for identification and quantitative determination of L-dopa in herbal extract and its formulations.

Inhibitory Mechanism of Melanin Synthesis by Glutathione

Glutathione dose dependently inhibited melanin synthesis in the reaction of tyrosinase and L-DOPA. The inhibition of melanin synthesis was recovered by increasing the concentration of L-DOPA, but not recovered by increasing tyrosinase. Glutathione inhibited the binding between tyrosinase and L-DOPA. Although the synthesized melanin was aggregated within 1 h, the aggregation was inhibited by the addition of glutathione. These results indicate that glutathione inhibits the synthesis and agglutination of melanin by interrupting the function of L-DOPA.

Simultaneous determination of serum concentrations of levodopa, dopamine, 3-O-methyldopa and α-methyldopa by HPLC

Levodopa is the medication of choice for Parkinson's disease. The biological complexity of levodopa and of its main derivatives makes their determination important in the clinical field. The aim of this study was to develop an HPLC method for the simultaneous determination of serum concentrations of levodopa, dopamine, 3-O-methyldopa and α-methyldopa. We compared UV and fluorimetric detection of native and derivatised compounds. Though less sensitive than other methods, UV detection is important to exclude naturally fluorescent, interfering substances. Fluorimetric detection of derivatised compounds is more sensitive than UV detection. Since 3-O-methyldopa does not react with the derivatising agent 1,2-diphenylethylenediamine, it cannot be detected. For simultaneous determination of the four compounds after pharmacological treatment of patients we therefore advise fluorimetric detection of the native compound.