Overcoming the challenge of potent endogenous interferences in limaprost quantification: An innovative methodology combining differential mobility spectrometry with LC-MS/MS for ultra-high sensitivity, selectivity and significantly enhanced throughput.

Limaprost, an orally administered analogue of prostaglandin E1, possesses potent vasodilatory, antiplatelet, and cytoprotective properties. Due to its extremely low therapeutic doses and exceedingly low plasma concentrations, the pharmacokinetic and bioequivalence studies of limaprost necessitate a highly sensitive quantitative method with a sub-pg/mL level of lower limit of quantification. Moreover, the intensity of endogenous interferences can even exceed the maximum concentration level of limaprost in human plasma, presenting further challenge to the quantification of limaprost. As a result, existing methods have not yet met the necessary level of sensitivity, selectivity, and throughput needed for the quantitative analysis of limaprost in pharmacokinetic and bioequivalence investigations. This study presents a new methodology that combines differential mobility spectrometry (DMS) with liquid chromatography-tandem mass spectrometry (LC-MS/MS) and utilizes a distinctive strategy to achieve more accurate DMS conditions. This integration yields a method that is currently the most sensitive and features the shortest analytical time, making it the sole technique capable of meeting the requirements for limaprost pharmacokinetic and bioequivalence investigations. This method demonstrates robustness and is successfully employed in a pharmacokinetic investigation of limaprost in human subjects, underscoring that the combination of DMS with LC-MS/MS serves as an efficacious strategy for overcoming the challenges inherent in analyzing biological samples afflicted by multiple interferences.

In Vivo Targeted Metabolomic Profiling of Prostanit, a Novel Anti-PAD NO-Donating Alprostadil-Based Drug.

Prostanit is a novel drug developed for the treatment of peripheral arterial diseases. It consists of a prostaglandin E1 (PGE1) moiety with two nitric oxide (NO) donor fragments, which provide a combined vasodilation effect on smooth muscles and vascular spastic reaction. Prostanit pharmacokinetics, however, remains poorly investigated. Thus, the object of this study was to investigate the pharmacokinetics of Prostanit-related and -affected metabolites in rabbit plasma using the liquid chromatography-mass spectrometry (LC-MS) approach. Besides, NO generation from Prostanit in isolated rat aorta and human smooth muscle cells was studied using the Griess method. In plasma, Prostanit was rapidly metabolized to 1,3-dinitroglycerol (1,3-DNG), PGE1, and 13,14-dihydro-15-keto-PGE1. Simultaneously, the constant growth of amino acid (proline, 4-hydroxyproline, alanine, phenylalanine, etc.), steroid (androsterone and corticosterone), and purine (adenosine, adenosine-5 monophosphate, and guanosine) levels was observed. Glycine, aspartate, cortisol, and testosterone levels were decreased. Ex vivo Prostanit induced both NO synthase-dependent and -independent NO generation. The observed pharmacokinetic properties suggested some novel beneficial activities (i.e., effect prolongation and anti-inflammation). These properties may provide a basis for future research of the effectiveness and safety of Prostanit, as well as for its characterization from a clinical perspective.

Physiological interrelationships between the venous shunts of penile corpus spongiosum and cavernous compartments during erection used in the pharmacotherapy of erectile dysfunction.

AIM OF THE STUDY: Alprostadil (A) induces smooth-muscle relaxation by stimulating the increasing of intracellular cAMP. Intra-urethral administration is an alternative to its intra-cavernose injection in inducing erection. In order to understand the functional and anatomical basis for drug transfer, the possible communications of the corpus spongiosum with the corpora cavernosa were studied. MATERIAL OF STUDY: A "Spongiogram" (SP) was performed in 44patients (pt.) by injection of radiological contrast into the glans. The SP visualized drainage of the glans into the deep dorsal vein, as well as that of the spongiosum into the circumflex veins, which in turn drained into the deep dorsal vein. RESULTS: In 34pt. (77,2%) filling of the corpora was also visualized; in 10pt.(22.8%), with ectasia of the deep dorsal vein, such a filling was not visualized. DISCUSSION: Filling of the corpora cavernosa, in 34 pt. demonstrated the presence of "spongiosal-cavernosal" shunts. CONCLUSIONS: The presence of "spongiosal-cavernosal" shunts justified the efficacy of A in term of complete penile erection, regarding also corpora cavernosa; in the other cases an increased speed of venous discharge justified the drug's lack of local efficacy, with poor results in terms of erection. KEY WORDS: Alprostadil, Erectile Dysfunction, Spongiosography.

Prostaglandin transporter (OATP2A1/SLCO2A1) contributes to local disposition of eicosapentaenoic acid-derived PGE3.

Eicosapentaenoic acid (EPA)-derived prostaglandin E3 (PGE3) possesses an anti-inflammatory effect; however, information for transporters that regulate its peri-cellular concentration is limited. The present study, therefore, aimed to clarify transporters involved in local disposition of PGE3. PGE3 uptake was assessed in HEK293 cells transfected with OATP2A1/SLCO2A1, OATP1B1/SLCO1B1, OATP2B1/SLCO2B1, OAT1/SLC22A6, OCT1/SLC22A1 or OCT2/SLC22A2 genes, compared with HEK293 cells transfected with plasmid vector alone (Mock). PGE3 uptake by OATP2A1-expressing HEK293 cells (HEK/2A1) was the highest and followed by HEK/1B1, while no significantly higher uptake of PGE3 than Mock cells was detected by other transporters. Saturation kinetics in PGE3 uptake by HEK/2A1 estimated the Km as 7.202 ± 0.595 µM, which was 22 times higher than that of PGE2 (Km=0.331 ± 0.131 µM). Furthermore, tissue disposition of PGE3 was examined in wild-type (WT) and Slco2a1-deficient (Slco2a1(-/-)) mice after oral administration of EPA ethyl ester (EPA-E) when they underwent intraperitoneal injection of endotoxin (e.g., lipopolysaccharide). PGE3 concentration was significantly higher in the lung, and tended to increase in the colon, stomach, and kidney of Slco2a1(-/-), compared to WT mice. Ratio of PGE2 metabolite 15-keto PGE2 over PGE2 concentration was significantly lower in the lung and colon of Slco2a1(-/-) than that of WT mice, suggesting that PGE3 metabolism is downregulated in Slco2a1(-/-) mice. In conclusion, PGE3 was found to be a substrate of OATP2A1, and local disposition of PGE3 could be regulated by OATP2A1 at least in the lung.

Single- and multiple-dose pharmacokinetics and tolerability of limaprost in healthy Chinese subjects.

BACKGROUND AND OBJECTIVES: Limaprost, a prostaglandin E1 analogue, is used to treat various symptoms in patients with ischemic diseases. The present study was designed to determine the pharmacokinetics and tolerability of single and multiple oral doses of limaprost 5 µg tablets in healthy Chinese subjects. METHODS: Single and multiple doses of 5-µg limaprost were orally administered to 12 healthy Chinese subjects. There was a 2-week washout period between single and multiple dosing. Blood samples were collected at various times. Indomethacin and aspirin were added to the blood samples to inhibit the endogenous release of prostaglandins during the sample processing. Plasma limaprost was measured by a two-dimensional liquid chromatography-tandem mass spectrometry method. RESULTS: After single dosing, limaprost was rapidly absorbed (time to reach maximum plasma concentration [t max] = 22.50 min) and eliminated (elimination half-life [t ½] = 21.70 min), with the maximum plasma concentration (C max) being 2.56 pg/mL and area under the concentration-time curve (AUC) from time 0 to the last quantifiable time point (AUC0-t) being 70.68 pg·min/mL. There were significant inter-individual variations in the AUCs for both single- and multiple-dose regimens. The values of C max, AUC, t ½ and t max were not statistically different between single and multiple dosing. The accumulation factor R was 0.609 ± 0.432 (R < 1), indicating that there was no accumulation after multiple dosing. There were no statistically significant differences in pharmacokinetic parameters for both single and multiple dosing between female and male subjects. The drug was well tolerated, with no severe adverse events being observed. CONCLUSIONS: Limaprost is rapidly absorbed after oral administration and is rapidly eliminated, with no accumulation after multiple dosing. The drug is well tolerated and no serious adverse events occurred.

Efecto del alprostadil en los cultivos de músculo esquelético isquémico. Estudio proteómico comparativo frente a músculo sano / The effect of alprostadil on ischemic skeletal muscle cultures. Comparative proteomic study with healthy muscle

INTRODUCCIÓN: El alprostadil tiene efecto vasodilatador y antiagregante. Es bien conocido su efecto endotelial, pero se desconocen sus posibles efectos pleiotrópicos sobre el músculo esquelético y si estos difieren en el músculo isquémico. OBJETIVO: Determinar el efecto del alprostadil sobre el metabolismo del músculo esquelético y valorar diferencias en su acción sobre el músculo isquémico frente al sano. MATERIAL Y MÉTODOS: Se obtuvieron muestras de tejido de 10 pacientes con isquemia irreversible intervenidos de amputación supracondílea, tanto de músculo isquémico (extensor corto de los dedos del pie, grupo I) como de músculo sano (músculo cuádriceps del borde de amputación, grupo S). Ambos grupos se cultivaron basalmente y con 5 ng de alprostadil. Se analizó la expresión proteómica de las siguientes enzimas: triosa-fosfato-isomerasa (TPI), malato deshidrogenasa (MDH), lactato deshidrogenasa (LDH) y piruvato carboxilasa (PC). Se determinaron también sus productos, lactato y piruvato. RESULTADOS: La MDH presentó una disminución en el grupo I en las muestras basales (2.196 ± 348 grupo S vs 644 ± 192 grupo I, p < 0,05). La PC estaba aumentada en el grupo I en ambos tipos de muestras (basal: 1,80 ± 1,27 vs 3,16 ± 2,25; alprostadil: 6,72 ± 2,13 vs 8,16 ± 3,63, grupo S vs grupo I, respectivamente, p < 0,05). No hubo diferencias significativas en la concentración de lactato ni en la de piruvato. CONCLUSIONES: La reducción de MDH en el músculo isquémico sugiere una reducción del ciclo de Krebs. El alprostadil estimula la expresión de PC, que induce la formación de oxalacetato; este se introduce en el ciclo de Krebs, permitiéndole funcionar parcialmente en el músculo isquémico y mejorando la obtención de energía

INTRODUCTION: Alprostadil has vasodilator properties and inhibits platelet aggregation. Its effects on endothelial wall have been widely studied, but there is no knowledge about possible skeletal muscle effects, and differences with ischemic muscle. OBJECTIVE: To determine the effects of alprostadil on skeletal muscle metabolism, and to investigate possible differences with ischemic muscle. METHODS: Samples were obtained in 10 patients with leg above-knee amputation due to severe irreversible ischemia, of ischemic muscle (extensor digitorum brevis, group I), and healthy muscle (quadriceps femoris, amputation edge, group S). Muscle segments were incubated with alprostadil 5 ng, or without it (baseline). Proteomic analysis of metabolic enzymes was performed: Triose-phosphate isomerase (TPI), malate dehydrogenase (MDH), lactate dehydrogenase (LDH) and pyruvate carboxylase (PC). Lactate and pyruvate was also determined. RESULTS: A decrease in malate dehydrogenase was observed in group I in the baseline samples (2196 ± 348 group S vs 644 ± 192 group I, P < 0.05). PC was increased in both samples in group I (baseline: 1.80 ± 1.27 vs 3.16 ± 2.25; alprostadil: 6.72 ± 2.13 vs 8.16 ± 3.63, group S vs group I, respectively, P < 0.05). No changes were observed in pyruvate and lactate. DISCUSSION: Decreased MDH in ischemic muscle suggests a Krebs cycle reduction. Alprostadil stimulates the expression of PC, which leads to oxaloacetate production. This product is inserted in Krebs cycle, improving energy obtaining. In this manner, Krebs cycle can work partially in ischemic muscle

Lubiprostone: pharmacokinetic, pharmacodynamic, safety and regulatory aspects in the treatment of constipation-predominant irritable bowel syndrome.

INTRODUCTION: Lubiprostone acts locally (apical membrane of human intestinal epithelial cells) as a highly selective type-2 chloride channel activator. It was approved in the USA for chronic idiopathic constipation (January 2006) and in women aged ≥ 18 years suffering from irritable bowel syndrome with constipation (IBS-C) (April 2008). So far, the only other pro-secretory medication approved in IBS-C and currently available in USA and Europe (since August and November 2012, respectively) is linaclotide. AREAS COVERED: This review outlines the regulatory history, pharmacokinetic, pharmacodynamic and safety data in the treatment of IBS-C with a European perspective. It is based on publicly available data, namely, published literature, drug labels and the FDA's spontaneous reporting system. EXPERT OPINION: Although interesting pharmacodynamic data suggest that lubiprostone may have additional mechanisms of action, its beneficial effects in IBS-C must be confirmed in the actual clinical scenario taking into account the new version of European Medicines Agency's guideline. This is especially important with regard to duration of studies (recommended to be at least 6 months) to adequately assess long-term sustained efficacy, withdrawal, rebound and safety. Further research is warranted in uncertain areas (i.e., males, pediatric and elderly patients). On the basis of current data, it is still too early to draw definite conclusions on the overall risk-benefit balance for IBS-C.

Determination of Prostaglandin E1 in dog plasma using liquid chromatography-tandem mass spectrometry and its application to a pharmacokinetic study.

The determination of Prostaglandin (PG) E1 in plasma is challenged by its low concentration (pg/mL) and endogenous interference. An LC-MS/MS method for the determination of PGE1 in dog plasma has been developed and validated. Plasma being sampled at 4°C and treated with indomethacin effectively inhibited interferents synthesized post-sampling. Samples were subjected to one-step extraction and separated by reversed phase HPLC with a short cycle time of 3min. An LLOQ of 10pg/mL was achieved with 500µl plasma. The method was applied to a pharmacokinetic study in beagle dogs involving an intravenous infusion of 3.2µg/kg PGE1. The half-life was recovered at 7min. The simple, sensitive and rapid method was suitable to be applied to pharmacokinetic studies of PGE1 at clinically relevant doses.

Prostaglandin E1 -containing nanoparticles improve walking activity in an experimental rat model of intermittent claudication.

OBJECTIVES: Due to the low stability of lipid emulsions, a lipid emulsion of prostaglandin E1 (Lipo-PGE1 ) necessitates daily intravenous drip infusions. To overcome this issue, we developed nanoparticles containing PGE1 (Nano-PGE1 ). Nano-PGE1 showed a good sustained-release profile of PGE1 from the nanoparticles in vitro, which may permit a longer-lasting therapeutic effect to be achieved. We here examined the pharmacological activity of Nano-PGE1 in a rat experimental model of intermittent claudication induced by femoral artery ligation. METHODS: The walking activity of the rat was tested on a rodent treadmill. Tissue levels of PGE1 were determined by enzyme immunoassay, and skeletal muscle angiogenesis (capillary growth) was monitored by immunohistochemical analysis. KEY FINDINGS: PGE1 could be detected in the lesion site one day after the intravenous administration of Nano-PGE1 but not of Lipo-PGE1 . An increased accumulation of Nano-PGE1 in the lesion site compared with control (unlesioned) site was also observed. The ligation procedure reduced the walking activity, which in turn was improved by a single administration of Nano-PGE1 but not of Lipo-PGE1 . The single administration of Nano-PGE1 also stimulated angiogenesis in the skeletal muscle around the ligated artery. CONCLUSIONS: The findings of this study suggest that Nano-PGE1 improves the walking activity of femoral artery-ligated rats through the accumulation and sustained release of PGE1 .

Pharmacokinetic and pharmacodynamic considerations for the current chronic constipation treatments.

INTRODUCTION: Chronic constipation is a frequent condition often treated pharmacologically. The laxatives available belong to very different pharmacologic groups. AREAS COVERED: This is a short but comprehensive review of the pharmacology, efficacy and safety of currently available laxatives for chronic constipation. Pertinent publications were retrieved from reference lists of publications and by literature searches via PubMed, lastly performed in November 2012. EXPERT OPINION: The most relevant laxative groups are the older representatives osmotic salts, sugars and sugar alcohols, macrogol, anthraquinones, diphenolic laxatives or diphenyl methanes (bisacodyl and sodium picosulfate) and the newer compounds prucalopride, lubiprostone and linaclotide. For all of these laxatives efficacy has been shown in controlled trials. Electrolyte losses do not occur when laxatives are given in therapeutic doses (rare exceptions with phosphate salts and salinic laxatives). The older laxatives are also safe regarding teratogenicity, abortion and lactation. For the newer compounds no respective data are available as yet. It is questionable whether the newer compounds offer advantages over the older ones. Unfortunately, comparative trials are lacking.

Safety evaluation of lubiprostone in the treatment of constipation and irritable bowel syndrome.

INTRODUCTION: Lubiprostone is approved in the United States for the treatment of chronic idiopathic constipation and constipation predominant irritable bowel syndrome (IBS-C). Lubiprostone causes secretion of fluid and electrolytes in the small bowel, through the activation of chloride channels, and thereby induces laxation and improvement of bowel functions. It is generally considered to be safe and effective. Common side effects of lubiprostone include nausea, diarrhea, abdominal pain and bloating, and the rare side effect dyspnea. Likely mechanisms for these side effects may be related to lubiprostone's primary action on small bowel secretion and the associated intestinal distension, as well as smooth muscle contraction. AREAS COVERED: This article reviews the pharmacokinetic and safety profile of lubiprostone, with particular relevance to the two FDA-approved dosages. EXPERT OPINION: Lubiprostone acts topically in the gut lumen and is almost completely metabolized in the gut lumen. Lubiprostone's M3 metabolite can be detected in low concentrations in the serum and may be responsible for some of its side effects. However, the exact mechanisms by which the side effects are produced are currently unknown.

Discovery of orally available 8-aza-5-thiaProstaglandin E1 analogs as highly selective EP4 agonists.

Analogs 8-aza-16-aryl prostaglandin E(1) (PGE(1)) and 8-aza-5-thia-16-arylPGE(1) were synthesized and evaluated with respect to their subtype receptor affinity and EP4 agonist activity for the purposes of identifying subtype-selective EP4 agonists that demonstrate oral efficacy. Using an inhibition assay of lipopolysaccharide (LPS)-induced tumor necrosis factor (TNF)-α production in rats, representative compounds were evaluated for their pharmacokinetic profiles and in vivo efficacy. Structure-activity relationships (SARs) were characterized and presented. Of the compounds tested, several demonstrated better oral exposure and/or in vivo efficacy compared with the previously reported analog 2a.

Accumlation of lipoprostaglandin E1 at the anastomotic region after systemic application.

Lipoprostaglandin E1 (lipo-PGE1) has been found accumulating in injured vascular regions. This study examined the localization of lipo-PGE1 in the anastomotic region. The study was divided into three parts. First, we performed anastomosis of the rat femoral artery and vein (n = 17). Lipo-PGE1 labeled with 1,1'-dioctadecyl-1,3,3',3'-tetramethyl-indocarbocyanine was infused intravenously. Hematoxylin-Eosin staining and fluorescence microscopic findings showed that lipo-PGE1 markedly accumulated at the anastomotic site when compared to the contralateral non anastomotic region. Then, we measured laser Doppler flow (LDF) of a lower leg before and after infusion of lipo-PGE1 (n = 7) and saline (n = 7). Increase of blood flow was maintained 1 hour after the infusion of lipo-PGE1 (144% ± 25.0%) when compared to saline infusion. Finally, we performed immunohistochemical and electron microscopic examinations and found that Lipo-PGE1 was incorporated in vascular smooth muscle cells of the anastomotic region. These findings suggest selective accumulation of the lipo-PGE1 in the vascular anastomosis site and affect on the blood flow of repaired vessels.

Influence of PEI as a core modifying agent on PLGA microspheres of PGE1, a pulmonary selective vasodilator.

This study tests the hypothesis that large porous poly (lactic-co-glycolic acid) (PLGA) microparticles modified with polyethyleneimine (PEI) are viable carriers for pulmonary delivery of prostaglandin E(1) (PGE(1)) used in the treatment of pulmonary arterial hypertension (PAH), a pulmonary vascular disorder. The particles were prepared by a double-emulsion solvent evaporation method with PEI-25 kDa in the internal aqueous phase to produce an osmotic pressure gradient. Polyvinyl alcohol (PVA) was used for external coating of the particles. The particles were examined for morphology, size, aerodynamic diameter, surface area, pore volume and in-vitro release profiles. Particles with optimal properties for inhalation were tested for in-vivo pulmonary absorption, metabolic stability in rat lung homogenates, and acute toxicity in rat bronchoalveolar lavage fluid and respiratory epithelial cells, Calu-3. The micromeritic data indicated that the PEI-modified particles of PGE(1) are optimal for inhalation. Incorporation of PEI in the formulations resulted in an increased entrapment efficiency - 83.26 ± 3.04% for particles with 1% PVA and 95.48 ± 0.46% for particles with 2% PVA. The amount of cumulative drug released into the simulated interstitial lung fluid was between 50.8 ± 0.76% and 55.36 ± 0.06%. A remarkable extension of the circulation half-life up to 6.0-6.5h was observed when the formulations were administered via the lungs. The metabolic stability and toxicity studies showed that the optimized formulations were stable at physiological conditions and relatively safe to the lungs and respiratory epithelium. Overall, this study demonstrates that large porous inhalable polymeric microparticles can be a feasible option for non-invasive and controlled release of PGE(1) for treatment of PAH.

Nano PGE1 promoted the recovery from spinal cord injury-induced motor dysfunction through its accumulation and sustained release.

The effect of Nano PGE(1) (nanoparticles containing prostaglandin E(1)) on spinal cord injury (SCI) was investigated in rat model. Nano PGE(1) significantly and dose-dependently promoted the recovery from SCI-induced motor dysfunction, and the potency of Nano PGE(1) was comparable with successive treatment of Lipo PGE(1), and was superior to single treatment of Lipo PGE(1). Distribution study revealed that Nano PGE(1) sustained longer in the blood. In the injured spinal cord, gradual accumulation and longer retention were observed. Lipo PGE(1) was also accumulated with time, but over 10 fold less. It should be noted that over 80 fold more of PGE(1) were detected in Nano PGE(1)-treated injured spinal cord as compared with that in normal ones. Nano PGE(1)-treated injured spinal cord had less lesion cavity with increased MBP expression. Also, HGF production significantly increased as compared with that of SCI control. These findings could lead to the conclusion that Nano PGE(1) had the therapeutic potential for SCI, which might be partly ascribed by the efficient distribution of Nano PGE(1) to the injured spinal cord. The sustained release of PGE(1) would have increased HGF production, and both would have promoted cell survival and endogenous repair.

Pharmacokinetic characteristics of a vasodilatory and antiplatelet agent, limaprost alfadex, in the healthy Korean volunteers.

Limaprost, a prostaglandin E1 analogue, with a strong vasodilatory and antiplatelet activity has been used to release from the symptoms of thromboangiitis obliterans (TAO), which is more prevalent in Korea and Japan, and lumbar spinal canal stenosis (LSCS). In spite of many uses of limaprost, the pharmacokinetics (PK) of it has not been studied in the Korean population. Therefore, a preliminary PK study was designed at a clinical oral dosage of 30-microg limaprost in 5 healthy Korean volunteers. Blood samples were obtained at 14 consecutive time points for 12 hours after dosing and analyzed by liquid chromatography-tandem mass spectrometry with electrospray ionization (LC-ESI/MS/ MS) at a very low detection limit of 0.5 pg/mL of limaprost in human plasma with considerably short run time (18 minutes). Pharmacokinetic characteristics resulted in ''time for maximal concentrations (T(max) 0.5 hour),'' ''elimination half-life (T(1/2) 1.64 hours),'' ''maximal concentration (C(max) 13.37 pg/mL),'' ''area under the curve (AUC(12 hours) 18.60 pg . h/mL),'' ''AUC extrapolated to infinity (AUC(infinity) 22.98 pg . h/mL),'' ''extrapolation (AUC(infinity - 12 hours)/AUC(infinity) 0.15%),'' ''elimination rate constant (k(e) 0.68 h(-1)),'' ''systemic clearance (CL 1.77 L/h),'' and ''mean residence time (MRT 1.74 hours).'' These results showed that orally administered 30-microg limaprost was rapidly and highly absorbed, and it was considerably eliminated fast from the blood stream in the healthy Korean volunteers.

Feasibility study of aerosolized prostaglandin E1 microspheres as a noninvasive therapy for pulmonary arterial hypertension.

This study was designed to test the feasibility of polymeric microspheres as an inhalable carrier for prostaglandin E(1) (PGE(1)) for treatment of pulmonary arterial hypertension. Poly(lactic-co-glycolic acid) (PLGA) microspheres were prepared by a double emulsion-solvent evaporation method. Six different microspheric formulations were prepared using two different blends of PLGA (50:50 and 85:15) and varying concentrations of polyvinyl alcohol (PVA) in the external aqueous phase (EAP). The particles were characterized for morphology, size, aerodynamic diameter, entrapment efficiency, release patterns, and metabolic stability. Pulmonary absorption was studied in a rat model, and safety of the formulations was evaluated by measuring cytotoxicity in Calu-3 cells and assessing injury markers in bronchoalveolar lavage (BAL) fluid. Both actual particle size and aerodynamic diameter of the formulations decreased with increasing PVA concentration. The mass median aerodynamic diameter of the particles was within the respirable range. Entrapment efficiency increased with increasing PVA concentration; PLGA 85:15 showed better entrapment due to hydrophobic interactions with the drug. Compared to intravenously administered PGE(1), microspheres prepared with PLGA 85:15 produced a 160-fold increase in the half-life of PGE(1) following pulmonary administration. Although plain PGE(1) showed rapid degradation in rat lung homogenate, PGE(1) entrapped in the particles remained intact for about 8 h. Optimized formulations were demonstrated to be safe, based on analysis of cytotoxicity and lung-injury markers in BAL fluid. Overall, the data suggest that microspheric PGE(1) formulations have the potential to be used as a noninvasive and controlled-release alternative to the current medications used for treatment of pulmonary arterial hypertension that are administered by continuous infusion or require multiple inhalations.

Determination of alprostadil in rat plasma by ultra performance liquid chromatography-electrospray ionization-tandem mass spectrometry after intravenous administration.

A rapid, highly selective ultra performance liquid chromatography-electrospray ionisation-tandem mass spectrometry method (UPLC-ESI-MS/MS) was developed and validated for the determination and pharmacokinetic investigation of alprostadil in rat plasma. After a simple sample preparation procedure involving a one-step liquid-liquid extraction, alprostadil and the internal standard, diphenhydramine, were chromatographed on an ACQUITY UPLC BEH C(18) column with gradient elution using a mobile phase consisting of acetonitrile and water (containing 0.1% formic acid) at a flow rate of 0.25 mL min(-1). The detection was performed on a triple quadrupole tandem mass spectrometer in multiple reaction monitoring (MRM) mode via an electrospray ionization (ESI) source. The calibration curve was linear (r(2)=0.99) over the concentration range 0.4-250.0 ng mL(-1), with a lower limit of quantification of 0.4 ng mL(-1) for alprostadil. The inter- and intra-day precision (%R.S.D.) was less than 8.5% and 2.4%, respectively, and the accuracy (RE%) was between 9.3% and 1.0% (n=6). Alprostadil in rat plasma was stable when stored at room temperature for 0.5h and at -20 degrees C for two weeks. The method was very rapid, simple and reliable, and was employed for the first time for the pharmacokinetic studies of alprostadil in rats after a single intravenous administration of 50 microg kg(-1).

Lubiprostone for chronic idiopathic constipation and irritable bowel syndrome with constipation.

Lubiprostone, a locally acting highly selective type-2 chloride channel activator, has been US FDA approved since January 2006 for the treatment of adults with chronic idiopathic constipation and FDA approved since April 2008 for the treatment of woman aged 18 years or older suffering from irritable bowel syndrome (IBS) with constipation. Through activation of the type-2 chloride channels located on the luminal side of intestinal epithelial cells, it promotes fluid secretion, increasing the liquid content of stool and accelerating small bowel as well as colonic transit. Lubiprostone has demonstrated efficacy with respect to increasing weekly spontaneous bowel movements and improving stool consistency, straining and constipation severity, both in short- and long-term studies. It has also demonstrated efficacy in the treatment of IBS with constipation, with beneficial effects on global symptoms, abdominal pain, constipation-related symptoms and overall quality of life. There is no evidence of a rebound in constipation or IBS symptoms following cessation of lubiprostone. In general, lubiprostone is well tolerated, with the most common side effects including nausea, headache and diarrhea.

Long beneficial effects after lipo-prostaglandin E1 administration on the ischemic gastric tube in pigs.

Lipo-prostaglandin E1 (PGE1) is a new preparation of PGE1 in which it is bound to lipids in order to slow PGE1 release and delay its rate of metabolism. We investigated how long the beneficial effects of intravenous administration of lipo-PGE1 on the ischemic gastric tube continue. The gastric tube was constructed using 15 domestic pigs under general anesthesia and saline, unmodified PGE1 and lipo-PGE1 were infused continuously at a rate of 0.05 microg/kg/min for 10 minutes. Tissue blood flow (TBF) was analyzed from before administration to 120 minutes after the end of administration. There were no obvious changes in TBF during the administration of saline. However, TBF during treatment with unmodified PGE1 and lipo-PGE1 was significantly increased to 13.1 +/- 1.3 and 13.5 +/- 1.4 mL/min/100 g, respectively (paired t-test; P < 0.01). Although TBF was significantly decreased to 8.0 +/- 1.0 mL/min/100 g on 10 minutes after the end of unmodified PGE1 administration (paired t-test; P < 0.01), it was maintained over 10 mL/min/100 g until 120 minutes in lipo-PGE1 group. Lipo-PGE1 infusion leads to the objectively measurable improvement and the prolonged action in the blood perfusion of the gastric tube in pigs.