Determination of olanzapine in serum by high-performance liquid chromatography using ultraviolet detection considering the easy oxidability of the compound and the presence of other psychotropic drugs

A method for determination of the atypical neuroleptic drug olanzapine in serum was developed. After a single-step liquid–liquid extraction, the compound was separated from other constituents on a normal-phase silica gel column using a buffer–methanol mobile phase and measured by UV absorption at 270 nm. Addition of 0.25% ascorbic acid to serum protects olanzapine against oxidation during extraction and stabilizes the easily oxidised compound during storage. Inter-day variation was <8% at serum levels found in olanzapine treated patients. Analytical interference from coadministered psychoactive drugs and their metabolites were studied. Only risperidone, also a relatively newly developed antipsychotic drug, interfered, but the most commonly used antidepressants and traditional antipsychotics and their metabolites did not interfere.     

Determination of risperidone and 9-hydroxyrisperidone in human plasma by liquid chromatography: application to the evaluation of CYP2D6 drug interactions

A high-pressure liquid chromatography with ultra-violet detection method for the simultaneous determination of risperidone and 9-hydroxyrisperidone in plasma after liquid-liquid extraction has been developed. The limit of quantitation was 5 nmol/L, and the inter-day coefficient of variation was less than 8% for both compounds. The mean recoveries of risperidone and 9-hydroxyrisperidone added to plasma were 96.8 and 99.4%, with an intra-day coefficient of variation of under 5 and 6%, respectively. Studies of analytical interference showed that the most commonly co-administered antidepressants and benzodiazepines did not interfere. The method was used for the determination of the plasma concentrations of a schizophrenic patient treated daily with an oral dose of 4.5 mg risperidone. The patient suffered severe extrapyramidal side-effects after adding risperidone to his previous medication of haloperidol and levomepromazine. The risperidone plasma concentration was well above the average (182 nmol/L), which suggests that a pharmacokinetic interaction occurred, presumably due to inhibition of the enzyme CYP2D6.     

The D2 dopamine receptor occupancy of risperidone and its relationship to extrapyramidal symptoms: A pet study

Risperidone is a recently introduced neuroleptic distinguished by a decreased incidence of extrapyramidal side effects (EPS). The mechanism of its low EPS is unclear. Since it has been shown that EPS is related to the level of D₂ receptor occupancy, we studied nine patients receiving 2–6 mg/day of risperidone using [¹¹C]-raclopride PET scans in order to determine the in vivo D₂ receptor binding characteristics of risperidone. The mean level of receptor occupancy was 66% at 2 mg; 73% at 4 mg; and 79% at 6 mg. Three patients, those with the highest receptor occupancies, exhibited mild EPS, though none required anitparkinsonian medications. Our results suggest that at doses of 4–6 mg the in vivo D₂ receptor occupancy of risperidone is similar to that of typical neuroleptics and higher than that of clozapine. This would suggest that the EPS benefits of risperidone cannot be explained by a low D₂ binding but may be related to its high 5-HT₂ affinity. However, the emergence of EPS at higher levels of D₂ receptor occupancy, in this study and in previous clinical trials, would suggest that risperidone's high 5-HT₂ affinity provides only a relative protection from EPS. and once the D₂ occupancy exceeds a certain threshold this ‘relative’ 5-HT₂-mediated protection from EPS may be lost.     

Effect of the atypical neuroleptic risperidone on morphology and S100B secretion in C6 astroglial lineage cells

We investigated the effect of the atypical neuroleptic risperidone on morphology and S100B secretion in C6 glioma cells, considering the putative involvement of astroglial cells in neuropsychiatric disorders. In the presence of high experimental doses of risperidone, C6 cells become stellate, with process-bearing cells and partial retraction of the cell body followed by detachment from the adhesion surface with practically no cell death. These results indicate that risperidone is able to interfere with C6 cell adhesion without toxic effects. RhoA activator LPA prevented the effects of risperidone on cell morphology. From 6 h risperidone induced a statistically significant increment of about 80% in S100B secretion. These data contribute to the proposal that glial cells are targets of risperidone, which could be involved in the therapeutic response of risperidone to improve autism symptoms.     

The influence of several nutritional supplements on the rational use of cabozantinib

To promote the rational use of cabozantinib (CBZ), this paper studied the influence of several nutritional supplements on the interaction between CBZ and bovine serum albumin (BSA), an appropriate alternative model for human serum albumin (HSA) that is one of the important transporter proteins in plasma, by fluorescence spectroscopy and UV–vis spectroscopy. The results showed that CBZ could quench the fluorescence of BSA via a dynamic–static quenching process, and the six nutritional supplements did not change the quenching mode of BSA by CBZ. However, all of them could reduce the binding constant of the CBZ–BSA system at 293 K and increase the polarity around tryptophan residues. Among them, nicotinamide and vitamin B₁₂ (VB₁₂) had a greater effect on the binding constants of the CBZ–BSA system. In the meantime, the thermodynamic parameters of the CBZ–BSA system were examined, indicating that the interaction of CBZ with BSA was spontaneous and dominated by hydrophobic forces. Further research discovered that the combining of CBZ with BSA was primarily located within Site I of BSA, and the binding distance r was 2.48 nm. Consequently, while taking CBZ, patients should use VB₁₂ and nicotinamide carefully, which may interfere with the transport of drugs.     

Serum IL-1beta,sIL-2R,IL-6, IL-8 and TNF-alpha in schizophrenic patients,relation with symptomatology and responsiveness to risperidone treatment.

Activation of the inflammatory response system and varied levels of cytokines in acute schizophrenia have been suggested by recent studies. Psychopharmacologic agents can differentially effect cytokine production, which suggests that therapeutic function of neuroleptics may involve immunomodulation. The present study was carried out to examine: (i) serum concentrations of interleukin (IL)-1beta, soluble interleukin-2 receptor (sIL-2R), IL-6, IL-8 and tumour necrosis factor (TNF)-alpha in schizophrenic patients; (ii) their relation with psychopathological assessment; and (iii) the relation of the initial cytokine levels with responsiveness to risperidone therapy. Thirty-four drug-free schizophrenic patients with acute exacerbation and 23 age- and gender-matched healthy controls were recruited for this study. Psychopathological assessments at admission and throughout risperidone treatment for 60 days were recorded. Serum cytokine concentrations were determined with chemilumunescence assays. According to our results, serum IL-1beta, sIL-2R, IL-6, IL-8 and TNF-alpha concentrations adjusted for age, gender, body mass index and smoking were no different in patients with schizophrenia and controls and among subtypes of schizophrenia. However, the initial TNF-alpha concentrations had a significant effect on Brief Psychiatric Rating Scale and Scale Assessment of Positive Symptoms scores. The initial cytokine concentrations of the patients responsive to risperidone were not significantly different from those of non-responsive patients. The present study demonstrates that plasma levels of IL-1beta, sIL-2R, IL-6, IL-8 and TNF-alpha adjusted for confounding factors are not altered in drug-free schizophrenic patients at acute exacerbation. We suggest that, if cytokine production is altered in schizophrenia, these alterations may not be detectable in systemic circulation. According to our results, the therapeutic effect of risperidone is not related to basal levels of the aforementioned cytokines. However, serum TNF-alpha may contribute to symptomatology in schizophrenia     

High-resolution preparative gel electrophoresis: separation and recovery of functional messenger RNA species

Certain open-chain polyols were shown to interfere with the determination of phosphorus of the Lowry-Lopez method by forming a complex with Mo₇O₂₄^6?. The ability to interfere with the assay increased with increasing chain length of the polyols: Ethylene glycol and glycerol did not react at all; i-erythritol reacted to a small extent, but hexitols and perseitol formed stronger complexes. Depending on the polyol, interference occurred even at 0.2 mm (hexitols) or 2 mm (xylitol) concentrations. At these concentrations the polyols interfered only to a small extent with the phosphorus assays based on the use of Triton X-100 and molybdate. The complex formation was exploited in the development of a colorimetric polyol assay.     

A sensitive method for measuring pyrophosphate in the presence of a 10,000-fold excess of orthophosphate using inorganic pyrophosphatase

A simple method for measuring PP_i at concentrations down to 2 μm has been devised using the ability of inorganic pyrophosphatase to be inactivated by fluoride in the presence of PP_i. Orthophosphate (20 mm) and a number of other compounds did not interfere with the assay. The applicability of the method for direct measurement of PP_i in urine is demonstrated.     

Inducible expression of maize polyamine oxidase in the nucleus of MCF-7 human breast cancer cells confers sensitivity to etoposide

Summary. In this study, polyamine oxidase from maize (MPAO), which is involved in the terminal catabolism of spermidine and spermine to produce an aminoaldehyde, 1,3-diaminopropane and H₂O₂, has been conditionally expressed at high levels in the nucleus of MCF-7 human breast cancer cells, with the aim to interfere with polyamine homeostasis and cell proliferation. Recombinant MPAO expression induced accumulation of a high amount of 1,3-diaminopropane, an increase of putrescine levels and no alteration in the cellular content of spermine and spermidine. Furthermore, recombinant MPAO expression did not interfere with cell growth of MCF-7 cells under normal conditions but it did confer higher growth sensitivity to etoposide, a DNA topoisomerase II inhibitor widely used as antineoplastic drug. These data suggest polyamine oxidases as a potential tool to improve the efficiency of antiproliferative agents despite the difficulty to interfere with cellular homeostasis of spermine and spermidine. Authors’ address: Dr. Paraskevi Tavladoraki, Department of Biology, University ‘Roma Tre’, Viale G. Marconi 446, 00146 Rome, Italy     

Analysis of cisapride in neonatal plasma using high-performance liquid chromatography with a base-stable column and fluorescence detection

A simple, selective, sensitive and precise high-performance liquid chromatographic plasma assay for the prokinetic drug cisapride is described. Alkalinised samples of plasma (100 μl) were extracted with 1.0 ml of 10% (v/v) isopropanol in chloroform, dried, redissolved in mobile phase and injected. Chromatography was performed at 20°C by pumping a mobile phase of acetonitrile (370 ml) in pH 5.2, 0.02 M phosphate buffer (630 ml) at 1.0 ml/min through a C₈ Symmetry column. Cisapride and the internal standard were detected by fluorescence monitoring at 295 nm (excitation) and 350 nm (emission), and were eluted 5 min and 8 min, respectively, after injection. Calibration plots in bovine serum albumin (3% w/v) were linear (r > 0.999) from 5 to 250 ng/ml. Intra-day and inter-day precision (C.V.) was 9.5%, or less, and the accuracy was within 5.5% of the nominal concentration over the range 8–200 ng/ml. Total assay recovery was above 82%. Endogenous plasma components, major cisapride metabolite (norcisapride), and other durgs used in neonatal pharmacotherapeutics did not interfere.     

Development of Controlled-Release Matrix Tablet of Risperidone: Influence of Methocel®- and Ethocel®-Based Novel Polymeric Blend on <Emphasis Type="Italic">In Vitro</Emphasis> Drug Release and Bioavailability

Controlled-release (CR) matrix tablet of 4 mg risperidone was developed using flow bound dry granulation–slugging method to improve its safety profile and compliance. Model formulations F1, F2, and F3, consisting of distinct blends of Methocel® K100 LV-CR and Ethocel® standard 7FP premium, were slugged. Each batch of granules (250–1,000 μm), obtained by crushing the slugs, was divided into three portions after lubrication and then compressed to 9-, 12-, and 15-kg hard tablets. In vitro drug release studies were carried out in 0.1 N HCl (pH 1.2) and phosphate buffer (pH 6.8) using a paddle dissolution apparatus run at 50 rpm. The CR test tablet, containing 30% Methocel® and 60% Ethocel® (F3) with 12-kg hardness, exhibited pH-independent zero-order release kinetics for 24 h. The drug release rate was inversely proportional to the content of Ethocel®, while the gel layer formed of Methocel® helped in maintaining the integrity of the matrix. Changes in the hardness of tablet did not affect the release kinetics. The tablets were reproducible and stable for 6 months at 40 ± 2°C/75 ± 5% relative humidity. Risperidone and its active metabolite, 9-hydroxyrisperidone, present in the pooled rabbit’s serum, were analyzed with HPLC-UV at λ_max 280 nm. The CR test tablet exhibited bioequivalence to reference conventional tablet in addition to the significantly (p < 0.05) optimized peak concentration, C_max, and extended peak time, T_max, of the active moiety. There was a good association between drug absorption in vivo and drug release in vitro (R² = 0.7293). The successfully developed CR test tablet may be used for better therapeutic outcomes of risperidone.KEY WORDS: controlled release, dry granulation slugging method, risperidone     

Influence of selenium and zinc on performance,blood constituents,and immune response in stressed calves

Eighty weanling beef calves were used to determine the effects of zinc and selenium supplementation on performance, immune response, and blood characteristics during stress. Treatments included: (1) control; (2) control diet with selenium injection [15 mg/hd (head)] (3) zinc diet (25 mg Zn/hd/d, added as ZnO); and (4) zinc diet with selenium injection. Feed intake and weight gains were not affected by zinc throughout the 28-d study. However, selenium improved weight gains in calves with low initial selenium status in the first 14 d of the study. Glutathione peroxidase (GSH-Px) was increased by selenium and decreased by zinc on d 19. Zinc appeared to interfere with the role of selenium in GSH-Px activity at the cellular level. Plasma zinc was not affected by treatment. Zinc supplementation resulted in increased serum sodium and decreased serum total protein and glutamic-oxaloacetic transaminase. Immune response was measured via antibody titers to Infectious Bovine Rhinotracheitis (IBR) and Para-influenza₃ (PI₃) viruses 19 d postvaccination. Levels of titers were not affected by either zinc or selenium, however, titers did reflect a response to vaccination between sampling dates. On d 19, zinc increased the percentage of leukocytes that were monocytes. Total leukocyte, neutrophil and lymphocyte numbers did not differ across treatments. These results suggest that selenium or zinc supplementation may individually improve an animal’s response to stress.     

Treatment with the Antipsychotic Agent,Risperidone, Reduces Disease Severity in Experimental Autoimmune Encephalomyelitis

Recent studies have demonstrated that atypical antipsychotic agents, which are known to antagonize dopamine D2 and serotonin 5-HT_2a receptors, have immunomodulatory properties. Given the potential of these drugs to modulate the immune system both peripherally and within the central nervous system, we investigated the ability of the atypical anti-psychotic agent, risperidone, to modify disease in the animal model of multiple sclerosis (MS)⁴, experimental autoimune encephalomyelitis (EAE). We found that chronic oral administration of risperidone dose-dependently reduced the severity of disease and decreased both the size and number of spinal cord lesions. Furthermore, risperidone treatment substantially reduced antigen-specific interleukin (IL)-17a, IL-2, and IL-4 but not interferon (IFN)-γ production by splenocytes at peak disease and using an in vitro model, we show that treatment of macrophages with risperidone alters their ability to bias naïve T cells. Another atypical antipsychotic agent, clozapine, showed a similar ability to modify macrophages in vitro and to reduce disease in the EAE model but this effect was not due to antagonism of the type 1 or type 2 dopamine receptors alone. Finally, we found that while risperidone treatment had little effect on the in vivo activation of splenic macrophages during EAE, it significantly reduced the activation of microglia and macrophages in the central nervous system. Together these studies indicate that atypical antipsychotic agents like risperidone are effective immunomodulatory agents with the potential to treat immune-mediated diseases such as MS.     

Inhibition of complex I by neuroleptics in normal human brain cortex parallels the extrapyramidal toxicity of neuroleptics

There is increasing evidence that a defect of the mitochondrial respiratory chain is implicated in the development of Parkinson disease. Decreased complex I activity of the mitochondrial respiratory chain has been reported in platelets, muscle, and brain of patients with Parkinson disease. Extrapyramidal symptoms (e.g. parkinsonism and dystonic reactions) are major limiting side effects of neuroleptics. Experimental evidence suggests that neuroleptics inhibit complex I in rat brain. There has not been a study of the effects of neuroleptics in human tissue, however. We therefore analyzed the activities of complexes I + III, complexes II + III, succinate dehydrogenase, complex IV (cytochrome c oxidase), and of citrate synthase in normal human brain cortex after the addition of haloperidol and chlorpromazine and the atypical neuroleptics risperidone, zotepine, and clozapine. Activity of complex I was progressively inhibited by all neuroleptics. Half maximal inhibition (IC₅₀) was 0.1 mM fo r haloperidol, 0.4 mM for chlorpromazine, and 0.5 mM for risperidone and zotepine. Clozapine had no effect on enzyme activity at concentrations up to 0.5 mM, followed by a slow decline with a maximum inhibition of 70% at 10 mM. IC50 was at about 2.5 mM. Thus, the concentration of clozapine needed to cause 50% inhibition of the activity of complexes I and III was about 5 times that of zotepine and risperidone, about 6 times that of chlorpromazine, and 25 times that of haloperidol. The inhibition thus paralleled the incidence of extrapyramidal effects caused by the different neuroleptics as they are known from numerous clinical studies. Our data support the hypothesis that neuroleptic-induced extrapyramidal side effects may be due to inhibition of the mitochondrial respiratory chain. (Mol Cell Biochem 174: 255–259, 1997)     

Lipid binding to sterol carrier protein-2 is inhibited by ethanol

Sterol carrier protein-2 (SCP-2) is an intracellular lipid carrier protein that binds cholesterol, phospholipids, fatty acids and other ligands. It has been reported that expression of SCP-2 was increased in brain nerve endings or synaptosomes of chronic ethanol-treated mice and it was shown that cholesterol homeostasis was altered in brain membranes of chronic ethanol-treated animals. Ethanol may interfere with the capacity of SCP-2 to bind cholesterol as well as other lipids. This hypothesis was tested using recombinant SCP-2 and fluorescent-labeled cholesterol, phosphatidylcholine (PC), and stearic acid. The association constants (K_a) of the ligand-SCP-2 complex were in the following order: NBD-cholesterol>NBD-PC>NBD-stearic acid. Ethanol, beginning at a concentration of 25 mM, significantly reduced the affinity of NBD-cholesterol and NBD-PC for SCP-2. Effects of ethanol on the K_a of NBD-stearic acid was significant only at the highest concentration that was examined (200 mM). Ethanol significantly increased the B_max of NBD-cholesterol for SCP-2 but did not have a significant effect on the B_max of NBD-PC. Similar results were found for effects of ethanol on the K_as and B_maxs using pyrene-labeled cholesterol and PC. In conclusion, ethanol beginning at a physiological concentration of 25 mM inhibited binding of cholesterol and PC to SCP-2. However, effects of ethanol on lipid binding to SCP-2 were dependent on the type of lipid. Ethanol in vivo may interfere with lipid binding to SCP-2 and disrupt lipid trafficking within cells.     

Study of the role of iron in the anticryptococcal activity of human serum and fluconazole

Anticryptococcal activity of human serum and apotransferrin in RPMI 1640 was studied in vitro. The effects of varying concentrations of FeCl₃ on this activity was investigated. Possible synergy of serum and apotransferrin with fluconazole was also measured. The fungistatic activity of human serum, whether lyophilized, stored at 4 °C, fresh frozen or purchased from commercial sources vs. Cryptococcus neoformans was comparable. There was no significant loss of fungistatic activity after freezing and thawing the serum up to 10 times. The fungistatic activity of human serum was similar when tested in different tissue culture media with the exception of Medium 199. The addition of apotransferrin (2.0 or 0.2 mg/ml) to RPMI 1640 had an inhibitory effect on cryptococcal growth. This effect was reversed by 20 M of FeCl₃ at both apotransferrin concentrations. By contrast, addition of FeCl₃ to human serum and RPMI 1640 did not reverse inhibition of growth. Fluconazole synergized with the human serum preparations described, but not with pooled commercial serum, for fungicidal activity. Synergistic activity of fluconazole and human serum was not affected by the addition of FeCl₃. Apotransferrin did not show any synergistic fungicidal activity with fluconazole.     

三种非典型抗精神病药对儿童青少年精神分裂症患者血脂、肝功能和认知功能的影响

摘要 目的：探讨利培酮、阿立哌唑、奥氮平分别对儿童青少年精神分裂症患者肝功能、血脂和认知功能的影响。方法：选取2015年1月至2019年12月我院收治的84例儿童青少年精神分裂症患者,采用乱数表法随机分为阿立哌唑组（n=28,阿立哌唑治疗）、利培酮组（n=28,利培酮治疗）、奥氮平组（n=28,奥氮平治疗）,均治疗8周,对比三组患者症状评分、血脂、肝功能、认知功能以及不良反应。结果：三组治疗8周后阳性与阴性症状量表（PANSS）评分整体比较无差异（P>0.05）,三组治疗8周后PANSS评分均较治疗前降低（P<0.05）。奥氮平组、利培酮组治疗8周后三酰甘油(TG)、总胆固醇(TC)、低密度脂蛋白(LDL-C)高于阿立哌唑组,且奥氮平组高于利培酮组（P<0.05）;奥氮平组、利培酮组治疗8周后高密度脂蛋白(HDL-C)低于阿立哌唑组,且奥氮平组低于利培酮组（P<0.05）。三组不良反应发生率整体比较无差异（P>0.05）。阿立哌唑组治疗8周后延迟回忆数、即刻回忆数、回忆总数、再认数评分均高于利培酮组、奥氮平组（P<0.05）。利培酮组治疗8周后ALT、AST、TBIL高于治疗前（P<0.05）,利培酮组治疗8周后ALT、AST、TBIL高于阿立哌唑组、奥氮平组（P<0.05）。结论：利培酮、阿立哌唑、奥氮平应用于儿童青少年精神分裂症中,可获得相当的治疗效果,其中利培酮对肝功能影响较大,奥氮平对人体血脂影响较大,阿立哌唑对血脂、肝功能影响轻,改善认知功能效果优于利培酮、奥氮平。     

Determination of risperidone and its major metabolite 9-hydroxyrisperidone in human plasma by reversed-phase liquid chromatography with ultraviolet detection

A simple and sensitive high-performance liquid chromatographic (HPLC) method with UV absorbance detection is described for the quantitation of risperidone and its major metabolite 9-hydroxyrisperidone in human plasma, using clozapine as internal standard. After sample alkalinization with 1 ml of NaOH (2 M) the test compounds were extracted from plasma using diisopropyl ether–isoamylalcohol (99:1, v/v). The organic phase was back-extracted with 150 μl potassium phosphate (0.1 M, pH 2.2) and 60 μl of the acid solution was injected into a C₁₈ BDS Hypersil analytical column (3 μm, 100×4.6 mm I.D.). The mobile phase consisted of phosphate buffer (0.05 M, pH 3.7 with 25% H₃PO₄)–acetonitrile (70:30, v/v), and was delivered at a flow-rate of 1.0 ml/min. The peaks were detected using a UV detector set at 278 nm and the total time for a chromatographic separation was about 4 min. The method was validated for the concentration range 5–100 ng/ml. Mean recoveries were 98.0% for risperidone and 83.5% for 9-hydroxyrisperidone. Intra- and inter-day relative standard deviations were less than 11% for both compounds, while accuracy, expressed as percent error, ranged from 1.6 to 25%. The limit of quantitation was 2 ng/ml for both analytes. The method shows good specificity with respect to commonly prescribed psychotropic drugs, and it has successfully been applied for pharmacokinetic studies and therapeutic drug monitoring.     

ACTH-induced inhibition of endogenous rat brain protein phosphorylation in vitro: Structure activity

ACTH_1–24 inhibits the endogenous phosphorylation in vitro of distinct SPM protein bands. Using N-terminal fragments of ACTH, the structure-activity requirements for this effect were studied. A rather complex interaction of the ACTH fragments with endogenous SPM phosphorylation was observed. The effects were not only dependent on the primary structure of the peptide used, but also on the protein band studied and the ATP/SPM ratio used in the incubation system. ACTH_1–24 did not interfere with the ATP-hydrolyzing activity of the SPM preparation, nor did it influence the endogenous phosphatase activity. Therefore, a direct interaction of ACTH with SPM protein kinase(s) is likely to be responsible for its effect on phosphorylation.