Potentiometric sensor for methylene blue based on methylene blue–silicotungstate ion association and its pharmaceutical applications

A methylene blue (MB) poly(vinyl chloride) membrane sensor based on MB–silicotungstate (SLT) ion association as electroactive material was described. The linear response covered the range 1×10⁻³–1×10⁻⁶ mol dm⁻³ MB solution, with a slope 52.0±0.8 mV decade⁻¹ (pH range 3.0–10.0). The detection limit was 7.65×10⁻⁷ mol dm⁻³. The electrode showed stability, good reproducibility and fast response. Interferences from common inorganic cations, some organic base were negligible. These characteristics of the electrode enabled it to be used successfully for the determination of MB in injection. There was a good agreement for the results of MB content in injection between potentiometric method and USP standard procedure.     

High absolute bioavailability of methylene blue given as an aqueous oral formulation

Purpose  Methylene blue (MB) has recently been reevaluated for malaria treatment. With the aim of excluding treatment failures due to low bioavailability, we have investigated the absolute bioavailability of MB given as an aqueous oral formulation and its interaction with chloroquine (CQ). Methods  A phase I study in 16 healthy individuals was performed as a monocenter prospective open randomized intra-individual cross-over comparison of MB single doses [50 mg intravenous (i.v.), 500 mg orally, separated by a 1-week wash-out]. After a second week, the group was split for a randomized parallel group comparison of CQ 750 mg administered orally alone or combined with 500 mg MB orally. Results  Mean MB plasma area under the substrate concentration–time curve () was 7,639 ± 3,384 ng/mL*h and 51,171 ± 17,147 ng/mL*h after i.v. and oral administration, respectively (dosage 1:10), and 76,897 ± 46,037 ng/mL*h after MB combined with CQ. The absolute bioavailability was 72.3 ± 23.9%. Co-administration with CQ significantly increased MB plasma concentrations (p ≤ 0.016); CQ kinetics remained unaffected. Conclusion  The absolute bioavailability of MB is high. Co-administration of MB and CQ increases plasma, but not whole blood MB concentrations.     

The interaction between methylene blue and the cholinergic system

1. The inhibitory effects of methylene blue (MB) on different types of cholinesterases and [³H]-N-methylscopolamine ([³H]-NMS) binding to muscarinic receptors were studied.
2. Human plasma from young healthy male volunteers, purified human pseudocholinesterase and purified bovine true acetylcholinesterase were incubated with acetylcholine and increasing concentrations of MB (0.1–100 μmol l⁻¹) in the presence of the pH-indicator m-nitrophenol for 30 min at 25°C. The amount of acetic acid produced by the enzymatic hydrolysis of acetylcholine was determined photometrically.
3. Rat cardiac left ventricle homogenate was incubated with [³H]-NMS and with increasing concentrations of MB (0.1 nmol l⁻¹–100 μmol l⁻¹) at 37°C for 20 min. The binding of [³H]-NMS to the homogenate was quantified by a standard liquid scintillation technique.
4. MB inhibited the esterase activity of human plasma, human pseudocholinesterase and bovine acetylcholinesterase concentration-dependently with IC₅₀ values of 1.05±0.05 μmol l⁻¹, 5.32±0.36 μmol l⁻¹ and 0.42±0.09 μmol l⁻¹, respectively. MB induced complete inhibition of the esterase activity of human plasma and human pseudocholinesterase, whereas bovine acetylcholinesterase was maximally inhibited by 73±3.3%.
5. MB was able to inhibit specific [³H]-NMS binding to rat cardiac left ventricle homogenate completely with an IC₅₀ value of 0.77±0.03 μmol l⁻¹, which resulted in a K_i value for MB of 0.58±0.02 μmol l⁻¹.
6. In conclusion, MB may be considered as a cholinesterase inhibitor with additional, relevant affinity for muscarinic binding sites at concentrations at which MB is used for investigations into the endothelial system. In our opinion these interactions between MB and the cholinergic system invalidate the use of MB as a tool for the investigation of the L-arginine-NO-pathway, in particular when muscarinic receptor stimulation is involved.

     

亚甲蓝对感染性休克犬血液动力学和氧合功能的影响

目的　观察鸟苷酸环化酶抑制药亚甲蓝 (MB)对感染性休克犬血液动力学和氧合功能的影响。方法　采用内毒素静脉注入诱导的 1 1只感染性休克犬模型 ,输注 0 9%的氯化钠后 ,MB2mg/kg1 5分钟内静脉注入。经右颈内静脉放置的肺动脉导管测定基础、休克 1小时、输注生理盐水后和MB注入后 30、60、1 2 0、1 80分钟血液动力学 ,分析动脉、混合静脉血气和动脉血乳酸 ,计算氧合。结果　MB注入明显增加感染性休克犬平均动脉压 (MAP)、体血管阻力 (SVR)和左心室作功(LVSW) ,不影响肺动脉压 (PAP)、肺动脉嵌顿压 (PAOP)、心输出量 (CO)、氧输送 (DO2 )和氧摄取(VO2 ) (P >0 0 5)。MAP、SVR和LVSW的增高以 30分钟最为明显 ,分别从 (96 4± 1 6 6)mmHg、(4332 5± 75 8)dyne .sec .cm- 5、(1 4 0± 1 8) g .m升高到 (1 2 9 9± 1 8 3)mmHg、(52 67 1± 94 6)dyne.sec .cm- 5、(1 7 1± 2 6)g .m(P <0 0 1 ) ,1 80分钟基本回复。内毒素注入增加动脉血乳酸 (P <0 0 5) ;MB注入降低乳酸 ,1 2 0分钟最明显 ,从 (4 3± 1 3)mmol/L下降到 (3 4± 1 2 )mmol/L(P <0 0 5)。结论　MB注入改善感染性休克血液动力学 ,降低血乳酸 ,不影响氧合     

亚甲蓝类似物的合成及其与牛血清白蛋白相互作用的研究

目的 合成3种亚甲蓝类似物3,7-二(二正丙胺基)-吩噻嗪-5-鎓碘化物、3,7-二(二正丁胺基)-吩噻嗪-5-鎓碘化物和3,7-二(二正戊胺基)-吩噻嗪-5-鎓碘化物,并研究模拟生理条件下这3种亚甲蓝类似物与牛血清白蛋白的相互作用.方法 通过1H-NMR及MS对这3种亚甲蓝类似物进行结构表征,应用荧光光谱法研究了模拟生理条件下这3种亚甲蓝类似物与牛血清白蛋白的相互作用.结果 3种亚甲蓝类似物和BSA相互作用形成了蛋白质-药物复合物并猝灭其固有荧光,亚甲蓝类似物与BSA相互作用过程的ΔG0<0,亚甲蓝类似物与BSA荧光残基间的距离r均小于7 nm.结论 亚甲蓝类似物和BSA相互作用的猝灭机制为静态猝灭,二者之间的反应是自发进行的.非辐射能量转移理论表明BSA与亚甲蓝类似物之间存在非辐射能量转移.同步荧光光谱和三维荧光光谱的研究结果表明,亚甲蓝类似物与BSA的相互作用导致BSA构象发生变化.     

An alternative application for reuse of leaching residues: Determination of adsorption behaviour for methylene blue and process optimization

In this study, the potential efficiency in adsorption process, as a functional use route, of laterite ore residues (LR) produced from atmospheric acid leaching was evaluated for methylene blue (MB) removal representing cationic organic pollutants discharged in large amounts from industry into aquatic environment. Adsorption process was optimized by using Box-Behnken design based on Response Surface Methodology (RSM) to determine correlation between responses as, adsorption efficiency (%) and adsorption capacity (mg/g), and 3 independent variables with 3 levels (pH, initial MB concentration and LR amount). 98.29 ± 0.63% of adsorption efficiency and 3.072 ± 0.280 mg/g of adsorption capacity were achieved in determined optimum conditions; pH = 5.2, C₀ = 50 mg/L MB and m = 0.40 g. The good agreement between predicted and experimental values observed by comparison of values showed accuracy of model at 95% confidence level. Furthermore, it was determined that nature of adsorption equilibrium complied Freundlich isotherm model and adsorption mechanism was well defined with pseudo-second-order kinetics model. Thus, process exhibited a heterogeneous multilayer adsorption behavior and mainly controlled by chemical reactions. Furthermore, the adsorption mechanism was determined by characterization of residues, before and after adsorption by scanning electron microscopy with energy dispersive of X-ray (SEM?EDAX), Fourier-transform infrared (FTIR) spectroscopy, and X-ray diffraction (XRD).     

In vivo lymphatic targeting of methylene blue with microemulsion and multiple microemulsion

Three formulations including methylene blue solution (MB-S), MB water-in-oil microemulsion (MB-ME), and MB multiple microemulsion (MB-MME) were prepared with the aim to evaluate whether the three formulations can carry MB target to regional lymph nodes and show lymphatic tropism after subcutaneous administration. The pseudo-ternary phase diagrams were also studied. The morphology of MB-ME and MB-MME was examined by transmission electron microscopy to characterize the microstructure. The particle size and viscosity were also measured. MB concentrations in plasma, lymph nodes, and limb soles were quantitatively analyzed using HPLC. Results show that MME can target MB to regional lymph nodes, and can be employed as a potential lymph tracer in sentinel lymph node biopsy.     

腹腔镜下胆道美蓝显色对婴幼儿梗阻性黄疸的诊断研究

目的探讨腹腔镜下胆道美蓝显色在婴幼儿梗阻性黄疸诊断中的应用价值。方法梗阻性黄疸患者32例,男12例,女20例,年龄52～155 d,平均68d;术前行相关检查未能确定病因。在腹腔镜下行胆道美蓝显色、胆囊置管造影检查。结果32例患儿中10例因胆囊细小干瘪不能提出,造成胆囊穿刺困难,腹部开一小口完成胆囊穿刺,部分行置管照影,其它均在腹腔镜下完成操作,20例胆管、十二指肠美蓝未显色则诊断为先天性胆道闭锁,12例诊断为胆汁淤积症。胆道造影18例诊断为先天性胆道闭锁,其中9例开腹行肝门空肠吻合术（Kasai术）,14例诊断为胆汁淤积症,前后两次诊断结果进行卡方检验,P值〉0.05,结果无显著性差异。结论腹腔镜下胆道美蓝显色在梗阻性黄疸的诊断中具有操作简单、经济、安全、直观的优点,结合胆囊形态及穿刺结果,对于胆道闭锁与胆汁瘀积可以基本作出诊断,但相对胆道照影来仍有一定的局限性,对于没有床边X线机条件的基层医院,仍是诊断婴幼儿梗阻性黄疸的一种较好的方法。     

输尿管双J管在泌尿系疾病中的应用及亚甲蓝在置管时的辅助作用

目的 分析泌尿系疾病中输尿管双J管的应用及亚甲蓝在开放手术置双J管时的辅助作用.方法 326例患者分别采用3种途径(开放手术置入法74例、膀胱镜逆行置入法121例、肾镜/输尿管镜顺行置入法131例)内置双J管,67例开放手术以亚甲蓝辅助下放置双J管.结果 术后1～3个月膀胱镜下拔除双J管,随访患者吻合口通畅,肾积水、肾功能明显好转.部分患者长期留置、定期更换双J管.以亚甲蓝辅助置管者无一例出现拔管困难.结论 泌尿系疾病应用输尿管双J管内引流效果好、操作简单、并发症少,亚甲蓝协助开放手术中双J管放置安全有效.     

联合使用蓝染料和Tc同位素测定胃癌前哨淋巴结的临床意义

目的探讨胃癌中前哨淋巴结（sentinel lymphnode，SLN）概念的适用性，评估前哨淋巴结活检预测胃癌区域淋巴结转移状态的价值及其指导胃癌淋巴结清扫范围的临床意义。方法46例胃癌患者，术前经胃镜于病灶周围黏膜下注入锝标记的锡胶体，术中于病灶周同浆膜下分点注入亚甲蓝，待亚甲蓝显示淋巴结后，将γ探测仪检测到的放射活性最高的淋巴结视为胃癌前哨淋巴结，行术中冰冻免疫组化和常规病理检查或进一步行常规免疫组化染色，分别计算前哨淋巴结诊断胃癌淋巴结转移状态的准确性、敏感性、阴性预测值。结果胃癌前哨淋巴结的检出成功率为100％（46／46）。联合法测定SLN诊断胃癌周围淋巴结转移状态的准确性为100％．敏感性为100％，阴性预测值为100％。结论前哨淋巴结概念适合于胃癌；联合使用蓝染料和锝标锡胶体示踪检测胃癌前哨淋巴结可准确预测胃癌周围淋巴结的转移状态，并可能用于指导胃癌的淋巴结清扫范围。     

滤除白细胞与MB-P法病毒灭活对血浆生化指标的影响

目的探讨应用滤除白细胞及MB-P病毒灭活技术对血浆生化指标的影响。方法随机采集无偿献血合格者血液标本20份,400ml/份。将每份新鲜血浆在制备过程中平均分为试验组与对照组。所有标本同时进行相关生化指标检测。结果试验组经滤除白细胞后FRUC浓度、FHb及CO2增高明显,与对照组比较有显著性差异(P<0.01)。试验组经MB-P法病毒灭活后FRUC与P3+浓度明显增高,FHb下降较快,与对照组及病毒灭活后比较均有显著性差异(P<0.01)。其它生化指标变化不大。结论滤除白细胞及病毒灭活后的血浆制作,对血液生化指标影响均较小或轻,适用于临床输用血(浆)需求。     

亚甲蓝和泛影葡胺定位法切除乳腺微小病变

目的探讨B超下亚甲蓝和泛影葡胺定位法在乳腺微小病灶（直径≤1．0cm）切除和诊断中的作用。方法对136例患者212个乳腺病灶在B超下进行亚甲蓝和泛影葡胺染色定位并手术切除。结果136例患者212病灶全部成功切除,其中7例为早期浸润型导管癌,1例为导管内癌。结论亚甲蓝和泛影葡胺技术可有效提高对乳腺微小病灶切除的准确性以及早期乳腺癌的诊断率,安全性好。     

Toxicity and carcinogenicity studies of methylene blue trihydrate in F344N rats and B6C3F1 mice

Methylene blue trihydrate has a variety of biomedical and biologically therapeutic applications. Groups of 50 male and 50 female rats and mice were administered methylene blue trihydrate in 0.5% aqueous methylcellulose solution by gavage at doses of 0, 5, 25, or 50 mg/kg bw/day (rats) or 0, 2.5, 12.5, and 25 mg/kg bw/day (mice), 5 days per week for 2 years. In rats survival of all dosed groups was similar to that of the vehicle controls, whereas mice exhibited a dose-dependent increase in survival. Rats receiving 25 and 50 mg/kg bw/day and mice receiving 25 mg/kg bw/day developed mild anemia. The incidences of pancreatic islet cell adenoma and adenoma or carcinoma (combined) were increased in all dosed groups of male rats, but increases were statistically significant in 25 mg/kg bw/day males only and the dose–response was non-linear. There was a corresponding increase in the incidence of pancreatic islet cell hyperplasia but statistically significant only in the 50 mg/kg bw/day male rats. There were no significant increases in neoplastic transformation observed in the mice; however, positive trends were noted for adenoma or carcinoma (combined) of the small intestine and malignant lymphoma.     

Preparation and characterization of biochar derived from the fruit seed of Cedrela odorata L and evaluation of its adsorption capacity with methylene blue

Biochar has proven to be an effective tool in several aspects of environmental management; as well as many other applications. Herein we report biochar derived from the capsule-like interior of the seeds of Cedrela odorata L, prepared via pyrolysis at 400 °C. The biochar was characterized using: Elemental analysis, Fourier-transform infrared spectroscopy (FT-IR), Scanning electron microscopy (SEM), X-ray diffraction (XRD), Thermal gravimetric analysis (TGA) and Raman Spectroscopy. Results demonstrate that a biochar of high thermal stability and porosity was obtained, with surface functional groups that enhance the adsorption process. Equilibrium adsorption data were consistent with the Langmuir isotherm model, indicating surface homogeneity, with maximum adsorption capacity of 158.8 mg/g. These results demonstrate that the biochar is an efficient adsorbent for the removal of MB dye from aqueous solution; and suggests its utility for further exploration in removing heavy metals from water as well as its use as a solid support in catalysis.     

Green and ecofriendly bio-removal of methylene blue dye from aqueous solution using biologically activated banana peel waste

Using of bio-wastes in dye adsorption is one of the greenest influential applied techniques for the removal of dyestuff from the industrial effluents and it considered as waste management. In the current study, banana peel waste (BPW) was used as an inexpensive and eco-friendly adsorbent for methylene blue (MB) dye. Mechanical pretreatment of BPW was carried out and followed by biological activation using Rhizopus microspores. MB dye was adsorbed by mechanically pretreated BPW (mBPW) by 31%. Moreover, the day nine fermented BPW fibers (mbBPW) is the best time for R. microspores to complete activation, where adsorption ratio reached to about 96.5%. Likewise, enzymes activity was recorded the highest activity after this period of fermentation, where enzymes activity of cellulase, xylanase, lignin peroxidase, poly phenol oxidase and laccase were 0.75, 0.68, 0.38, 0.55 and 0.32 U/ml, respectively. The FT-IR, SEM and BET were used to observe the effect of treatment on the BPW. Otherwise, the kinetics study is illustrated that the adsorption of MB with mbBPW fitted with pseudo-first-order kinetic models. However, the adsorption parameters indicated that the Langmuir model is better to describe the adsorption of dye with excellent maximum adsorption capacity 991 mg/g. In conclusion, biologically activated BPW is very efficient for dye adsorption as well as waste management.     

离子交换树脂和活性炭分离提取地黄寡糖工艺探讨

目的:考察阴、阳离子交换树脂和活性炭分离提取地黄寡糖成分,优选提取纯化地黄寡糖的最佳工艺。方法:分别应用阴、阳离子交换树脂联用活性炭及单独应用活性炭分离地黄寡糖成分,以重蒸水和不同体积分数的乙醇溶液进行洗脱;并采用HPLC法测定寡糖含量,分析比较HPLC色谱图。结果:阳离子树脂-阴离子树脂-活性炭联用,10%乙醇洗脱物中主要成分为水苏糖;而单用活性炭分离,15%乙醇洗脱物中主要成分是甘露三糖。结论:阳离子树脂-阴离子树脂-活性炭联用及单用活性炭采用不同体积分数乙醇溶液洗脱均可以较好地分离提取寡糖。     

Action of methylene blue and haemoglobin in rabbit aorta in relation to the surface of drug entry

Summary The effect of methylene blue and haemoglobin has been evaluated by using a procedure of restricting drug entry to one surface of rabbit aortic rings. In preparations precontracted with noradrenaline, methylene blue and haemoglobin produced, via the intima, an additional rapid and endothelium-dependent contraction. While haemoglobin was without effect via the adventitia, methylene blue produced a slow endothelium-independent contraction that was diminished but not abolished in reserpine-pretreated animals. This augmentation of the noradrenaline-induced contraction produced by methylene blue via the adventitia in reserpine-pretreated rabbits was observed also with phenylephrine but not with methoxamine. Methylene blue, when added to aortic rings incubated with ³H-(–)-noradrenaline, interfered with the neuronal uptake of this amine and enhanced the outflow of ³H. In conclusion, methylene blue and haemoglobin enhanced noradrenaline-induced contraction by an endothelium-dependent mechanism when applied via the intima. Haemoglobin is devoid of effects via the adventitia but methylene blue has indirect sympathomimetic effects. In experiments using intact rings with no restriction to drug entry it was found that the enhancement by haemoglobin of the tyramine-induced response was markedly less than that observed for noradrenaline for the same level of plateau contraction. This finding indicates that the endothelium-derived relaxing factor is more effective on smooth muscle cell layers in the vicinity of intima than on those close to adventitia. Send offprint requests to E. Morcillo at the above address     

Spectrophotometric determination of methylene blue in biological fluids after ion-pair extraction and evidence of its adsorption on plastic polymers

Methylene blue has gained a renewed interest in medicine. It is now used in combination with visible light as a photoinactivating agent of membrane viruses such as HIV and HBV in single-donor fresh frozen plasma. MB has also been proposed for the prophylaxis and reversal of ifosfamide induced encephalopathy and in septic shock. In spite of its wide use, its pharmacokinetics in humans are little known and an improved methodology for its quantitation in biological fluids was therefore required. We report an improved spectrophotometric determination of methylene blue in whole blood, plasma and urine. Liquid-liquid ion-pair extraction of MB from the biological matrix is achieved by addition of 500 μl (to 1.2 ml whole-blood aliquot) or 125 μl (to 1 ml plasma or 250 μl urine) of 5% sodium hexanesulfonate solution and subsequent extraction with 2.0 ml 1,2-dichloroethane. After centrifugation, the organic layer is assayed at 657 nm. In urine samples, the reduced form leucomethylene blue is also assayed after oxidation to methylene blue (addition of 38 μl HCl 0.5M to 250 μl urine sample and heating on a water bath at 70°C for 1 min) prior to the ion-pairing extraction. Methylene blue can be accurately determined in spiked whole blood and plasma over the working range 0.1–9.1 μg/ml. At higher concentrations, likely to occur in urine (up to 55 μg/ml), the measurements deviate more than 10% from the nominal values. Analysis of spiked samples shows that methylene blue is adsorbed to some extent to polypropylene and polystyrene plastic tubes. This phenomenon is influenced by the biological matrix and is more marked with low volumes of concentrated (>3 μg/ml) samples. At low concentrations, the adsorption is likely to occur but is more difficult to quantitate due to assay variability. This phenomenon should be kept in mind since it could potentially affect the biological activity of methylene blue.     

滤除白细胞与病毒灭活对血浆酶等生化指标的影响

目的探讨滤除白细胞及MB-P病毒灭活技术对血浆酶学及部分生化指标的影响。方法随机采集无偿献血合格者血液标本20份,400ml/份。将每份新鲜血浆在制备过程中平均分为滤白与灭活实验组与对照组。所有标本同时进行相关酶学及部分生化指标检测。结果经滤除白细胞后,实验组LDH、HBDH、TBA下降明显,与对照组比较有显著性差异(P<0.05)。经MB-P法病毒灭活后,实验组CPK、LDH、HBDH、TBIL、IBIL、DBIL、TBA、CREA、URIC下降较快,与对照组比较均有显著性差异(P<0.05)。结论滤除白细胞及病毒灭活后的血浆制作,对血液酶学及部分生化指标影响均较小,符合临床输用血(浆)的要求。     

亚甲蓝和纳米炭混悬液在乳腺癌前哨淋巴结活检中的临床应用

目的:比较亚甲蓝和纳米炭混悬液行乳腺癌前哨淋巴结活检(sentinel lymph nodes biopsy,SLNB)的成功率及临床值.方法:从2007年5月我院收治乳腺癌患者中抽取86例(T1 T2),按注射染料的类型不同随机分为亚甲蓝组和纳米炭混悬液组,其中亚甲蓝组44例和纳米炭混悬液组42例.染料被分4点注入,乳晕周围皮下和乳腺组织中.30分钟后行前哨淋巴结活检术,再行腋窝淋巴结清扫术,并将前哨淋巴结及腋窝淋巴结送病理检查.结果:亚甲蓝组44例检出前哨淋巴结(Sentiel Lymph Nodes,SLN)41例(91.18％),SLNB的灵敏度为92.31％(12/13),准确率为90.91％(40/44),假阴性率为7.69％(1/13),假阳性率为0.纳米炭混悬液组42例,检出SLN40例(95.24％),SLNB的灵敏度为91.67％(11/12),准确率为92.86％(39/42),假阴性率为8.33％(1/12),假阳性率为0.两组间差异无显著性(P＞0.05).结论:采用亚甲蓝行乳腺癌SLNB的准确率和灵敏度与纳米炭混悬液相当,而亚甲蓝费用低廉更具有临床应用前景.