HPAEC-PAD quantification of Haemophilus influenzae type b polysaccharide in upstream and downstream samples

Due to the rapidly increasing introduction of Haemophilus influenzae type b (Hib) and other conjugate vaccines worldwide during the last decade, reliable and robust analytical methods are needed for the quantitative monitoring of intermediate samples generated during fermentation (upstream processing, USP) and purification (downstream processing, DSP) of polysaccharide vaccine components. This study describes the quantitative characterization of in-process control (IPC) samples generated during the fermentation and purification of the capsular polysaccharide (CPS), polyribosyl-ribitol-phosphate (PRP), derived from Hib. Reliable quantitative methods are necessary for all stages of production; otherwise accurate process monitoring and validation is not possible. Prior to the availability of high performance anion exchange chromatography methods, this polysaccharide was predominantly quantified either with immunochemical methods, or with the colorimetric orcinol method, which shows interference from fermentation medium components and reagents used during purification. Next to an improved high performance anion exchange chromatography-pulsed amperometric detection (HPAEC-PAD) method, using a modified gradient elution, both the orcinol assay and high performance size exclusion chromatography (HPSEC) analyses were evaluated. For DSP samples, it was found that the correlation between the results obtained by HPAEC-PAD specific quantification of the PRP monomeric repeat unit released by alkaline hydrolysis, and those from the orcinol method was high (R² = 0.8762), and that it was lower between HPAEC-PAD and HPSEC results. Additionally, HPSEC analysis of USP samples yielded surprisingly comparable results to those obtained by HPAEC-PAD. In the early part of the fermentation, medium components interfered with the different types of analysis, but quantitative HPSEC data could still be obtained, although lacking the specificity of the HPAEC-PAD method. Thus, the HPAEC-PAD method has the advantage of giving a specific response compared to the orcinol assay and HPSEC, and does not show interference from various components that can be present in intermediate and purified PRP samples.     

Preliminary characterization,antioxidant activity in vitro and hepatoprotective effect on acute alcohol-induced liver injury in mice of polysaccharides from the peduncles of Hovenia dulcis

The fresh fleshy peduncles of Hovenia dulcis have been used as a food supplement and traditional herbal medicine for the treatment of liver diseases and alcoholic poisoning for more than a millennium. The objectives of the present study, therefore, were to determine the antioxidant activity of polysaccharides from the peduncles of H. dulcis (HDPS) and to evaluate its hepatoprotective effect on acute alcohol-induced liver injury in mice. HDPS, prepared by hot water extraction, ethanol precipitation and treatment of macroporous resin, was found to be non-starch polysaccharide and mainly composed of galactose, arabinose, rhamnose and galacturonic acid. In in vitro antioxidant assay, HDPS exhibited high superoxide radical scavenging activity, strong inhibition effect on lipid peroxidation and a medium ferrous ion-chelating activity. For hepatoprotective activity in vivo, the administration of HDPS significantly decreased the serum levels of alanine aminotransferase and aspartate aminotransferase, significantly decreased the liver level of malondialdehyde and remarkably restored the liver activities of superoxide dismutase and glutathione peroxidase in alcohol-induced liver injury mice. The results suggested that HDPS had a significant protective effect against acute alcohol-induced liver injury possibly via its antioxidant activity to protect biological systems against the oxidative stress.     

翻白草多糖提取工艺研究

目的优化翻白草多糖的提取工艺。方法采用正交实验方法,以粗多糖得率和总糖含量为评价指标,对翻白草中多糖提取工艺进行筛选。结果综合考虑,最佳提取工艺为煎煮2次,温度70℃,加水12倍量,煎煮时间1 h。结论优选得到的工艺经济、简单、稳定、可行。     

茜草多糖RPS—1,RPS—2和RPS—3组成研究

从茜草中分离得到三种多糖RPS—1、RPS—2和RPS—3,将水解后的多糖转化为糖腈乙酰酯衍生物,用气相色谱分析确定RPS—1、RPS—2和RPS—3的糖基的组成为鼠李糖、阿拉伯糖、木糖、甘露糖、葡萄糖和半乳糖,摩尔比分别为鼠李糖:阿拉伯糖:木糖:甘露糖:葡萄糖:半乳糖=1.0:1.19:0.750.81:1.38:2.60;0.23:0.47:1.47:0.32:2.33:1.0和0.40:1.0:0.58:9.12:11.08:6.13。     

Mitigating base-catalysed degradation of periodate-oxidized capsular polysaccharides: Conjugation by reductive amination in acidic media

Reductive amination coupling an aldehyde-containing polysaccharide, generated by periodate oxidation, with the amino groups in protein has been widely used in the synthesis of glycoconjugate vaccines. The conjugation is often achieved under slightly basic conditions via a Schiff’s base intermediate followed by its reduction with sodium cyanoborohydride. We observed that oxidized capsular polysaccharides such as Streptococcus pneumoniae type 6B (Pn-6B) and Haemophilus influenzae type a (HiA) underwent significant degradation during the conjugation in slightly basic media leading to sub-optimal glycoconjugates. Further study on oxidized Pn-3, Pn-6A, Pn-6C, Pn-2 polysaccharides and dextran provided evidence that the degradation is a result of base-catalysed β-elimination. In contrast to HiA, Pn-2, Pn-3, Pn-6B polysaccharides and dextran, oxidized Pn-6A and Pn-6C polysaccharides were stable under basic conditions due to lack of the leaving group at the β-position of the aldehyde. By performing conjugation of oxidized polysaccharides to bovine serum albumin (BSA) in phosphate buffer at pH 6.0, 6.8, 7.2 and 8.0, we concluded that the reductive amination proceeds best in slightly acidic media, particularly with those β-elimination susceptible polysaccharides.     

黄芪多糖对大鼠慢性难愈合创面的作用及其对PTEN、AKT和VEGF蛋白表达的影响

目的:观察黄芪多糖对慢性难愈合创面的促进愈合作用及其对同源性磷酸酶-张力蛋白(phos-phatase and tension homology deleted on chromosome ten,PTEN)、丝-苏氨酸蛋白激酶(serine-threonine ki-nase,AKT)和血管内皮生长因子(vascul...     

Potentilla anserine L. polysaccharide protects against cadmium-induced neurotoxicity

Cadmium is a toxic metal that can damage the brain and other organs. This study aimed to explore the protective effects of Potentilla anserine L. polysaccharide (PAP) against CdCl₂-induced neurotoxicity in N2a and SH-SY5Y cells and in the cerebral cortex of BALB/c mice. In addition, we aimed to identify the potential mechanisms underlying these protective effects. Relative to CdCl₂ treatment alone, pretreatment with PAP prevented the reduction in cell viability evoked by CdCl₂, decreased rates of apoptosis, promoted calcium homeostasis, decreased ROS accumulation, increased mitochondrial membrane potential, inhibited cytochrome C and AIF release, and prevented the cleavage of caspase-3 and PARP. In addition, PAP significantly decreased the CdCl₂-induced phosphorylation of CaMKII, Akt, and mTOR. In conclusion, PAP represents a potential therapeutic agent for the treatment of Cd-induced neurotoxicity, functioning in part via attenuating the activation of the mitochondrial apoptosis pathway and the Ca²⁺-CaMKII-dependent Akt/mTOR pathway.     

Natural immunity against capsular group X N. meningitidis following an outbreak in Togo, 2007

BackgroundCapsular group X N. meningitidis (MenX) has emerged as a cause of localized disease outbreaks in sub-Saharan Africa, but the human immune response following exposure to MenX antigens is poorly described. We therefore assessed the natural immunity against MenX in individuals who were living in an area affected by a MenX outbreak during 2007 in Togo, West Africa. During 2009, 300 healthy individuals (100 aged 3–5 years, 100 aged 13–19 years and 100 aged 20–25 years) were included in the study, and serum responses were compared with sera from age-matched controls from the U.K. and Burkina Faso.MethodsMenX serum bactericidal antibody (SBA) was measured using rabbit complement, and antibodies against MenX polysaccharide (XPS) and outer membrane vesicles (XOMVs) were quantified by ELISA.ResultsThe proportion of Togolese individuals with an SBA titer of ≥8 against the MenX strain was 29% (95% confidence interval (CI) 18–41) among those aged 3–5 years, 34% (95% CI 9–60) among those aged 13–19 years and 32% (95% CI 24–40) among those aged 20–25 years. These were significantly higher than observed in the control populations from the U.K (range 13–16%) and Burkina Faso (range 2–6%).ConclusionIn Togolese individuals, the concentration of serum IgG against XPS was higher among the two older age groups as compared to the youngest age group. Antibody concentrations against MenX PS correlated significantly with SBA titers. This supports further development of a MenX PS based conjugate vaccine. Further studies are needed to verify the ability of MenX PS to induce SBA in humans.     

桑叶多组分协同降血糖作用

目的 探讨桑叶多糖、黄酮、1 - 脱氧野尻霉素（1 - deoxynojirimycin, DNJ）混合物对糖尿病小鼠降血糖作用。方法 采用链脲佐菌素诱导糖尿病小鼠模型,灌胃桑叶混合物,4周后测定小鼠血糖、胰岛素水平等,观察肝脏、胰腺组织病理变化。结果 桑叶混合物高剂量组具有明显降血糖效应,该组小鼠血糖、血清胰岛素、胰岛素抵抗指数及敏感指数分别是模型组的59.8%、63.5%、30.9%及1.24倍（P＜0.05）,且与阳性对照无统计学差异（P＞0.05）;该组小鼠血清抗氧化酶T - SOD、GSH - Px、CAT活性分别是模型组的1.34倍、1.24倍、2.07倍（P＜0.05）,MDA含量是模型组的79.0%（P＜0.05）,且优于阳性对照组（P＜0.05）。结论 桑叶多糖、黄酮、DNJ混合物可能通过减轻糖尿病小鼠体内氧化损伤,修复受损胰岛细胞、改善机体胰岛素效应细胞的抵抗作用,达到协同降血糖效应。