基于UPLC-Q-Exactive-MS的菲牛蛭酶解物肽段鉴定与分子特性表征

目的 基于超高效液相色谱-四极杆-静电场轨道阱高分辨质谱（UPLC-Q-Exactive-MS）技术对菲牛蛭酶解物中肽段进行鉴定与表征。方法 采用胰蛋白酶对菲牛蛭进行酶解,利用UPLC-Q-Exactive-MS技术结合Maxquant软件对菲牛蛭酶解物进行分析并对其肽段进行序列鉴定,同时采用生物信息学平台对这些肽段进行生物活性与不良反应预测以及相对分子质量、等电点（pI）、净电荷、亲水性氨基酸比例、不稳定指数等基本分子特性的预测。结果 利用UPLC-Q-Exactive-MS技术联合Maxquant软件在菲牛蛭酶解物中共鉴定出32条肽段,其中肽段AGFAGDDAPR的各项肽段分子特性符合目前已知抗血栓肽的共性特征,鉴定分数为103.83,可信度高;活性概率为0.56,相对分子质量976.01,肽链长度为10;平均亲水性为0.5,亲水性氨基酸残基比例为30%,水溶性良好;pI值为4.21,净电荷为-1;不稳定性指数为20.72,在水中稳定性强;具有抗血栓活性的可能性较大。同时6条肽段SSGETSSIIRR、AGFAGDDAPR、SSGETSSIIR、DSYVGDEAQSKR、GARRER、SIEDQVKR被预测具有抗血管生成活性且无溶血现象,但仍需进一步的合成与活性验证。结论 以菲牛蛭为例,提供了一种快速从酶解物中鉴定动物药肽段序列的方法,以期为动物药的肽类成分的研究提供思路。     

Preventive effect of feeding high-risk infants a casein hydrolysate formula or an ultrafiltrated whey hydrolysate formula. A prospective, randomized, comparative clinical study

In a prospective study of a 1-year birth cohort of 158 high-risk infants the effect of feeding breastmilk, a casein hydrolysate (Nutramigen®) or a new ultrafiltrated whey hydrolysate (Profylac®) on the development of cow milk protein allergy/intolerance (CMPA/CMPI) was assessed and compared. All the infants had biparental or severe single atopic predisposition, the latter combined with cord blood IgE ≥ 0. 5 kU/L. At birth all infants were randomized to Nutramigen or Profylac, which was used when breastfeeding was insufficient or not possible during the first 6 months of life. During the same period this regimen was combined with avoidance of solid foods and cow milk protein. All mothers had unrestricted diets and were encouraged to do breastfeeding only. Moreover, avoidance of daily exposure to tobacco smoking, furred pets and dust-collecting materials in the the bedroom was advised. The infants were followed prospectively from birth to 18 months of age. All possible atopic symptoms were registered and controlled elimination/challenge studies were performed when symptoms suggested CMPA/CMPI. A total of 154 (97%) were followed up and 141 followed the diet strictly. Eighty-eight (62%) of the infants were breastfed for at least 6 months, 20 (14%) were breastfed exclusively, 59 and 62 had varying amounts of Nutramigen or Profylac respectively. CMPA/CMPI was diagnosed in 1/20, 1/59 and 3/62 in the breastfed, the Nutramigen and Profylac groups respectively, but 1 of the latter also had Nutramigen. None of the infants showed reactions against Nutramigen or Profylac. In 4 infants symptoms were provoked by breastmilk when the mother ingested cow milk and in 1 only by cow milk. The incidence of CMPA/CMPI among the infants who followed the dietary prevention programme was 3. 6% (5/141) which was a significant reduction compared to 20% (15/75) in an identically defined high-risk group without dietary preventive measures. None of the infants in the prevention group developed CMPA/CMPI after the age of 6 months. We conclude that feeding breastmilk, an extensively hydrolysed casein formula (Nutramigen) or an ultrafiltrated whey hydrolysate (Profylac) combined with avoidance of solid foods during the first 6 months of life in high-risk infants significantly reduced the cumulative incidence of CMPA/CMPI during the first 18 months of life. No difference was noted whether the infants were fed breastmilk, Nutramigen or Profylac and a diet period of 6 months seems sufficient. Both formulae were well tolerated and accepted by the infants.     

Food sensitive enteropathy: Overview and update

There are two types of food sensitive enteropathy; permanent and temporary. Celiac disease belongs to the former, the temporary food sensitive enteropathies of early childhood to the latter. A food sensitive enteropathy is characterized by an abnormal small intestinal mucosa while having the offending food in the diet; the abnormality is reversed by an elimination diet, only to recur once more on challenge with the relevant food. These disorders are temporary and may follow gastroenteritis. Cow's milk sensitive enteropathy is the most frequent and best known example but soy protein, egg, fish, chicken meat, ground rice and probably gluten may also temporarily damage the small intestinal mucosa in infancy. Treatment is with an elimination diet and protein hydrolysates as a cow's milk substitute. The reason why these enteropathies are temporary has not yet been established.     

Evaluation of a casein and a whey hydrolysate for treatment of cow's-milk-sensitive enteropathy

A casein and a whey hyrolysate were evaluated in the management of 18 children with cow's-milk-sensitive enteropathy. This diagnosis was based upon clinical features, an abnormal small intestinal mucosa, i.e. an enteropathy, and a clinical response to cow's milk elimination. Two infants refused to take the whey hydrolysate. The median weight gain was higher in children given whey hydrolysate (19.4 g/day) than the casein hydrolysate (9.8 g/day). All children responded to cow's milk elimination and most has a significant improvement in small intestinal morphology after a cow's-milk-free diet. There was some advantage for the whey hydrolysate on morphometric analysis of their small intestinal mucosal response.     

Acute and repeated dose (4 weeks) oral toxicity studies of two antihypertensive peptides,RYLGY and AYFYPEL,that correspond to fragments (90–94) and (143–149) from αs1-casein

The Lowpept® is a powdered casein hydrolysate containing the antihypertensive peptides RYLGY and AYFYPEL, two sequences that correspond to α_s1-casein f (90–94) (RYLGY) ¹ and α_s1-casein f (143–149) (AYFYPEL) ¹. To support the safety, Lowpept® has been examined in an acute and in a 4-week repeated dose oral toxicity studies in rats. Powdered casein hydrolysate administered in a single oral gavage dose of 2000 mg/kg resulted in no adverse events or mortality. Also, casein hydrolysate administered as a daily dose of 1000 mg/kg for 4 weeks by gavage resulted in no adverse events or mortality. No evidence or treatment-related toxicity was detected during both studies. Data analysis of body weight gain, food consumption, clinical observations, blood biochemical, haematology, organ weight ratios and histopathological findings did not show significant differences between control and treated groups. It is concluded that the casein hydrolysate containing the peptides RYLGY and AYFYPEL orally administered to rats was safe and that not treatment-related toxicity was detected even at the highest doses investigated in both acute (2000 mg/kg of body weight) and repeated dose (4 weeks) oral (1000 mg/kg of body weight) toxicity studies.     

Collagen hydrolysate for the treatment of osteoarthritis and other joint disorders:a review of the literature

ABSTRACT

Background: There is a need for an effective treatment for the millions of people in the United States with osteoarthritis (OA), a degenerative joint disease. The demand for treatments, both traditional and non-traditional, will continue to grow as the population ages.

Scope: This article reviews the medical literature on the preclinical and clinical research on a unique compound, collagen hydrolysate. Articles were obtained through searches of the PubMed database (www.pubmed.gov) through May 2006 using several pairs of key words (collagen hydrolysate and osteoarthritis; collagen hydrolysate and cartilage; collagen hydrolysate and chondrocytes; collagen hydrolysate and clinical trial) without date limits. In addition, other sources of information, such as abstracts presented at scientific congresses and articles in the German medical literature not available on PubMed, were reviewed and included based on the authors’ judgment of their relevance to the topic of the review.

Findings: According to published research, orally administered collagen hydrolysate has been shown to be absorbed intestinally and to accumulate in cartilage. Collagen hydrolysate ingestion stimulates a statistically significant increase in synthesis of extracellular matrix macromolecules by chondrocytes (?p < 0.05 compared with untreated controls). These findings suggest mechanisms that might help patients affected by joint disorders such as OA. Four open-label and three double-blind studies were identified and reviewed; although many of these studies did not provide key information – such as the statistical significance of the findings – they showed collagen hydrolysate to be safe and to provide improvement in some measures of pain and function in some men and women with OA or other arthritic conditions.

Conclusion: A growing body of evidence provides a rationale for the use of collagen hydrolysate for patients with OA. It is hoped that ongoing and future research will clarify how collagen hydrolysate provides its clinical effects and determine which populations are most appropriate for treatment with this supplement.     

桃红四物汤合五苓散联合人工全髋关节置换治疗陈旧性髋臼骨折并髋关节中心性脱位1例报告

报告一例桃红四物汤合五苓散联合人工全髋关节置换治疗陈旧性髋臼骨折并髋关节中心性脱位。40岁女性,车祸致多发骨折遗留左髋关节疼痛,功能受限,行左侧人工全髋关节置换术。术后48h内头孢替安预防感染,术后当天即兰索拉唑预防应激性溃疡,曲克芦丁脑蛋白水解物预防创伤性水肿,术后第2天加用丹参川芎嗪活血化瘀,低分子肝素钠预防下肢深静脉血栓,患肢保持外展中立位,加压泵促进下肢回流;术后第3天左下肢出现水肿,皮肤小片瘀斑,至第5天下肢肿胀明显,且出现大片瘀斑,按压皮肤有凹陷,恢复缓慢。辨证属瘀水互结,血瘀气滞,水停湿阻,治以活血化瘀,利水消肿,桃红四物汤合五苓散(酒当归、赤芍、三七粉、川芎、桃仁、藏红花、桂枝、炒白术、茯苓、泽泻、牛膝),口服7剂,外用自制冰硝散外敷3天,服药第5剂水肿明显消退,但瘀斑未见明显改善,7剂服毕瘀斑范围略见缩小,水肿基本消退,继续丹参川芎嗪活血化瘀。术后第7天指导患者功能锻炼,术后两周刀口拆线并指导患者在双拐辅助下患肢禁负重下地行走锻炼,术后三周出院,患者术后2个月复查,X线示假体位置良好,术后3个月复查指导患者负重锻炼,关节功能良好,无不适。此例同时应用曲克芦丁脑蛋白水解物、低分子肝素钠等,下肢水肿的治疗效果应为多种药物共同作用,中药具体效果还需进一步研究认证。     

Fast Anxiolytic-Like Effect Observed in the Rat Conditioned Defensive Burying Test,after a Single Oral Dose of Natural Protein Extract Products

Anxiety appears among the most frequent psychiatric disorders. During recent years, a growing incidence of anxiety disorders can be attributed, at least in part, to the modification of our eating habits. To treat anxiety disorders, clinicians use benzodiazepines, which unfortunately display many side effects. Herein, the anxiolytic-like properties of two natural products (αS1–casein hydrolysate and Gabolysat^®) were investigated in rats and compared to the efficacy of benzodiazepine (diazepam). Thus, the conditioned defensive burying test was performed after a unique oral dose of 15 mg/kg, at two time-points (60 min and then 30 min post oral gavage) to show potential fast-onset of anxiolytic effect. Both natural products proved to be as efficient as diazepam to reduce the time rats spent burying the probe (anxiety level). Additionally, when investigated as early as 30 min post oral gavage, Gabolysat^® also revealed a fast-anxiolytic activity. To date, identification of bioactive peptide, as well as how they interact with the gut–brain axis to sustain such anxiolytic effect, still remains poorly understood. Regardless, this observational investigation argues for the consideration of natural compounds in care pathway.     

脑蛋白水解物治疗帕金森病50例效果评估

目的观察脑蛋白水解物治疗帕金森病(PD)的效果。方法 100例PD患者随机分为对照组与治疗组各50例,两组均给予抗胆碱能药物、左旋多巴等药物治疗,治疗组加用脑蛋白水解物120 mg/d。观察两组治疗前后帕金森病Hoehn-Yahr分期、Schwab和英格兰日常生活活动量表及帕金森病综合评分量表(UPDRS)积分改善情况。结果治疗组有效率较对照组明显提高(p<0.01),治疗后的UPDRS积分较治疗前明显下降(p<0.05)。结论脑蛋白水解物治疗PD患者疗效显著。