WO2018188443A1 - 小檗碱或其活性代谢产物在制备预防和/或治疗苯丙酮尿症药物中的应用 - Google Patents

小檗碱或其活性代谢产物在制备预防和/或治疗苯丙酮尿症药物中的应用 Download PDF

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WO2018188443A1
WO2018188443A1 PCT/CN2018/078910 CN2018078910W WO2018188443A1 WO 2018188443 A1 WO2018188443 A1 WO 2018188443A1 CN 2018078910 W CN2018078910 W CN 2018078910W WO 2018188443 A1 WO2018188443 A1 WO 2018188443A1
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berberine
phenylketonuria
phenylalanine
blood
oral
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PCT/CN2018/078910
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French (fr)
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王琰
蒋建东
赵朕雄
马殊荣
寿伽文
李晓阳
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中国医学科学院药物研究所
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Priority to US16/604,794 priority Critical patent/US20200230118A1/en
Priority to EP18785064.9A priority patent/EP3610873B1/en
Publication of WO2018188443A1 publication Critical patent/WO2018188443A1/zh
Priority to ZA2019/07334A priority patent/ZA201907334B/en

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/4353—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom ortho- or peri-condensed with heterocyclic ring systems
    • A61K31/4375—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom ortho- or peri-condensed with heterocyclic ring systems the heterocyclic ring system containing a six-membered ring having nitrogen as a ring heteroatom, e.g. quinolizines, naphthyridines, berberine, vincamine
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/47—Quinolines; Isoquinolines
    • A61K31/4738—Quinolines; Isoquinolines ortho- or peri-condensed with heterocyclic ring systems
    • A61K31/4745—Quinolines; Isoquinolines ortho- or peri-condensed with heterocyclic ring systems condensed with ring systems having nitrogen as a ring hetero atom, e.g. phenantrolines
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P3/00—Drugs for disorders of the metabolism

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  • the present invention relates to the use of berberine or an active metabolite thereof, and a pharmaceutically acceptable salt thereof for the preparation of a medicament for preventing and/or treating phenylketonuria; and belongs to the technical field of medicine.
  • Phenylketonuria is a common amino acid metabolic disease caused by the defect of phenylalanine hydroxylase in the phenylalanine metabolic pathway, which prevents phenylalanine from being converted into tyrosine, resulting in styrene-acrylic acid. Amino acid and phenylpyruvate accumulate and are excreted in large amounts from the urine. Pediatric phenylketonuria is a relatively common autosomal recessive disorder. The child is normal at birth, and after the milk is introduced, the symptoms usually appear at 3 to 6 months, and the symptoms are obvious at the age of 1 year.
  • the main clinical features are mental retardation, mental and neurological symptoms, eczema, skin scratch marks and pigment loss and rat odor, EEG abnormalities. If early diagnosis and early treatment are available, the aforementioned clinical manifestations may not occur, intelligence is normal, and EEG abnormalities can be restored. At present, most of the treatment methods are to reduce the content of phenylalanine in breast milk or infant formula, and there is no clinically preventable/therapeutic drug.
  • Berberine is a natural compound isolated from many different plants, such as berberine, hedgehog, white pheasant, and cork.
  • BBR Berberine
  • berberine has been used as an over-the-counter drug to treat diarrhea without significant adverse effects in patients.
  • berberine has been shown to have a good effect in improving cognition and increasing learning and memory.
  • oral berberine can reduce the content of phenylpyruvate in the blood of suckling rats induced by ⁇ -methylphenylalanine, and increase the content of tyrosine in blood and blood.
  • the ratio of phenylalanine/tyrosine was down-regulated; accordingly, the content of dopa and dopamine in the brain of SD rats with phenylketonuria after treatment was also significantly increased.
  • the intraperitoneal injection of berberine group and pseudo-sterile animal oral berberine group had no obvious effect.
  • the berberine analogs jatrorrhizine and dihydroberberine can also reduce the ratio of phenylpyruvate and phenylalanine/tyrosine in the blood of ICR mice after oral administration.
  • Oral berberine increases the expression of dopa and dopamine in the brain of the phenylketonuria model, and dopamine is one of the most important neurotransmitters in humans.
  • oral berberine can improve the metabolic pathway of phenylalanine, increase the biosynthesis of phenylalanine to tyrosine, and reduce phenylalanine.
  • Biotransformation to phenylpyruvate; an increase in tyrosine synthesis activates the dopa-dopamine synthesis pathway, resulting in a significant increase in dopamine expression in the brain.
  • oral berberine or an active metabolite thereof, and a pharmaceutically acceptable salt thereof can reduce the phenylpyruvate content in the blood of SD suckling mice or ICR mice, phenylalanine in blood/ The ratio of tyrosine, thereby increasing the levels of brain dopa and dopamine, suggesting that berberine or its active metabolites, and pharmaceutically acceptable salts thereof, have the utility of preventing and/or treating phenylketonuria, and their effects
  • the mechanism may be related to the intestinal-brain axis pathway under the regulation of intestinal bacteria.
  • the technical problem to be solved by the present invention is to provide a new class of drugs for preventing and/or treating phenylketonuria.
  • the present invention provides the following technical solutions:
  • the present invention provides the use of berberine or an active metabolite thereof as shown in Structural Formula (I), and a pharmaceutically acceptable salt thereof for the preparation of a medicament for preventing and/or treating phenylketonuria
  • the berberine active metabolite comprises a jatrorrhizine, dihydrogen as shown in structural formula (II), (III), (IV), (V), (VI), (VII), (VIII) Berberine, T. chinensis, berberine, trichostatin, palmatine, arsenic,
  • the pharmaceutically acceptable salt includes a hydrochloride, a sulfate, a hydrobromide, a hydroiodide, a formate, an acetate, an oxalate.
  • phenylketonuria is caused by an increase in phenylpyruvate in the blood.
  • the model of phenylketonuria induced by ⁇ -methylphenylalanine is one of the important experiments for evaluating phenylketonuria.
  • the concentration of phenylpyruvate in the blood of the suckling rats induced by ⁇ -methylphenylalanine was significantly higher than that of the normal control group, and the ratio of phenylalanine/tyrosine was significantly higher than that of the control group; these results indicated that benzene
  • the phenylalanine metabolism of the mice with acetaminophen is abnormal and cannot be metabolized to tyrosine, producing a large amount of phenylpyruvate.
  • the phenylpyruvate in the blood of the suckling mice after oral administration of berberine was significantly lower than that of the model group, and the phenylalanine/tyrosine ratio was significantly lower than that of the model group.
  • Oral berberine could improve the metabolism of phenylalanine, further Can prevent / treat phenylketonuria.
  • Example 1 Therapeutic effect of berberine in alpha-methylphenylalanine induced phenylketonuria model in suckling mice.
  • the concentration of phenylpyruvate in blood and the ratio of phenylalanine/tyrosine in blood are one of the important indicators for evaluating the therapeutic effect of phenylketonuria in suckling mice.
  • SD suckling rats (1 day old) were purchased from Beijing Weitong Lihua Animal Technology Co., Ltd., and animals and mothers were raised together in SPF environment (21 ⁇ 2°C, 12-hour photoperiod), free diet and water during the experiment. .
  • Berberine purchased from the company. Phenylpyruvate, phenylalanine, and tyrosine were purchased from Beijing Suobao Technology Co., Ltd. Alpha-methylphenylalanine was purchased from Nanjing Snow Company.
  • High-performance liquid chromatography-triple quadrupole tandem mass spectrometry (LC-MS/MS 8050, Shimadzu Corporation, Japan) was used to quantitatively determine dopa, phenylalanine, tyrosine, and phenylpyruvate.
  • the experiment used an Alltima C 18 (5 ⁇ m, 4.6 x 150 mm) column with the column temperature maintained at 40 °C.
  • the mobile phase of levodopa was determined to be water-formic acid (100:0.2 v/v) and acetonitrile.
  • mice were divided into 6 groups, including the control group, the model group, the model low-dose oral treatment group, the model high-dose oral treatment group, the model intraperitoneal administration treatment group, and the model pseudo-sterile oral treatment group.
  • Model establishment was started from 2 days old SD suckling mice for 10 days and then treated for 7 days.
  • Control group subcutaneous injection of normal saline
  • Model group subcutaneous injection of ⁇ -methylphenylalanine (50 mg/kg/day) + subcutaneous injection of phenylalanine (200 mg/kg/day);
  • Model low-dose treatment group subcutaneous injection of ⁇ -methylphenylalanine (50 mg/kg/day) + subcutaneous injection of phenylalanine (200 mg/kg/day), oral berberine (100 mg/kg/ day);
  • Model high-dose treatment group subcutaneous injection of ⁇ -methylphenylalanine (50mg/kg/day) + subcutaneous injection of phenylalanine (200mg/kg/day), oral berberine (200mg/kg/ day);
  • Model intraperitoneal administration group subcutaneous injection of ⁇ -methylphenylalanine (50mg/kg/day) + subcutaneous injection of phenylalanine (200mg/kg/day), intraperitoneal injection of berberine (20mg/ Kg/day);
  • Model pseudo-sterile treatment group subcutaneous injection of ⁇ -methylphenylalanine (50mg/kg/day) + phenylalanine (200mg/kg/day), oral berberine (200mg/kg/day) Antibiotic dose, cefadroxil (100 mg/kg/day) + oxytetracycline (300 mg/kg/day) + erythromycin (300 mg/kg/day).
  • the phenylpyruvate content in the blood of the model group was significantly higher than that of the normal control group, indicating that the model was successfully established.
  • the phenylpyruvate was significantly reduced in the blood of the suckling mice 7 days after oral berberine treatment, and the results of oral berberine 100, 200 mg/kg/day were dose-dependent.
  • the phenylalanine/tyrosine ratio in the model group was significantly higher than that in the normal control group, indicating that the model was successfully established.
  • the phenylalanine/tyrosine ratio was significantly reduced in the blood of suckling mice after oral berberine treatment in a dose-dependent manner. There was no significant change in the ratio of phenylalanine/tyrosine in the blood of the intraperitoneal injection group and the pseudo-sterile treatment group.
  • the concentration of dopa in the brain of the model group was significantly lower than that of the normal control group, indicating that the neurotransmitter metabolism in the brain of the SD group in the model group was abnormal.
  • dopa significantly increased in a dose-dependent manner. There was no significant change in the concentration of dopa in the brain between the intraperitoneal injection group and the pseudo-sterile treatment group.
  • the concentration of dopamine in the brain of the model group was significantly lower than that of the normal control group, indicating that the neurotransmitter metabolism in the brain of the SD rats in the model group was abnormal.
  • Dopamine was significantly increased in the brain of suckling mice after oral berberine treatment in a dose-dependent manner. There was no significant change in dopamine concentration in the brain of the intraperitoneal injection group and the pseudo-sterile treatment group.
  • Example 2 Medicinal base reduces the effect of phenylpyruvate in ICR mice.
  • the concentration of phenylpyruvate in blood and the concentration of tyrosine in blood are one of the important indicators for evaluating the therapeutic effect of phenylketonuria in children.
  • ICR mice male, 20 ⁇ 2 g
  • the animals were housed in an SPF environment (21 ⁇ 2 ° C, 12-hour photoperiod), free diet and water during the experiment.
  • Berberine purchased from the company.
  • Phenylpyruvate, phenylalanine, and tyrosine were purchased from Beijing Suobao Technology Co., Ltd.
  • Experimental design The experimental animals were divided into 3 groups, including the control group, oral roots 200mg/kg 12h group, and 24h after oral administration of 200ml/kg.
  • Control group oral saline 0.2mL
  • the concentration of phenylpyruvate in blood and the concentration of tyrosine in blood are one of the important indicators for evaluating the therapeutic effect of phenylketonuria.
  • ICR mice male, 20 ⁇ 2 g were purchased from Beijing Vital Lihua Animal Technology Co., Ltd., and the animals were housed in an SPF environment (21 ⁇ 2 ° C, 12-hour photoperiod), free diet and water during the experiment.
  • Berberine was purchased from the company, and dihydroberberine was purchased from Chengdu Mansite. Phenylpyruvate, phenylalanine, and tyrosine were purchased from Beijing Suobao Technology Co., Ltd.
  • Experimental design The experimental animals were divided into 3 groups, including the control group, the 12h group after oral administration of dihydroberberine 200mg/kg, and the 24h group after oral administration of dihydroberberine 200mg/kg.
  • Control group oral saline 0.2mL

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Abstract

本申请涉及式(I)所示的小檗碱或其活性代谢产物、以及其药学上可接受的盐在制备预防和/或治疗苯丙酮尿症药物中的应用。

Description

小檗碱或其活性代谢产物在制备预防和/或治疗苯丙酮尿症药物中的应用 技术领域
本发明涉及小檗碱或其活性代谢产物、以及其药学上可接受的盐在制备预防和/或治疗苯丙酮尿症药物中的应用；属于医药技术领域。
背景技术
苯丙酮尿症(PKU)是一种常见的氨基酸代谢病，是由于苯丙氨酸代谢途径中的苯丙氨酸羟化酶缺陷，使得苯丙氨酸不能转变成为酪氨酸，导致苯丙氨酸及苯丙酮酸蓄积，并从尿中大量排出。小儿苯丙酮尿症是较为常见的一种常染色体隐性遗传性疾病。患儿出生时正常，随着进奶以后，一般在3～6个月时，即可出现症状，1岁时症状明显。临床主要特征为智力低下、精神神经症状、湿疹、皮肤抓痕征及色素脱失和鼠气味、脑电图异常等。如果能得到早期诊断和早期治疗，则前述临床表现可不发生，智力正常，脑电图异常也可得到恢复。目前，治疗方法大多数是降低母乳或婴儿奶粉中苯丙氨酸的含量，临床上尚无可预防/治疗的药物。
最近的证据表明，肠道菌的变化可以影响动物中枢神经系统的活动，导致脑功能和行为的改变。实验结果支持了肠-脑轴的假说。但是，有关宿主与细菌之间通信的分子机制与化学基础仍不清楚。肠道菌可能是通过微生物产生代谢产物调控大脑功能，这些代谢物影响了神经递质的生物合成，如5-羟色胺，从而影响宿主生理功能。因此，阐明肠-脑通路之间的作用方式，可能会帮助我们了解肠道细菌新功能并有利于发现治疗中枢神经系统紊乱疾病的新方法。
小檗碱(Berberine,BBR)是一个天然化合物，从多种不同植物中分离得到，如黄连，刺檗，白毛茛，黄柏等。几十年来，小檗碱一直作为非处方药物治疗腹泻而病人没有明显的不良反应。自2004年以来，我们发现了小檗碱能够作为新药用于治疗高脂血症和II型糖尿病，其临床疗效已经被国内和国外多个研究组证实。同时，在啮齿动物模型的实验中，小檗碱已经被证明有提高认知和增加学习记忆的良好作用。本专利描述的是口服小檗碱能够降低α-甲基苯丙氨酸诱导的苯丙酮尿症模型乳鼠血中的苯丙酮酸的含量，同时升高血中酪氨酸的含量、使血中苯丙氨酸/酪氨酸比值下调；相应地，治疗后的苯丙酮尿症模型SD乳鼠脑中多巴和多巴胺的含量也显著性升高。而腹腔注射小檗碱组和伪无菌动物口服小檗碱组则没有明显药效。小檗碱类似物药根碱和二氢小檗碱经口服后，同样可以降低ICR小鼠血中苯丙酮酸、苯丙氨酸/酪氨酸的比值。
口服小檗碱增加了苯丙酮尿症模型乳鼠大脑中多巴和多巴胺的表达，而多巴胺是人类最重要的神经递质之一。目前，临床上用于治疗小儿苯丙酮尿症缺乏有效药物，而口服小檗碱可以改善苯丙氨酸的代谢通路，增加了苯丙氨酸向酪氨酸的生物合成、减少苯丙氨酸向苯丙酮酸的生物转化；酪氨酸合成的增加，激活了多巴-多巴胺的合成通路，使得大脑中多巴胺的表达显著性增大。因此，在本专利中描述了口服小檗碱或其活性代谢产物、以及其药学上可接受的盐可以降低SD乳鼠或ICR小鼠血中苯丙酮酸的含量、血中苯丙氨酸/酪氨酸的比值，从而增加大脑多巴和多巴胺的含量，提示小檗碱或其活性代谢产物、以及其药学上可接受的盐具有预防和/或治疗苯丙酮尿症的应用，而其作用机制可能与肠道菌调控下的肠-脑轴通路有关。
发明内容
本发明要解决的技术问题是提供一类新的预防和/或治疗苯丙酮尿症的药物。
为解决本发明的技术问题，本发明提供如下技术方案：
本发明提供了如结构式(I)所示的小檗碱或其活性代谢产物、以及其药学上可接受的盐在制备预防和/或治疗苯丙酮尿症药物中的应用
Figure PCTCN2018078910-appb-000001
Figure PCTCN2018078910-appb-000002
其中，所述的小檗碱活性代谢产物包括如结构式(II)、(III)、(IV)、(V)、(VI)、(VII)、(VIII)所示的药根碱、二氢小檗碱、唐松草分定、小檗红碱、曲亚甲基小檗碱、巴马汀、非洲防己碱，
Figure PCTCN2018078910-appb-000003
其中，所述的药学上可接受的盐包括盐酸盐、硫酸盐、氢溴酸盐、氢碘酸盐、甲酸盐、乙酸盐、草酸盐。
进一步的，所述的苯丙酮尿症是由血中苯丙酮酸升高引起的。
有益技术效果
使用α-甲基苯丙氨酸诱导的苯丙酮尿症模型乳鼠是评价苯丙酮尿症的重要实验之一。
使用α-甲基苯丙氨酸诱导后的乳鼠，其血中苯丙酮酸浓度显著性高于正常对照组，苯丙氨酸/酪氨酸比值显著性高于对照组；这些结果说明苯丙酮尿症乳鼠的苯丙氨酸代谢出现异常，不能代谢为酪氨酸，生成大量苯丙酮酸。口服小檗碱后的乳鼠血中苯丙酮酸显著性低于模型组，苯丙氨酸/酪氨酸比值显著性低于模型组，口服小檗碱可以改善苯丙氨酸的代谢，进一步的可以预防/治疗苯丙酮尿症。
附图说明：
图1.苯丙酮尿症乳鼠经小檗碱治疗后血中苯丙酮酸
图2.苯丙酮尿症乳鼠经小檗碱治疗后血中苯丙氨酸/酪氨酸比值测定
图3.苯丙酮尿症乳鼠经小檗碱治疗后脑中多巴结果
图4.苯丙酮尿症乳鼠经小檗碱治疗后脑中多巴胺结果
图5.ICR小鼠口服药根碱后血中酪氨酸
图6.ICR小鼠口服药根碱后血中苯丙酮酸
图7.ICR小鼠口服二氢小檗碱后血中苯丙酮酸
图8.ICR小鼠口服二氢小檗碱后血中酪氨酸
具体实施方式
实施例1.小檗碱在α-甲基苯丙氨酸诱导的苯丙酮尿症模型乳鼠中的治疗作用。
血中苯丙酮酸浓度和血中苯丙氨酸/酪氨酸比值是评价苯丙酮尿症乳鼠的治疗效果的重要指标之一。
1、实验动物、仪器及药品试剂
SD乳鼠(1日龄)从北京维通利华动物技术有限公司购买，动物与母鼠共同饲养在SPF级环境下(21±2℃，12小时光照周期)，在实验期间自由饮食和饮水。小檗碱、从百灵威公司购买。苯丙酮酸、苯丙氨酸、酪氨酸从北京索莱宝科技公司购买。α-甲基苯丙氨酸从南京斯诺凯公司购买。
2、实验仪器和分析方法
采用高效液相色谱-三重四级杆串联质谱(LC-MS/MS 8050，日本岛津公司)，定量测定多巴、苯丙氨酸、酪氨酸、苯丙酮酸。实验使用Alltima C 18(5μm，4.6x150mm)色谱柱，柱温保持在40℃。测定左旋多巴的流动相为水-甲酸(100:0.2v/v)和乙腈。采用梯度洗脱(A:B,0min,90:10；2.5min,80:20；2.51min,5:95；5min,5:95；5.01min,90:10；8min 90:10)，流速为0.8mL/min。定量采用多反应监测(MRM)模式，多巴定量离子对为198.15→152.05(m/z)。测定苯丙氨酸、酪氨酸、苯丙酮酸采用Alltima C 18(5μm，4.6x150mm)色谱柱，柱温为40℃。流动相为含有水-甲酸(100:0.1v/v)和甲醇。采用梯度洗脱(A:B,0min,90:10；1min,90:10；1.01min,60:40；5min,5:95；7min,5:95；7.01min 90:10；10min 90:10)流速为0.8mL/min。定量采用多反应监测(MRM)模式，定量离子对为苯丙氨酸165.85→120.20；酪氨酸182.00→136.10；苯丙酮酸163.00→91.00。
3.实验设计及动物分组
实验设计：实验动物分为6组，包括对照组、模型组、模型低剂量口服治疗组、模型高剂量口服治疗组、模型腹腔给药治疗组、模型伪无菌口服治疗组。模型建立从2日龄SD乳鼠开始给药10天，然后治疗7天。
动物分组：
(1)对照组：皮下注射生理盐水
(2)模型组：皮下注射α-甲基苯丙氨酸(50mg/kg/天)+皮下注射苯丙氨酸(200mg/kg/天)；
(3)模型低剂量治疗组：皮下注射α-甲基苯丙氨酸(50mg/kg/天)+皮下注射苯丙氨酸(200mg/kg/天)，口服小檗碱(100mg/kg/天)；
(4)模型高剂量治疗组，皮下注射α-甲基苯丙氨酸(50mg/kg/天)+皮下注射苯丙氨酸(200mg/kg/天)，口服小檗碱(200mg/kg/天)；
(5)模型腹腔给药治疗组，皮下注射α-甲基苯丙氨酸(50mg/kg/天)+皮下注射苯丙氨酸(200mg/kg/天)，腹腔注射小檗碱(20mg/kg/天)；
(6)模型伪无菌治疗组，皮下注射α-甲基苯丙氨酸(50mg/kg/天)+苯丙氨酸(200mg/kg/天)，口服小檗碱(200mg/kg/天)；抗生素剂量，头孢羟氨苄(100mg/kg/天)+土霉素(300mg/kg/天)+红霉素(300mg/kg/天)。
4.结果
如图1所示，建模10天后，模型组乳鼠血中苯丙酮酸含量显著性高于正常对照组，说明模型建立成功。在口服小檗碱治疗7天后的乳鼠血中，苯丙酮酸显著性降低，并且口服小檗碱100、200mg/kg/天两组结果呈剂量依赖性。腹腔注射治疗组(20mg/kg/天)和伪无菌治疗组(同时口服小檗碱)血中的苯丙酮酸无显著性变化，说明只有口服治疗有效，可能是小檗碱刺激肠道菌后产生的作用。
如图2所示，模型组血中苯丙氨酸/酪氨酸比值显著性高于正常对照组，说明模型建立成功。同样，在口服小檗碱治疗后的乳鼠血中，苯丙氨酸/酪氨酸比值显著性降低，并且呈剂量依赖性。腹腔注射治疗组和伪无菌治疗组血中的苯丙氨酸/酪氨酸比值无显著性变化。
如图3所示，模型组脑中多巴浓度显著性低于正常对照组，说明模型组SD乳鼠脑中神经递质代谢异常。在口服小檗碱治疗后的乳鼠脑中，多巴显著性回升，并且呈剂量依赖性。腹腔注射组和伪无菌治疗组脑中多巴浓度无显著性变化。
如图4所示，模型组脑中多巴胺浓度显著性低于正常对照组，说明模型组SD乳鼠脑中神经递质代谢异常。在口服小檗碱治疗后的乳鼠脑中，多巴胺显著性回升，并且呈剂量依赖性。腹腔注射组和伪无菌治疗组脑中多巴胺浓度无显著性变化。
实施例2.药根碱降低ICR小鼠体内苯丙酮酸的作用。
血中苯丙酮酸浓度和血中酪氨酸浓度是评价小儿苯丙酮尿症治疗效果的重要指标之一。
1、实验动物、仪器及药品试剂
ICR小鼠(雄性，20±2g)从北京维通利华动物技术有限公司购买，动物饲养在SPF级环境下(21±2℃，12小时光照周期)，在实验期间自由饮食和饮水。小檗碱、从百灵威公司购买。苯丙酮酸、苯丙氨酸、酪氨酸从北京索莱宝科技公司购买。
2.实验设计及动物分组
实验设计：实验动物分为3组，包括对照组、口服药根碱200mg/kg 12h组，口服药根碱200mg/kg后24h组。
动物分组：
(1)对照组：口服生理盐水0.2mL；
(2)单次口服药根碱200mg/kg后12h组：口服药根碱200mg/kg；
(3)单次口服药根碱200mg/kg后24h组：口服药根碱200mg/kg。
3.实验结果
如图5所示，ICR小鼠口服药根碱后，血中酪氨酸显著性升高，说明药根碱可以改善苯丙氨酸的代谢，有治疗苯丙酮尿症的作用。
如图6所示，ICR小鼠口服药根碱后，血中苯丙酮酸显著性降低，说明药根碱可以改善苯丙氨酸的代谢，有治疗苯丙酮尿症的作用。
实施例3.二氢小檗碱降低ICR小鼠体内苯丙酮酸的作用。
血中苯丙酮酸浓度和血中酪氨酸浓度是评价苯丙酮尿症治疗效果的重要指标之一。
1、实验动物、仪器及药品试剂
ICR小鼠(雄性，20±2g)从北京维通利华动物技术有限公司购买，动物饲养在SPF级环境下(21±2℃，12小时光照周期)，在实验期间自由饮食和饮水。小檗碱从百灵威公司购买，二氢小檗碱从成都曼思特公司购买。苯丙酮酸、苯丙氨酸、酪氨酸从北京索莱宝科技公司购买。
2.实验设计及动物分组
实验设计：实验动物分为3组，包括对照组、口服二氢小檗碱200mg/kg后12h组，口服二氢小檗碱200mg/kg后24h组。
动物分组：
(1)对照组：口服生理盐水0.2mL；
(2)单次口服二氢小檗碱100mg/kg后12h组：口服药根碱100mg/kg；
(3)单次口服二氢小檗碱100mg/kg后24h组：口服药根碱100mg/kg。
3.实验结果
如图7所示，ICR小鼠口服二氢小檗碱后，血中酪氨酸显著性升高，说明二氢小檗碱可以改善苯丙氨酸的代谢，有治疗苯丙酮尿症的作用。
如图8所示，ICR小鼠口服二氢小檗碱后，血中苯丙酮酸显著性降低，说明二氢小檗碱可以改善苯丙氨酸的代谢，有治疗苯丙酮尿症的作用。

Claims (4)

  1. 如结构式(I)所示的小檗碱或其活性代谢产物、以及其药学上可接受的盐在制备预防和/或治疗苯丙酮尿症药物中的应用
    Figure PCTCN2018078910-appb-100001
  2. 根据权利要求1的应用，其特征在于，所述的小檗碱活性代谢产物包括如结构式(II)、(III)、(IV)、(V)、(VI)、(VII)、(VIII)所示的药根碱、二氢小檗碱、唐松草分定、小檗红碱、去亚甲基小檗碱、巴马汀、非洲防己碱，
    Figure PCTCN2018078910-appb-100002
  3. 根据权利要求1的应用，其特征在于，所述的药学上可接受的盐包括盐酸盐、硫酸盐、氢溴酸盐、氢碘酸盐、甲酸盐、乙酸盐、草酸盐。
  4. 根据权利要求1的应用，其特征在于，所述的苯丙酮尿症是由血中苯丙酮酸升高引起的。
PCT/CN2018/078910 2017-04-11 2018-03-14 小檗碱或其活性代谢产物在制备预防和/或治疗苯丙酮尿症药物中的应用 WO2018188443A1 (zh)

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