CN104507321B - 奶粉类及其制造方法 - Google Patents
奶粉类及其制造方法 Download PDFInfo
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- CN104507321B CN104507321B CN201280074987.0A CN201280074987A CN104507321B CN 104507321 B CN104507321 B CN 104507321B CN 201280074987 A CN201280074987 A CN 201280074987A CN 104507321 B CN104507321 B CN 104507321B
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- Prior art keywords
- angiogenin
- hydrolysate
- milk powder
- cystatin
- powder class
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Abstract
本发明涉及一种奶粉类，其包含1.4～24mg/15g的血管生成素和/或血管生成素的水解产物，以及相对于血管生成素和/或血管生成素水解产物的质量比为0.03～1.3的抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物。
Description
技术领域
本发明涉及新的奶粉类及其制造方法。奶粉类包含特定的乳成分，并且可用于预防和治疗各种骨疾病例如骨质疏松、骨折、风湿症和关节炎。
背景技术
近年来，在全球范围内伴随着社会老龄化等，各种骨疾病，例如骨质疏松、骨折和腰疼增加，并且已变为严重的社会问题。这些疾病由钙摄入不足，钙吸收能力下降，闭经期后激素失衡等引起。据认为，从生命早期开始通过活化成骨细胞和骨形成尽可能地增加体内骨量，并且增加最大骨量和骨强度(骨密度+骨质)在预防各种骨疾病，例如骨质疏松、骨折和腰疼方面是有效的。注意术语“骨质”涉及骨显微结构、代谢更新(metabolicturnover)、微裂纹和钙化。据称各种骨疾病，例如骨质疏松、骨折和腰疼可通过抑制破骨细胞的骨吸收来预防。骨始终以平衡的方式反复吸收和形成(重塑)。然而，当由于闭经期后激素平衡变化等引起骨吸收超过骨形成时，可发生各种骨疾病，例如骨质疏松、骨折和腰疼。因此，通过抑制破骨细胞的骨吸收和维持骨强度在恒定水平可使骨得到强化。
考虑到上述情形，为了强化骨，已摄取了单独添加钙盐，例如碳酸钙、磷酸钙或乳酸钙的钙盐或者天然钙制品，例如乳清钙、牛骨粉或鸡蛋壳的药物、饮食品或饲料等。还使用包含此类钙制品以及具有钙吸收促进效果的物质，例如酪蛋白磷肽或寡聚多糖的药物、饮食品或饲料等来强化骨。然而，当摄取包含钙盐或天然钙制品的饮食品时钙吸收率为50％以下，并且摄取的大部分钙可能排出体外而没有被吸收。另外，即使钙被吸收入体内，由于对骨的亲和性可根据其形式或同时摄取的营养成分的类型而不同，也未必显示骨代谢改善效果或骨强化效果。作为用于治疗骨质疏松或强化骨的药物已知雌激素制品、活性维生素D3制品、维生素K2制品、双膦酸盐制品和降钙素制品等，并且还开发了新型药物例如抗RANKL抗体。然而，这些药物可具有副作用例如耳鸣、头痛或食欲不振。另外，从安全和成本等观点，目前上述物质处于不能将其添加到饮食品的状况。因此，已需要开发根据各种骨疾病例如骨质疏松、骨折和腰疼的性质，可长期经口摄取，通过促进骨形成和抑制骨吸收来增加骨强度，并且可期望具有预防或治疗各种骨疾病的效果的此类饮食品。
现有技术文献
专利文献
[专利文献1]JP-A-H08-151331
[专利文献2]JP-A-H10-7585
[专利文献3]JP-A-2000-281587
发明内容
发明要解决的问题
本发明的目的是提供可用于预防和治疗各种骨疾病例如骨质疏松、骨折、风湿症和关节炎的奶粉类。
用于解决问题的方案
本发明人已发现通过摄取包含血管生成素和/或血管生成素的水解产物，并相对于血管生成素和/或血管生成素的水解产物以特定的质量比包含抑半胱氨酸蛋白酶蛋白(cystatin)和/或抑半胱氨酸蛋白酶蛋白的水解产物的奶粉类可有效地增加骨密度。该发现导致发明的完成。
具体地，本发明包括以下方面：
(1)一种奶粉类，其包含1.4～24mg/15g的量的血管生成素和/或血管生成素的水解产物，和与血管生成素和/或血管生成素的水解产物的质量比为0.03～1.3的抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物。
(2)一种预防骨疾病的方法，其包括以15g/天以上的量摄取根据(1)所述的奶粉类。
(3)根据(1)所述的奶粉类的制造方法，其包括将血管生成素和/或血管生成素的水解产物以及抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物与乳原料均匀地混合。
(4)根据(1)所述的奶粉类的制造方法，其包括将血管生成素和/或血管生成素的水解产物以及抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物与乳原料混合，和造粒。
发明的效果
本发明的奶粉类显示骨强化效果，并且可用于预防和治疗各种骨疾病例如骨质疏松、骨折、风湿症和关节炎。
具体实施方式
本发明的奶粉类的特征在于奶粉类以特定量包含血管生成素和/或血管生成素的水解产物，并且相对于血管生成素和/或血管生成素的水解产物以特定质量比进一步包含抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物。
奶粉类一般包含约22.5～90μg/g(0.34～1.35mg/15g)的量的血管生成素和/或血管生成素的水解产物，和约45～113μg/g(0.68～1.70mg/15g)的量的抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物。
相反，发明的奶粉类添加有血管生成素和/或血管生成素的水解产物以及抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物，并且所述奶粉类包含1.4～24mg/15g的量的血管生成素和/或血管生成素的水解产物，和相对于血管生成素和/或血管生成素的水解产物的质量比为0.03～1.3的抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物。
作为本发明的奶粉类中包含的血管生成素和/或血管生成素的水解产物以及抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物，可使用由哺乳动物，例如人、奶牛(cow)、水牛(buffalo)、山羊或绵羊的乳制备的包含血管生成素和/或血管生成素的水解产物的级分；由哺乳动物，例如人、奶牛、水牛、山羊或绵羊制备的包含抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物的级分；由基因工程制造的包含血管生成素和/或血管生成素的水解产物的级分；从血液或器官纯化的血管生成素和/或血管生成素的水解产物；从血液或器官纯化的抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物等。还可使用商购可得的纯化的血管生成素或抑半胱氨酸蛋白酶蛋白试剂。
本发明的奶粉类可包含通过使用一种或多种蛋白酶消化包含血管生成素的级分、血管生成素试剂、包含抑半胱氨酸蛋白酶蛋白的级分、抑半胱氨酸蛋白酶蛋白试剂等获得的血管生成素的水解产物或抑半胱氨酸蛋白酶蛋白的水解产物。
本发明的奶粉类可包含通过直接从乳或来源于乳的材料，例如脱脂乳或乳清提取包含血管生成素和/或血管生成素的水解产物以及抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物的级分制备的蛋白材料。例如，该蛋白材料可如下制备。具体地，使乳或来源于乳的材料与阳离子交换树脂接触，并以0.1～2.0M的盐浓度洗脱吸附到树脂的乳来源的蛋白，使用反渗透膜、电渗析膜、超滤膜或微滤膜等脱盐和浓缩，并任选地使用蛋白酶，例如胰蛋白酶、胰酶、胰凝乳蛋白酶、胃蛋白酶、木瓜蛋白酶、激肽释放酶、组织蛋白酶、嗜热菌蛋白酶或V8蛋白酶进行蛋白水解至分子量为8000以下。当使用蛋白酶进行蛋白水解时，分子量的下限优选为500以上。由此获得的蛋白材料可通过冷冻干燥或喷雾干燥等干燥，并且干燥产物可并入奶粉类中。
本发明的奶粉类通过将血管生成素和/或血管生成素的水解产物以及抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物、包含血管生成素和/或血管生成素的水解产物以及抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物的蛋白材料等添加到乳原料，以使奶粉类包含1.4～24mg/15g的量的血管生成素和/或血管生成素的水解产物，并且相对于血管生成素和/或血管生成素的水解产物以0.03～1.3的质量比包含抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物。
如下述试验例中所示，当如上所述奶粉类包含血管生成素和/或血管生成素的水解产物以及抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物时，可获得与单独摄取血管生成素和/或血管生成素的水解产物或抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物的情况相比更加有效的骨强化效果。
本发明的奶粉类可以常规方式来制造，只要奶粉类以各自特定的量包含血管生成素和/或血管生成素的水解产物以及抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物即可。根据本发明制造的奶粉类可包含所有奶粉类，例如，脱脂奶粉、部分脱脂奶粉、奶油粉(cream powder)、全脂奶粉、乳清粉、乳矿物浓缩物(milk mineralconcentrate)、干酪粉、WPI、WPC、调制奶粉(modified milk powder)、特殊奶粉(specialmilk powder)等。
例如，本发明的奶粉类通过将血管生成素和/或血管生成素的水解产物添加到乳原料以使奶粉类包含特定量的血管生成素和/或血管生成素的水解产物，将抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物添加到混合物以使抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物与血管生成素和/或血管生成素的水解产物的质量比在上述特定范围内，并且均匀地混合所得混合物来制造。乳原料的实例包括脱脂乳、脱脂奶粉、部分脱脂乳、部分脱脂奶粉、奶油、奶油粉、牛乳、全脂奶粉、脱脂浓缩乳、乳清粉、乳矿物浓缩物、干酪粉、酪蛋白、WPI、WPC、调制奶粉、特殊奶粉等。
本发明的奶粉类还可通过以特定量混合血管生成素和/或血管生成素的水解产物和抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物与乳原料，均匀地混合所得混合物，并以常规方式，例如，通过喷雾干燥、冷冻干燥、真空干燥等浓缩或干燥混合物从混合物除去水来制造。
在该情况下，可以调节奶粉类中的乳脂含量和乳蛋白含量至每非脂固体(non-fat-solid)的乳蛋白含量为34％以下，并且通过浓缩和干燥使奶粉类中的含水量为5％以下。可将造粒步骤等并入本发明中以便改善奶粉类的溶解性。
除了血管生成素和/或血管生成素的水解产物、抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物外，在上述乳原料之外，本发明的奶粉类可添加有通常用于饮食品的原料等，例如糖类、脂质、蛋白、维生素、矿物质、香料、稳定剂、pH调节剂、抗结剂等，并且还可添加有其它骨强化成分例如钙、维生素D、维生素K、异黄酮等。
如下述动物试验中所示，当以每kg体重15g以上的量经口摄取时本发明的奶粉类可强化骨。由于实验动物的摄入相当于根据血液药物浓度的成人摄入(参见MitsuyoshiNakajima(1993)，“Yakkou Hyoka Vol.8”，Hirokawa-Shoten Ltd.，第2-18页)，所以期望可强化骨，并且特别地可通过以每个成人15g/天以上的量摄食本发明的奶粉类可预防或治疗各种骨疾病，例如骨质疏松、骨折、风湿症和关节炎。
以下通过参考例、实施例和试验例更详细地进一步描述本发明。注意以下实施例仅用于说明性目的，不应解释为限制本发明。
参考例1
血管生成素级分的制备(1)
用去离子水彻底洗涤填充有30kg阳离子交换树脂(磺化Chitopearl；由FujiSpinning Co.,Ltd.制造)的柱，然后将1000升未杀菌的脱脂乳(pH 6.7)施加到柱。在用去离子水彻底洗涤柱之后，用0.1～2.0M线性梯度的氯化钠洗脱吸附的蛋白。使用S-Sepharose阳离子交换色谱(由Amersham Bioscientific制造)分级包含血管生成素的洗脱级分，并在90℃下热处理所得包含血管生成素的级分10分钟，并离心除去沉淀物。将包含血管生成素的级分进一步进行凝胶过滤色谱(柱：Superose 12)。使用反向渗透膜使获得的洗脱物脱盐，并冷冻干燥脱盐的洗脱物以获得16.5g血管生成素级分，其血管生成素纯度为90％。重复这些连续操作30次。
参考例2
血管生成素级分的制备(2)
用去离子水彻底洗涤填充有10kg肝素Sepharose(由GE Healthcare制造)的柱，然后将500升未杀菌的脱脂乳(pH 6.7)施加到柱。在用0.5M氯化钠溶液彻底洗涤柱之后，用1.5M氯化钠溶液洗脱吸附的蛋白。使用反渗透膜使洗脱物脱盐，并冷冻干燥脱盐的洗脱物以获得18g血管生成素级分，其血管生成素纯度为5％。重复上述连续操作50次。
参考例3
抑半胱氨酸蛋白酶蛋白级分的制备
在90℃下热处理十万升(100,000升)5％乳清蛋白溶液，并通过离心除去沉淀物。用通过将羧甲基化的木瓜蛋白酶结合至Tresyl-Toyopearl(由Tosoh Corporation制造)制备的载体填充柱。用0.5M氯化钠溶液平衡化后，将上述乳清蛋白溶液施加到柱。然后用0.5M氯化钠溶液和包含Tween20(0.1％)的0.5M氯化钠溶液顺序洗涤柱。之后，用20mM乙酸-0.5M氯化钠溶液洗脱包含抑半胱氨酸蛋白酶蛋白的级分。用1M氢氧化钠溶液立即中和洗脱物。然后使用反渗透膜使洗脱物脱盐，并冷冻干燥脱盐的洗脱物以获得9.6g抑半胱氨酸蛋白酶蛋白级分，其抑半胱氨酸蛋白酶蛋白纯度为90％。重复上述连续操作20次。
包含于奶粉类中的血管生成素和抑半胱氨酸蛋白酶蛋白的测量
根据更改的JP-A-2008-164511中所述的方法测量奶粉类中包含的血管生成素、血管生成素的水解产物、抑半胱氨酸蛋白酶蛋白和抑半胱氨酸蛋白酶蛋白的水解产物的含量。具体地，将二十五毫克(25mg)奶粉类添加到5ml超纯水，并将1/1000当量的甲酸添加到混合物以制备样品溶液。干燥十微升(10μl)样品溶液，并溶解于20μl含8M脲和1mM三(羧乙基)膦(TCEP)的0.1M碳酸氢铵中。在56℃下加热溶液30分钟。在溶液回到室温后，将5μl100mM碘乙酰胺溶液添加到溶液，并在暗处使混合物反应30分钟。在添加54μl超纯水后，将10μl的0.1μg/ml胰蛋白酶和10μl的0.1μg/ml赖氨酰基肽链内切酶添加到混合物。在37℃下使混合物反应16小时。然后通过添加3μl甲酸终止反应，并用作测量用的样品肽溶液。用包含0.1％甲酸、0.02％三氟乙酸(TFA)和2％乙腈的10fmol/μl内标肽溶液稀释样品溶液6倍，并将2.5μl稀释的溶液进行LC/MS/MS分析。
通过使用HPLC系统梯度洗脱分离肽。更具体地，使用在MAGIC2002HPLC系统上的配备有5μl肽捕获(trap)的柱(MAGICC18,0.2mm(ID)×50mm)以2μl/分钟的流速分离肽。使用溶液A(2％乙腈-0.05％甲酸)和溶液B(90％乙腈-0.05％甲酸)作为HPLC洗脱液。在2至65％溶液B的洗脱条件下经20分钟实施梯度洗脱。
作为测量抑半胱氨酸蛋白酶蛋白的对象离子，母体离子(parent ion)为NH2-QVVSGMNYFLDVELGR-COOH(m/z914.4)，和MS/MS靶离子为NH2-FLDVELGR-COOH(m/z948.7)。作为测量血管生成素的对象离子，母体离子为NH2-YIHFLTQHYDAK-COOH(m/z768.8)，和MS/MS靶离子为NH2-FLTQHYDAK-COOH(m/z1122.8)。关于内标肽，母体离子为NH2-ETTVFENLPEK-COOH(其中，P标记有13C和15N)(m/z656.9)，和MS/MS靶离子为NH2-FENLPEK-COOH(其中，P标记有13C和15N)(m/z882.4)。
将系统“LCQ Advantage”用于MS。各蛋白的峰面积由所得色谱计算，并且由相对于内标肽的比计算浓度。
实施例1
将十五克(15g)脱脂奶粉溶解于热水(50℃)中。接下来，将26mg参考例1中获得的血管生成素级分和0.05mg参考例3中获得的抑半胱氨酸蛋白酶蛋白级分与溶液均匀混合，并喷雾干燥混合物以获得奶粉类(实施例制品1)。获得的奶粉类以24mg/15g的量包含血管生成素和/或血管生成素的水解产物，并且奶粉类中抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物与血管生成素和/或血管生成素的水解产物的质量比为0.03。
实施例2
将十五克(15g)脱脂奶粉溶解于热水(50℃)中。接下来，将20mg参考例2中获得的血管生成素级分和1.2mg参考例3中获得的抑半胱氨酸蛋白酶蛋白级分与溶液均匀混合，并喷雾干燥混合物以获得奶粉类(实施例制品2)。获得的奶粉类以1.4mg/15g的量包含血管生成素和/或血管生成素的水解产物，并且奶粉类中抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物与血管生成素和/或血管生成素的水解产物的质量比为1.3。
实施例3
将十五克(15g)脱脂奶粉溶解于热水(50℃)中。接下来，将20mg参考例1中获得的血管生成素级分和1.2mg参考例3中获得的抑半胱氨酸蛋白酶蛋白级分与溶液均匀混合，并喷雾干燥混合物以获得奶粉类(实施例制品3)。获得的奶粉类以18mg/15g的量包含血管生成素和/或血管生成素的水解产物，并且奶粉类中抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物与血管生成素和/或血管生成素的水解产物的质量比为0.1。
比较例1
将十五克(15g)脱脂奶粉溶解于热水(50℃)中。接下来，将18mg参考例2中获得的血管生成素级分和3.2mg参考例3中获得的抑半胱氨酸蛋白酶蛋白级分与溶液均匀混合，并喷雾干燥混合物以获得奶粉类(比较例制品1)。获得的奶粉类以1.3mg/15g的量包含血管生成素和/或血管生成素的水解产物，并且奶粉类中抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物与血管生成素和/或血管生成素的水解产物的质量比为2.8。
比较例2
将十五克(15g)脱脂奶粉溶解于热水(50℃)中。接下来，将30mg参考例1中获得的血管生成素级分和0.02mg参考例3中获得的抑半胱氨酸蛋白酶蛋白级分与溶液均匀混合，并喷雾干燥混合物以获得奶粉类(比较例制品2)。获得的奶粉类以27mg/15g的量包含血管生成素和/或血管生成素的水解产物，并且奶粉类中抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物与血管生成素和/或血管生成素的水解产物的质量比为0.025。
试验例1
通过动物试验测定实施例制品1至3以及比较例制品1和2的骨强化效果。将C3H/HeJ小鼠(5周龄，雄性)用于动物试验。将实施例制品1至3以及比较例制品1和2的每一种添加到热水(50℃)以使奶粉类的含量为30％，并且均匀地搅拌混合物。1周驯化后，将小鼠分为六组(10只小鼠/组)。使用管以每1kg小鼠重量15g/天的量、一天一次2周向小鼠经口投与实施例制品1至3以及比较例制品1和2。对照组不投与任何实施例制品1至3以及比较例制品1和2。投与完成后(第二周)，使用micro-CT(由Rigaku Corporation制造)测量各小鼠右胫骨的骨密度。结果示于表1中。如表1所示，经口投与实施例制品1至3的组与对照组和经口投与比较例制品1或2的比较例组相比显示骨密度显著增加。
表1
| 骨密度(mg/cm3) | |
| 对照组 | 1239±9 |
| 实施例制品1 | 1264±11 |
| 实施例制品2 | 1271±13 |
| 实施例制品3 | 1269±12 |
| 比较例制品1 | 1242±5 |
| 比较例制品2 | 1243±7 |
参考例4
用去离子水彻底洗涤填充有400g阳离子交换树脂(磺化Chitopearl；由FujiSpinning Co.,Ltd.制造)的柱(直径：4cm，高度：30cm)，并以25ml/分钟的流速将40升未杀菌的脱脂乳(pH 6.7)施加到柱。用去离子水彻底洗涤柱后，使用包含0.78M氯化钠的0.02M碳酸盐缓冲液(pH 7.0)洗脱吸附到树脂上的蛋白。使用反向渗透膜脱盐洗脱物，并冷冻干燥脱盐的洗脱物以获得18g粉状蛋白材料(参考例制品4)。
参考例5
将参考例制品4的四克(4g)蛋白材料溶解于800ml水。在添加作为蛋白酶的胰蛋白酶(由Sigma制造)后，从而获得0.03质量％的终浓度，在37℃下使混合物进行酶促处理8小时。通过90℃下热处理5分钟失活蛋白酶后，冷冻干燥混合物以获得3.0g粉状蛋白材料(参考例制品5)。
实施例4
将四十毫克(40mg)参考例制品4与15g脱脂奶粉均匀混合，并使混合物造粒以获得奶粉类(实施例制品4)。获得的奶粉类以2.5mg/15g的量包含血管生成素和/或血管生成素的水解产物，并且奶粉类中抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物与血管生成素和/或血管生成素的水解产物的质量比为0.29。
实施例5
将四十毫克(40mg)参考例制品5与15g脱脂奶粉均匀混合，并使混合物造粒以获得奶粉类(实施例制品5)。获得的奶粉类以2.4mg/15g的量包含血管生成素和/或血管生成素的水解产物，并且奶粉类中抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物与血管生成素和/或血管生成素的水解产物的质量比为0.30。
比较例3
将三十毫克(30mg)参考例制品4和10mg参考例3中获得的抑半胱氨酸蛋白酶蛋白级分与15g脱脂奶粉均匀混合，并使混合物造粒以获得奶粉类(比较例制品3)。获得的奶粉类以2.0mg/15g的量包含血管生成素和/或血管生成素的水解产物，并且奶粉类中抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物与血管生成素和/或血管生成素的水解产物的质量比为5.0。
试验例2
通过动物试验测定实施例制品4和5以及比较例制品3的骨强化效果。将四十只SD雌性大鼠(51周龄)用于动物试验。将实施例制品4和5以及比较例制品3的每一种添加到热水(50℃)以使奶粉类的含量为30％，并均匀搅拌混合物。将大鼠分为五组(8只大鼠/组)。四组进行卵巢切除，剩余一组假手术(sham surgery)。4周恢复期后，使用管以每1kg小鼠重量15g/天、每日六次分剂量向切除卵巢的大鼠经口投与实施例制品4和5以及比较例制品3的每一种。对照组不投与实施例制品4和5和比较例制品3的任一种。4周恢复期后，以与对照组相同的方式饲养进行假手术的大鼠16周。投与完成后(第十六周)，使用micro-CT(由RigakuCorporation制造)测量各大鼠右胫骨的骨密度。结果示于表2中。如表2所示，经口投与实施例制品4和5的组与对照组和经口投与比较例制品3的比较例组相比显示骨密度显著增加。另外，骨密度接近假手术组的骨密度。
表2
| 骨密度(mg/cm3) | |
| 对照组 | 552±9 |
| 假手术组 | 600±10 |
| 实施例制品4 | 597±12 |
| 实施例制品5 | 594±11 |
| 比较例制品3 | 554±10 |
Claims (4)
1.一种奶粉类，其包含1.4～24mg/15g的量的血管生成素和/或血管生成素的水解产物，和与血管生成素和/或血管生成素的水解产物的质量比为0.03～1.3的抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物。
2.根据权利要求1所述的奶粉类在制备通过预防骨疾病的方法预防骨疾病的组合物中的用途，所述方法包括以15g/天以上的量摄取根据权利要求1所述的奶粉类。
3.根据权利要求1所述的奶粉类的制造方法，其包括将血管生成素和/或血管生成素的水解产物以及抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物与乳原料均匀地混合。
4.根据权利要求1所述的奶粉类的制造方法，其包括将血管生成素和/或血管生成素的水解产物以及抑半胱氨酸蛋白酶蛋白和/或抑半胱氨酸蛋白酶蛋白的水解产物与乳原料混合，和将所述混合物造粒。
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| JP2000281587A (ja) * | 1999-03-30 | 2000-10-10 | Snow Brand Milk Prod Co Ltd | 骨吸収抑制剤 |
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| CN104507321A (zh) | 2015-04-08 |
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| PH12015500052A1 (en) | 2015-03-02 |
| BR112015002046A2 (pt) | 2017-07-04 |
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| US20170319666A1 (en) | 2017-11-09 |
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| EP2880983B1 (en) | 2017-07-26 |
| MX366535B (es) | 2019-07-12 |
| KR20150036677A (ko) | 2015-04-07 |
| MY167577A (en) | 2018-09-20 |
| TW201408211A (zh) | 2014-03-01 |
| US20190269763A1 (en) | 2019-09-05 |
| WO2014020681A1 (ja) | 2014-02-06 |
| CA2879995A1 (en) | 2014-02-06 |
| TWI626009B (zh) | 2018-06-11 |
| CA2879995C (en) | 2019-09-03 |
| AU2012386764A1 (en) | 2015-03-05 |
| EP2880983A4 (en) | 2016-01-13 |
| US20160015790A1 (en) | 2016-01-21 |
| PH12015500052B1 (en) | 2015-03-02 |
| KR20190095541A (ko) | 2019-08-14 |
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| EP2880983A1 (en) | 2015-06-10 |
| AU2012386764B2 (en) | 2016-05-05 |
| JPWO2014020681A1 (ja) | 2016-07-11 |
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