JP2022532003A - クガイソウ抽出物を有効成分として含む炎症、アレルギー及び喘息の予防用または治療用の組成物、及びその用途 - Google Patents
クガイソウ抽出物を有効成分として含む炎症、アレルギー及び喘息の予防用または治療用の組成物、及びその用途 Download PDFInfo
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Abstract
Description
Balb/c雄マウスを、各群当たり6匹にし、正常群を除いた全ての群に、微細ほこりの構成成分である0.25mg/ml石炭(coal)、10mg/mlフライアッシュ(fly ash)、0.2mg/ml DEP(diesel exhaust particle)の混合物に、ミョウバン(alum)の最終濃度が8%になるように混合し、該微細ほこり混合物を、実験動物の気道及び鼻に、文献に記載されたINT(intra-nazal-trachea)注入(injection)法(Lim et al., Free Radic Biol Med. 25(6), 635-644. (1998))を利用し、実験開始の3日目、6日目に50μlずつ直接注入した。前述の陽性対照群(N-acetylcysteine、Sigma A7250)、実施例試料を、各群に200mg/kgの濃度で、0.5%ナトリウムCMC(carboxymethyl cellulose)、419273、Sigma-Aldrich)溶液に希釈し、毎日(10日)経口投与した。実験開始後11日目に、剖検を進め、気管支肺胞洗浄液(BAL fluid)を回収した。
前記試料に対する総気管支肺胞洗浄液(BAL:bronchoalveolar lavage)における、総細胞数に及ぼす影響を測定した結果を、下記表3に示した。総BAL細胞数は、誘発群に比べ、減少したことを示し、前記試料がいずれも炎症数値低下に寄与することを確認し、さらに比較例1より実施例1-1が、さらに低い総BAL細胞数を示し、実施例1-1に比べ、実施例2処置群において、さらに低い総BAL細胞数を示し、実施例2試料が、さらに高い気管支炎症抑制活性を有することを示した。
前記実験例2の方法と同一に進めた。回収した気管支肺胞洗浄液(BAL:bronchoalveolar lavage)において、ディフ・クイック(Diff-Qick)染色法(Takano et al., Am J Respir Crit Care Med, 156 (1), 36-42. (1997), Hemacolor Rapid staining of blood smear, 1.11661.0001, Merck)で好中球を染色した後で観察した。
前記試料に対する総気管支肺胞洗浄液(BAL:bronchoalveolar lavage)における、総細胞数対比の好中球細胞比率に及ぼす影響を測定した結果を、下記表4に示した。総好中球細胞比率は、誘発群に比べ、低下したことを示し、前記試料が、いずれも炎症数値低減に寄与することを確認し、さらに比較例1より実施例1-1において、低い総気管支肺胞洗浄液(BAL:bronchoalveolar lavage)における、総細胞数対比の好中球細胞比率を示し、実施例1-1に比べ、実施例2において、さらに低い総気管支肺胞洗浄液(BAL:bronchoalveolar lavage)における、総細胞数対比の好中球細胞比率を示し、実施例2試料が、さらに高い気管支炎症抑制活性を有することを示した(表4)。
前記実施例試料の気管支肺胞洗浄液(BAL fluid)における、好中球+/Gr-1+絶対(absolute)細胞数に及ぼす影響を確認するために、下記のように文献に記載された方法を応用し、実験した(Beutner EH., Bacteriological Reviews., 25(1):49-76, ((1961))。
気管支肺胞洗浄液(BAL:bronchoalveolar lavage)において、細胞数を測定することを除いては、前記実験例2の方法と同一に進めた。回収した気管支肺胞洗浄液(BAL fluid)を対象に、蛍光標識が結合されたGr-1抗体(553128、BD Biosciences、San Jose、CA、米国)を利用し、特異的蛍光抗体染色法(specific fluorescent antibody staining method)を進め、FACS(fluorescence-activated cell sorting、BD Biosciences、San Jose、CA、米国)法を利用し、全体白血球(leukocyte)における、好中球+/Gr-1+絶対(absolute)細胞数を測定した。
前記試料の気管支肺胞洗浄液(BAL fluid)における、好中球+/Gr-1+絶対(absolute)細胞数を測定した結果を、下記表5に示した。それらのいずれにおいても、誘発群に比べ、好中球+/Gr-1+絶対(absolute)細胞数が減少したことを確認し、さらに実施例1-1試料に比べ、実施例2試料処置群において、さらに低い好中球+/Gr-1+絶対(absolute)細胞数を示し、実施例2試料が、高い気管支炎症抑制活性を示すことを確認した(表5)。
前記実施例試料の肺(lung)細胞における、CD11b+/Gr-1+絶対(absolute)細胞数に及ぼす影響を確認するために、下記のように文献に記載された方法を応用し、実験した(Beutner EH., Bacteriological Reviews., 25(1):49-76, ((1961))。
気管支肺胞洗浄液(BAL:bronchoalveolar lavage)において、細胞数を測定することを除いては、前記実験例2の方法と同一に進めた。回収した肺(lung)を対象に、蛍光標識が結合されたCD11b抗体(553310、BD Biosciences、San Jose、CA、米国)及びGr-1抗体(553128、BD Biosciences、San Jose、CA、米国)を利用し、特異的蛍光抗体染色法(specific fluorescent antibody staining method)を進め、FACS(fluorescence-activated cell sorting、BD Biosciences、San Jose、CA、米国)法を利用し、全体肺細胞数におけるCD11b+/Gr-1+絶対(absolute)細胞数を測定した。
前記試料の肺(lung)細胞中のCD11b+/Gr-1+絶対(absolute)細胞数を測定した結果を、下記表6に示した。それらのいずれにおいても、誘発群に比べ、CD11b+/Gr-1+絶対(absolute)細胞数が減少したことを確認し、さらに実施例1-1に比べ、実施例2において、さらに低いCD11b+/Gr-1+絶対(absolute)細胞数を示し、実施例2試料が、高い気管支炎症抑制活性を示すことを確認した。
前記実施例試料の肺(lung)細胞における、CD4+/CD3+絶対(absolute)細胞数に及ぼす影響を確認するために、下記のように文献に記載された方法を応用し、実験した(Beutner EH., Bacteriological Reviews., 25(1):49-76, ((1961))。
気管支肺胞洗浄液(BAL:bronchoalveolar lavage)において、細胞数を測定することを除いては、前記実験例2の方法と同一に進めた。回収した肺(lung)を対象に、蛍光標識が結合されたCD4抗体(550280、BD Biosciences、San Jose、CA、米国)及びGr-1抗体(554829、BD Biosciences、San Jose、CA、米国)を利用し、特異的蛍光抗体染色法(specific fluorescent antibody staining method)を進め、FACS(fluorescence-activated cell sorting、BD Biosciences、San Jose、CA、米国)法を利用し、全体肺細胞数におけるCD4+/CD3+絶対(absolute)細胞数を測定した。
前記試料の肺(lung)細胞におけるCD4+/CD3+絶対(absolute)細胞数を測定した結果を、下記表7に示した。それらのいずれにおいても、誘発群に比べ、CD4+/CD3+絶対(absolute)細胞数が減少したことを確認し、さらに実施例1-1に比べ、実施例2において、さらに低いCD4+/CD3+絶対(absolute)細胞数を示し、実施例2試料が、高い気管支炎症抑制活性を示すことを確認した。
前記実施例試料の肺(lung)細胞における、大食細胞+/CD11b+絶対(absolute)細胞数に及ぼす影響を確認するために、下記のように文献に記載された方法を応用し、実験した(Beutner EH., Bacteriological Reviews., 25(1):49-76, ((1961))。
気管支肺胞洗浄液(BAL:bronchoalveolar lavage)において、細胞数を測定することを除いては、前記実験例2の方法と同一に進めた。回収した肺(lung)を対象に、蛍光標識が結合されたCD11b抗体(553310、BD Biosciences、San Jose、CA、米国)を利用し、特異的蛍光抗体染色法(specific fluorescent antibody staining method)を進め、FACS(fluorescence-activated cell sorting、BD Biosciences、San Jose、CA、米国)法を利用し、全体肺細胞数における大食細胞+/CD11b+絶対(absolute)細胞数を測定した。
前記試料の肺(lung)細胞における大食細胞+/CD11b+絶対(absolute)細胞数を測定した結果を、下記表8に示した。それらのいずれにおいても、誘発群に比べ、大食細胞+/CD11b+絶対(absolute)細胞数が減少したことを確認し、さらに実施例1-1試料に比べ、実施例2試料処置群において、さらに低い大食細胞+/CD11b+絶対(absolute)細胞数を示し、実施例2試料が、高い気管支炎症抑制活性を示すことを確認した。
前記実施例試料の気管支肺胞洗浄液(BAL fluid)における、炎症因子発現レベルに及ぼす影響を確認するために、下記のように文献に記載された方法を応用し、実験した(Brandt EB et al., J. Allergy Clin. Immunol., 132(5):1194-1204, (2013))。
気管支肺胞洗浄液(BAL:bronchoalveolar lavage)において、細胞数を測定することを除いては、前記実験例2の方法と同一に進めた。気管支肺胞洗浄液(BAL fluid)において、IL-17A、TNF-αMIP2及びCXCL-1のレベルをELISAで測定した。IL-17A抗体(M1700、R&D Systems、Minneapolis、米国)、TNF-α抗体(MTA00B、R&D Systems、Minneapolis、米国)、MIP2抗体(MM200、R&D Systems、Minneapolis、米国)及びCXCL-1抗体(MKC00B、R&D Systems、Minneapolis、米国)を緩衝溶液希釈し、微細ウェル(micro well)にコーティングした後、4℃で16時間培養した。各ウェル(well)を3回洗浄(washing)緩衝溶液で洗浄した後、10倍希釈した血清を100μlずつ分注した。
下記表9から分かるように、前記試料投与群の炎症因子(IL-17A、TNF-α、MIP2、CXCL-1)は、誘発群に比べて減少した。実施例1-1に比べ、実施例2試料処置群において、さらに低いIL-17A,TNF-α,MIP2,CXCL-1発現を示し、実施例2試料が、高い気管支炎症抑制活性を示すことを確認した。
抽出物(VSM): 20mg
乳糖: 100mg
タルク: 10mg
前述の成分を混合し、気密包に充填して散剤を製造する。
分画物(VSW): 10mg
とうもろこし澱粉: 100mg
乳糖: 100mg
ステアリン酸マグネシウム: 2mg
前述の成分を混合した後、通常の錠剤の製造方法によって打錠し、錠剤を製造する。
分画物(VS25E): 10mg
結晶性セルロース: 3mg
ラクトース: 14.8mg
ステアリン酸マグネシウム: 0.2mg 通常のカプセル剤製造方法により、前述の成分を混合し、ゼラチンカプセルに充填してカプセル剤を製造する。
分画物(VS50E): 10mg
マンニトール: 180mg
注射用滅菌蒸溜水: 2,974mg
Na2HPO4・12H2O: 26mg
通常の注射剤の製造方法により、1アンプル当たり(2ml)、前述の成分含量で製造する。
抽出物(VS75E): 20mg
異性化糖: 10g
マンニトール: 5g
精製水: 適量
通常の液剤の製造方法により、精製水にそれぞれの成分を加えて溶解させ、レモン香を適量加えた後、前述の成分を混合した後、精製水を加え、全体100mlに調節した後、褐色瓶に充填して滅菌させ、液剤を製造する。
製剤例6.健康食品の製造
抽出物(VSE): 1,000mg
ビタミン混合物: 適量
ビタミンAアセテート: 70μg
ビタミンE: 1.0mg
ビタミンB1: 0.13mg
ビタミンB2: 0.15mg
ビタミンB6: 0.5mg
ビタミンB12: 0.2μg
ビタミンC: 10mg
ビオチン: 10μg
ニコチン酸アミド: 1.7mg
葉酸: 50μg
パントテン酸カルシウム: 0.5mg
無機質混合物: 適量
硫酸第1鉄: 1.75mg
酸化亜鉛: 0.82mg
炭酸マグネシウム: 25.3mg
第1リン酸カリウム: 15mg
第2リン酸カルシウム: 55mg
クエン酸カリウム: 90mg
炭酸カルシウム: 100mg
塩化マグネシウム: 24.8mg
前述のビタミン及びミネラル混合物の組成比は、比較的健康食品にふさわしい成分を、望ましい実施例において混合組成したが、その配合比を、本発明の精神及び範囲を外れないと見なされる任意に変形実施してもよい。
抽出物(VSEA): 1,000mg
クエン酸: 1,000mg
オリゴ糖: 100g
梅濃縮液: 2g
タウリン: 1g
精製水加える: 全体900ml
通常の健康飲料製造方法により、前述の成分を混合した後、約1時間85℃で撹拌加熱した後、作られた溶液を濾過し、滅菌された2L容器に取り、密封滅菌した後、冷蔵保管した後、本発明の健康飲料組成物製造に使用する。
Claims (16)
- クガイソウ(Veronicastrum sibiricum L. Pennell,クガイソウ属(Veronicastrum))の抽出物を有効成分として含む、炎症、アレルギー及び喘息を予防または治療するために用いられる薬学組成物。
- 前記クガイソウは、韓国産、または中国産、ロシア産、日本産のような輸入産の、根、茎または花である請求項1に記載の薬学組成物。
- 前記クガイソウの抽出物は、粗抽出物、極性溶媒可溶抽出物または非極性溶媒可溶抽出物である請求項1または2に記載の薬学組成物。
- 前記粗抽出物は、精製水を含む水、メタノール・エタノール・ブタノールのようなC1-C4低級アルコール、またはそれらの混合溶媒から選択された溶媒に可溶された抽出物である請求項3に記載の薬学組成物。
- 前記非極性溶媒可溶抽出物は、ヘキサン、塩化メチレン、クロロホルムまたは酢酸エチルに可溶された抽出物である請求項3または4に記載の薬学組成物。
- 前記炎症は、皮膚炎、アトピー、結膜炎、歯周炎、鼻炎、中耳炎、咽喉炎、扁桃炎、肺炎、胃潰瘍、胃炎、クローン病、大腸カタル、痔疾、痛風、強直性脊椎炎、リウマチ熱、ループス、線維筋痛(fibromyalgia)、乾癬関節炎、骨関節炎、リウマチ関節炎、肩関節周囲炎、腱炎、腱鞘炎、腱周囲炎、筋肉炎、肝炎、膀胱炎、腎臓炎、シェーグレン症侯群(Sjogren's syndrome)、多発性硬化症、並びに急性及び慢性の炎症疾患からなる群のうちから選択された疾患である請求項1~5のいずれか一項に記載の薬学組成物。
- 前記アレルギーは、過敏症、アレルギー性鼻炎、喘息、アレルギー性結膜炎、アレルギー性皮膚炎、アトピー性皮膚炎、接触性皮膚炎、蕁麻疹、昆虫アレルギー、食品アレルギーまたは薬品アレルギーからなる群のうちから選択された疾患である請求項1~6のいずれか一項に記載の薬学組成物。
- 前記喘息は、室内塵、ダニ、花粉、動物毛、頭垢、ごきぶり、食品、薬物、風邪、タバコの煙、室内汚染、大気汚染、食品添加剤、身体的活動、気候変化、黄砂、またはストレスからなる群のうちから選択された要因に起因した気管支喘息である請求項1~7のいずれか一項に記載の薬学組成物。
- クガイソウ(Veronicastrum sibiricum L. Pennell,クガイソウ属(Veronicastrum))抽出物を有効成分として含む、炎症、アレルギー及び喘息の予防または改善するために用いられる健康機能食品。
- クガイソウ(Veronicastrum sibiricum L. Pennell,クガイソウ属(Veronicastrum))抽出物を有効成分として含む、炎症、アレルギー及び喘息を予防または改善するために用いられる健康機能食品。
- 散剤、顆粒剤、錠剤、カプセル剤、丸剤、懸濁液、エマルジョン、シロップ剤、ティーバッグ剤、浸出茶または健康飲料の形態である、請求項10記載の健康機能食品。
- クガイソウ(Veronicastrum sibiricum L. Pennell,クガイソウ属(Veronicastrum))抽出物を有効成分として含む、炎症、アレルギー及び喘息を予防または改善するために用いられる健康補助食品。
- クガイソウ(Veronicastrum sibiricum L. Pennell,クガイソウ属(Veronicastrum))抽出物を有効成分として含む炎症、アレルギー及び喘息を予防または改善するために用いられる食品。
- クガイソウ(Veronicastrum sibiricum L. Pennell,クガイソウ属(Veronicastrum))抽出物を有効成分として含む炎症、アレルギー及び喘息を予防または改善するために用いられる食品添加物。
- クガイソウ(Veronicastrum sibiricum L. Pennell,クガイソウ属(Veronicastrum))抽出物を、炎症患者、アレルギー患者及び喘息疾患患者に投与することを含む、炎症患者、アレルギー患者及び喘息疾患患者を治療するための治療方法。
- 炎症治療用、アレルギー治療用及び喘息疾患治療用の薬剤を製造するためのクガイソウ(Veronicastrum sibiricum L. Pennell,クガイソウ属(Veronicastrum))抽出物の用途。
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