JP5301192B2 - イグチ科ヌメリイグチ属担子菌培地培養物とその利用法 - Google Patents
イグチ科ヌメリイグチ属担子菌培地培養物とその利用法 Download PDFInfo
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- JP5301192B2 JP5301192B2 JP2008093863A JP2008093863A JP5301192B2 JP 5301192 B2 JP5301192 B2 JP 5301192B2 JP 2008093863 A JP2008093863 A JP 2008093863A JP 2008093863 A JP2008093863 A JP 2008093863A JP 5301192 B2 JP5301192 B2 JP 5301192B2
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- mycelium
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- melanin production
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- ZOKXTWBITQBERF-UHFFFAOYSA-N Molybdenum Chemical compound [Mo] ZOKXTWBITQBERF-UHFFFAOYSA-N 0.000 description 1
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- 150000007513 acids Chemical class 0.000 description 1
- 239000004480 active ingredient Substances 0.000 description 1
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- 125000002015 acyclic group Chemical group 0.000 description 1
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- 125000005907 alkyl ester group Chemical group 0.000 description 1
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- ARUVKPQLZAKDPS-UHFFFAOYSA-L copper(II) sulfate Chemical compound [Cu+2].[O-][S+2]([O-])([O-])[O-] ARUVKPQLZAKDPS-UHFFFAOYSA-L 0.000 description 1
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- MNNHAPBLZZVQHP-UHFFFAOYSA-N diammonium hydrogen phosphate Chemical compound [NH4+].[NH4+].OP([O-])([O-])=O MNNHAPBLZZVQHP-UHFFFAOYSA-N 0.000 description 1
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- GVVPGTZRZFNKDS-JXMROGBWSA-N geranyl diphosphate Chemical compound CC(C)=CCC\C(C)=C\CO[P@](O)(=O)OP(O)(O)=O GVVPGTZRZFNKDS-JXMROGBWSA-N 0.000 description 1
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- CDAISMWEOUEBRE-GPIVLXJGSA-N inositol Chemical compound O[C@H]1[C@H](O)[C@@H](O)[C@H](O)[C@H](O)[C@@H]1O CDAISMWEOUEBRE-GPIVLXJGSA-N 0.000 description 1
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- PNDPGZBMCMUPRI-UHFFFAOYSA-N iodine Chemical compound II PNDPGZBMCMUPRI-UHFFFAOYSA-N 0.000 description 1
- 229910052742 iron Inorganic materials 0.000 description 1
- BAUYGSIQEAFULO-UHFFFAOYSA-L iron(2+) sulfate (anhydrous) Chemical compound [Fe+2].[O-]S([O-])(=O)=O BAUYGSIQEAFULO-UHFFFAOYSA-L 0.000 description 1
- RUTXIHLAWFEWGM-UHFFFAOYSA-H iron(3+) sulfate Chemical compound [Fe+3].[Fe+3].[O-]S([O-])(=O)=O.[O-]S([O-])(=O)=O.[O-]S([O-])(=O)=O RUTXIHLAWFEWGM-UHFFFAOYSA-H 0.000 description 1
- 229910000359 iron(II) sulfate Inorganic materials 0.000 description 1
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- 230000007794 irritation Effects 0.000 description 1
- 229940119170 jojoba wax Drugs 0.000 description 1
- 239000008101 lactose Substances 0.000 description 1
- 150000002632 lipids Chemical class 0.000 description 1
- 239000011777 magnesium Substances 0.000 description 1
- 229910052749 magnesium Inorganic materials 0.000 description 1
- WRUGWIBCXHJTDG-UHFFFAOYSA-L magnesium sulfate heptahydrate Chemical compound O.O.O.O.O.O.O.[Mg+2].[O-]S([O-])(=O)=O WRUGWIBCXHJTDG-UHFFFAOYSA-L 0.000 description 1
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- 229940099596 manganese sulfate Drugs 0.000 description 1
- 239000011702 manganese sulphate Substances 0.000 description 1
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- WPBNNNQJVZRUHP-UHFFFAOYSA-L manganese(2+);methyl n-[[2-(methoxycarbonylcarbamothioylamino)phenyl]carbamothioyl]carbamate;n-[2-(sulfidocarbothioylamino)ethyl]carbamodithioate Chemical compound [Mn+2].[S-]C(=S)NCCNC([S-])=S.COC(=O)NC(=S)NC1=CC=CC=C1NC(=S)NC(=O)OC WPBNNNQJVZRUHP-UHFFFAOYSA-L 0.000 description 1
- SQQMAOCOWKFBNP-UHFFFAOYSA-L manganese(II) sulfate Chemical compound [Mn+2].[O-]S([O-])(=O)=O SQQMAOCOWKFBNP-UHFFFAOYSA-L 0.000 description 1
- 239000012046 mixed solvent Substances 0.000 description 1
- 229910052750 molybdenum Inorganic materials 0.000 description 1
- 239000011733 molybdenum Substances 0.000 description 1
- 239000001788 mono and diglycerides of fatty acids Substances 0.000 description 1
- 150000002772 monosaccharides Chemical class 0.000 description 1
- GOQYKNQRPGWPLP-UHFFFAOYSA-N n-heptadecyl alcohol Natural products CCCCCCCCCCCCCCCCCO GOQYKNQRPGWPLP-UHFFFAOYSA-N 0.000 description 1
- 229960003512 nicotinic acid Drugs 0.000 description 1
- 235000001968 nicotinic acid Nutrition 0.000 description 1
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- QJGQUHMNIGDVPM-UHFFFAOYSA-N nitrogen group Chemical group [N] QJGQUHMNIGDVPM-UHFFFAOYSA-N 0.000 description 1
- 235000015097 nutrients Nutrition 0.000 description 1
- CKQVRZJOMJRTOY-UHFFFAOYSA-N octadecanoic acid;propane-1,2,3-triol Chemical compound OCC(O)CO.CCCCCCCCCCCCCCCCCC(O)=O CKQVRZJOMJRTOY-UHFFFAOYSA-N 0.000 description 1
- 239000002674 ointment Substances 0.000 description 1
- 150000002894 organic compounds Chemical class 0.000 description 1
- 239000005416 organic matter Substances 0.000 description 1
- 239000003960 organic solvent Substances 0.000 description 1
- 239000003002 pH adjusting agent Substances 0.000 description 1
- 229940055726 pantothenic acid Drugs 0.000 description 1
- 235000019161 pantothenic acid Nutrition 0.000 description 1
- 239000011713 pantothenic acid Substances 0.000 description 1
- RUVINXPYWBROJD-UHFFFAOYSA-N para-methoxyphenyl Natural products COC1=CC=C(C=CC)C=C1 RUVINXPYWBROJD-UHFFFAOYSA-N 0.000 description 1
- 235000019319 peptone Nutrition 0.000 description 1
- 239000002304 perfume Substances 0.000 description 1
- 235000019271 petrolatum Nutrition 0.000 description 1
- 229940066842 petrolatum Drugs 0.000 description 1
- 239000000546 pharmaceutical excipient Substances 0.000 description 1
- 239000011574 phosphorus Substances 0.000 description 1
- 229910052698 phosphorus Inorganic materials 0.000 description 1
- 108010071062 pinene cyclase I Proteins 0.000 description 1
- 239000000419 plant extract Substances 0.000 description 1
- 229920000515 polycarbonate Polymers 0.000 description 1
- 239000004417 polycarbonate Substances 0.000 description 1
- 239000000256 polyoxyethylene sorbitan monolaurate Substances 0.000 description 1
- 235000010486 polyoxyethylene sorbitan monolaurate Nutrition 0.000 description 1
- 229920001282 polysaccharide Polymers 0.000 description 1
- 239000005017 polysaccharide Substances 0.000 description 1
- 150000003097 polyterpenes Chemical class 0.000 description 1
- 239000011591 potassium Substances 0.000 description 1
- 229910052700 potassium Inorganic materials 0.000 description 1
- 239000001103 potassium chloride Substances 0.000 description 1
- 235000011164 potassium chloride Nutrition 0.000 description 1
- 239000004323 potassium nitrate Substances 0.000 description 1
- 235000010333 potassium nitrate Nutrition 0.000 description 1
- 239000001965 potato dextrose agar Substances 0.000 description 1
- 230000002335 preservative effect Effects 0.000 description 1
- 235000013772 propylene glycol Nutrition 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- RADKZDMFGJYCBB-UHFFFAOYSA-N pyridoxal hydrochloride Natural products CC1=NC=C(CO)C(C=O)=C1O RADKZDMFGJYCBB-UHFFFAOYSA-N 0.000 description 1
- 235000008160 pyridoxine Nutrition 0.000 description 1
- 239000011677 pyridoxine Substances 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- CDAISMWEOUEBRE-UHFFFAOYSA-N scyllo-inosotol Natural products OC1C(O)C(O)C(O)C(O)C1O CDAISMWEOUEBRE-UHFFFAOYSA-N 0.000 description 1
- 229930000044 secondary metabolite Natural products 0.000 description 1
- 229930004725 sesquiterpene Natural products 0.000 description 1
- 150000004354 sesquiterpene derivatives Chemical class 0.000 description 1
- 229930002368 sesterterpene Natural products 0.000 description 1
- 150000002653 sesterterpene derivatives Chemical class 0.000 description 1
- 239000000377 silicon dioxide Substances 0.000 description 1
- 239000011734 sodium Substances 0.000 description 1
- 229910052708 sodium Inorganic materials 0.000 description 1
- 239000011684 sodium molybdate Substances 0.000 description 1
- 235000015393 sodium molybdate Nutrition 0.000 description 1
- TVXXNOYZHKPKGW-UHFFFAOYSA-N sodium molybdate (anhydrous) Chemical compound [Na+].[Na+].[O-][Mo]([O-])(=O)=O TVXXNOYZHKPKGW-UHFFFAOYSA-N 0.000 description 1
- 229910052938 sodium sulfate Inorganic materials 0.000 description 1
- 235000011152 sodium sulphate Nutrition 0.000 description 1
- 229960005078 sorbitan sesquioleate Drugs 0.000 description 1
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- 239000005720 sucrose Substances 0.000 description 1
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- 239000011593 sulfur Substances 0.000 description 1
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- 239000004094 surface-active agent Substances 0.000 description 1
- 208000024891 symptom Diseases 0.000 description 1
- 229940116411 terpineol Drugs 0.000 description 1
- 150000003535 tetraterpenes Chemical class 0.000 description 1
- 235000009657 tetraterpenes Nutrition 0.000 description 1
- 235000019157 thiamine Nutrition 0.000 description 1
- 239000011721 thiamine Substances 0.000 description 1
- KYMBYSLLVAOCFI-UHFFFAOYSA-N thiamine Chemical compound CC1=C(CCO)SCN1CC1=CN=C(C)N=C1N KYMBYSLLVAOCFI-UHFFFAOYSA-N 0.000 description 1
- 229960000790 thymol Drugs 0.000 description 1
- 150000003648 triterpenes Chemical class 0.000 description 1
- 229930195735 unsaturated hydrocarbon Natural products 0.000 description 1
- 235000010374 vitamin B1 Nutrition 0.000 description 1
- 239000011691 vitamin B1 Substances 0.000 description 1
- 239000011726 vitamin B6 Substances 0.000 description 1
- 235000019158 vitamin B6 Nutrition 0.000 description 1
- 239000012138 yeast extract Substances 0.000 description 1
- 239000011701 zinc Substances 0.000 description 1
- 229910052725 zinc Inorganic materials 0.000 description 1
- NWONKYPBYAMBJT-UHFFFAOYSA-L zinc sulfate Chemical compound [Zn+2].[O-]S([O-])(=O)=O NWONKYPBYAMBJT-UHFFFAOYSA-L 0.000 description 1
- 229910000368 zinc sulfate Inorganic materials 0.000 description 1
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Landscapes
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Cosmetics (AREA)
- Medicines Containing Plant Substances (AREA)
Description
bovinus)、チチアワタケ(Suillus granulatus)、シロヌメリイグチ(Suillus laricinus)、ヌメリツバイグチ(Suillus luteus)等が属する。しかし、その供給源としての天然の子実体は入手の時期が限定され、また入手した後も、子実体の個体差による有用性の変動や量産化等実用面での問題点が多い。そこで近年、イグチ科担子菌を含む外生菌根菌の菌糸体を人工的に培養する方法が提案されている(例えば、特許文献1)。
、更に好ましくは0.5〜10重量% である。0.5〜10重量% であれば、より高い美白効果が期待できる。配合量が0.1重量%未満の場合には十分な効果を発揮させることができない為に好ましくなく、20重量%より多い場合には高圧滅菌時または培養時の操作性が低下する為に好ましくない。
、更に好ましくは0.001〜0.01重量% である。0.001〜0.01重量%であれば、より高い美白効果が期待できる。配合量が0.0001重量%未満の場合には十分な効果を発揮させることができない為に好ましくなく、0.02重量%より多い場合には効果が平衡に達する。
g 、モルトエキス0.3g、塩化カルシウム2水和物0.0066g、塩化ナトリウム0.0025g、リン酸2水素カリウム0.05g、リン酸水素2アンモニウム0.025g、硫酸マグネシウム7水和物0.015g、塩化鉄(III)1%溶液0.12mL、チアミン塩酸塩0.01mg を精製水100mLの水に溶解させることで調製可能である。改変浜田培地は、例えばグルコース2.0g、乾燥酵母0.5gを精製水100mLに溶解させることで調製可能である。基本培地には炭素源、窒素源などイグチ科担子菌菌糸体を培養する際の生育に必要な栄養素が含まれており、さらにビタミン類やミネラルを添加することも可能である。
価アルコール;グリセリン、プロピレングリコール、1、3−ブチレングリコール等の液状多価アルコール;アセトン、メチルエチルケトン等のケトン;酢酸エチルなどのアルキルエステル;ベンゼン、ヘキサン等の炭化水素;ジエチルエーテル等のエーテル類;ジクロルメタン、クロロホルム等のハロゲン化アルカン等の1
種または2 種以上を用いることが出来る。そのうち、水、エチルアルコール、1、3−ブチレングリコールの1種または2種以上の混合溶媒が特に好適に示される。
皮膚の黒化には色素のメラニンが深く関与していると考えられる。即ち、メラニンが紫外線などの外的刺激を受けて肌の皮膚組織で生産され、その結果、肌の黒化が促進され、シミ・ソバカス・色黒等の症状が引き起こされると考えられている。よって、肌の美白作用の確認法としメラニン産生抑制効果に着目し、マウスメラノーマ細胞によるメラニン産生抑制試験を行った。以下、ハナイグチ培養菌糸抽出 物の美白効果に関するメラニン産生抑制試験について説明する。
本発明に係る液体培養培地を表1に示した組成にて作製した。グルコース10.0 g 、モルトエキス3.0g、塩化カルシウム2水和物0.066g、塩化ナトリウム0.025g、リン酸2水素カリウム0.5g、リン酸水素2アンモニウム0.25g、硫酸マグネシウム7水和物0.15g、塩化鉄(III)1%溶液1.2mL、チアミン塩酸塩0.1mg を1000mLの水に添加して基本の液体培地を作った。また、添加物として豆乳及び/またはモノテルペンを表1の量を添加して液体培地を調製後、オートクレーブを用いて高圧殺菌した。
Broth(Difco社製)2.4%で振とう培養(24℃、150rpm、2週間)したものを用いた。得られた菌糸をホモジネートしたものを5mLずつ培養瓶に接種した。接種後は24℃の条件下で通気しながら培養を行った。
ハナイグチ培養菌糸のエタノール抽出物は、ハナイグチ培養菌糸をエタノールで、60℃にて24時間抽出した。溶媒を留去し、1%濃度になるようにエタノールに溶解した。ハナイグチ培養菌糸の水抽出物は、ハナイグチ培養菌糸を精製水で、60℃にて24時間抽出した。溶媒を留去し、1%濃度になるように精製水に溶解した。ハナイグチ培養液は、培養した培養物をADVANTEC 5C定量ろ過を用いて吸引ろ過し調製した。陽性対照として用いたアルブチンは、市販の試薬を用いてPBS(−)溶液に溶解した。試料は全て、0.2μmメンブレンフィルターで除菌ろ過を行った。
培養液は、牛胎児血清5.0%を加えたダルベッコMEM(D−MEM)培地を用いた。細胞は、マウスメラノーマB−16 F−10を12ウェルプレートに植え付けた。植え付け量は5×104/wellとした。植え付けの翌日、抽出物試料を乾燥物濃度にして、エタノール抽出物は25ppm、水抽出物は200ppmの濃度になるよう添加し、添加後3日後に試験を終了した。
12ウェルプレートに増殖した細胞をPBS(−)溶液で洗浄後、2N−NaOH溶液に溶解し、405nmの吸光度を測定した。それぞれ吸光度をコントロールと比較することにより、8段階表示をした。
表2に結果の判定基準を示す。メラニン量はコントロールと比較した吸光度値の割合で示した。なおアルブチンは、メラニン産生抑制の陽性対照物質として示している。
bovinus)、チチアワタケ(Suillus granulatus)、シロヌメリイグチ(Suillus laricinus)、ヌメリツバイグチ(Suillus luteus)の培養においても、豆乳及び/またはモノテルペンの添加によって、菌糸水抽出液、菌糸エタノール抽出液、培養液のメラニン産生抑制効果が向上した。
化粧料の処方例
a) ミツロウ・・・2.0
b) ステアリルアルコール・・・5.0
c) ステアリン酸・・・8.0
d) スクワラン・・・10.0
e) 自己乳化型グリセリルモノステアレート・・・3.0
f) ポリオキシエチレンセチルエーテル(20E.O.)・・・1.0
g) ハナイグチ菌糸体水抽出物(ゲラニオール0.005%配合培養)・・・2.0
h) 1,3−ブチレングリコール・・・5.0
i) 水酸化カリウム・・・0.3
j) 防腐剤・酸化防止剤・・・適量
k) 精製水・・・残部
製法
a)〜f)までを加熱溶解し、80℃に保つ。h)
〜 k)までを加熱溶解し、80℃に保ち、a) 〜f)に加えて乳化し、40℃まで撹拌しながら冷却する。その後、g)を加え、攪拌し均一に溶解する。
a) ミツロウ・・・0.5
b) ワセリン・・・2.0
c) スクワラン・・・8.0
d) ソルビタンセスキオレエート・・・0.8
e) ポリオキシエチレンオレイルエーテル(20E.O.)・・・1.2
f) ハナイグチ菌糸体エタノール抽出物(ゲラニオール0.005%配合培養)・・・1.0
g) 1,3− ブチレングリコール・・・7.0
h) カルボキシビニルポリマー・・・0.2
i) 水酸化カリウム・・・0.1
j) 精製水・・・残部
k) 防腐剤・酸化防止剤・・・適量
l) エタノール・・・7.0
製法
a)〜e)までを加熱溶解し、80℃に保つ。g)〜k)までを加熱溶解し、80℃に保ち、a)〜e)に加えて乳化し、50℃
まで撹拌しながら冷却する。50℃ でf)、l)を添加し、40℃まで攪拌、冷却する。
a) ハナイグチ菌糸体エタノール抽出物(豆乳5%配合培養)・・・1.0
b) グリセリン・・・5.0
c) ポリオキシエチレンソルビタンモノラウレート(20E.O.)・・・1.0
d) エタノール・・・6.0
e) 香料・・・適量
f) 防腐剤・酸化防止剤・・・適量
g) 精製水・・・残部
製法
a)〜g)までを混合し、均一に溶解する。
a) ステアリン酸・・・12 .0
b) ミリスチン酸・・・14 .0
c) ラウリン酸・・・5.0
d) ホホバ油・・・3.0
e) グリセリン・・・10.0
f) ソルビトール・・・15.0
g) 1,3−ブチレングリコール・・・10.0
h) POE(20)グリセロールモノステアリン酸・・・2.0
i) 水酸化カリウム・・・5.0
j) 水・・・残部
k) キレート剤・・・適量
l) 香料・・・適量
m) ハナイグチ菌糸体水抽出物(豆乳5%配合培養)・・・2.0
製法
a)〜h)までを加熱溶解し70℃に保つ。j)にi)を溶解後a)〜h)に加えケン化する。その後k)、l)を入れ攪拌しながら冷却する。50℃でm)を添加し、40℃まで攪拌、冷却する。
(1)塗布によるヒトでの効果確認試験
被験者として、20〜50歳の女性15名に1日2回(朝、夜)連続2ヵ月間、本発明品と比較品のそれぞれを使用させ、塗布部位の状態を試験前後で比較し、改善効果を調べた。本試験には、実施例16
として処方例中の(1)化粧用クリームで示した化粧料を用い、比較例5には実施例16からハナイグチ菌糸体水抽出物(ゲラニオール0.005%配合培養)を除き、代わりに添加物無配合で培養したハナイグチ菌糸体水抽出物を配合した化粧料、比較例6には実施例1からハナイグチ菌糸体水抽出物を除いた化粧料を作成し、その使用による効果について調べた。本発明の有効成分を配合した化粧料を毎日使用しながら肌の美白効果及び使用後の肌の感触について塗布開始前及び2ヶ月塗布後におけるアンケートで集計し、効果の確認を行った。
著効:しみ・ソバカスが消えた部分がある。
有効:しみ・ソバカスが目立たなくなった。
やや有効:しみ・ソバカスがやや目立たなくなった。
無効:変化が認められなかった。
尚、本試験において実施例16を使用して何らかの刺激があったと答えた人はいなかった。
Claims (3)
- 豆乳を培地全量の0.5〜10重量%及び/又は非環状のモノテルペンを0.001〜0.01重量%配合した培地で培養したイグチ科ヌメリイグチ属(Suillus)担子菌培養物。
- イグチ科ヌメリイグチ属(Suillus)担子菌が、ハナイグチ(Suillus grevillei)、ヌメリイグチ(Suillus luteus)、アミタケ(Suillus bovinus)、チチアワタケ(Suillus granulatus)、シロヌメリイグチ(Suillus laricinus)、ヌメリツバイグチ(Suillus luteus)であることを特徴とする請求項1記載の培養物。
- 請求項1乃至請求項2記載のイグチ科ヌメリイグチ属(Suillus)担子菌培養物を配合することを特徴とする皮膚外用剤。
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