JP2020185012A - 藻類細胞培養液およびバイオマス、脂質化合物および組成物、ならびに関連製品の製造方法 - Google Patents
藻類細胞培養液およびバイオマス、脂質化合物および組成物、ならびに関連製品の製造方法 Download PDFInfo
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Abstract
Description
第一級アミノ基の窒素(L−アルギニンおよびグルタミン酸を含む)、尿素およびアンモニアなどの生物学的に利用可能な総窒素量は、K−PANOPA、K−LargeおよびK−Glut生化学キット(Megazyme、アイルランド)を使用して測定した。有機認証済のビートジュースの発酵物、トマトジュース、有機認証済のにんじんジュース、有機認証済のりんごジュース、およびココナッツォーターの5種のジュースを試験した。各ジュースは、低温殺菌し、10%濃度(vol/vol)に希釈してから試験した。測定値は100%ジュースの値に換算した。
オーランチオキトリウム・リマシナムを、pHを調節していないジュース培地中で増殖させた。(藻類の播種および培養の両方を使用目的とした)合計15個の培地を、以下の各ジュース濃度となるように調製した:(1)10%濃度の有機認証済のビートジュース、(2)25%濃度の有機認証済のビートジュース、(3)50%濃度の有機認証済のビートジュース、(4)10%濃度の有機認証済のりんごジュース、(5)25%濃度の有機認証済のりんごジュース、(6)50%濃度の有機認証済のりんごジュース、(7)10%濃度のトマトジュース、(8)25%濃度のトマトジュース、(9)50%濃度のトマトジュース、(10)10%濃度のココナッツォーター、(11)25%濃度のココナッツォーター、(12)50%濃度のココナッツォーター、(13)10%濃度の有機認証済のにんじんジュース、(14)25%濃度の有機認証済のにんじんジュース、および(15)50%濃度の有機認証済のにんじんジュース。各培地は、塩分濃度が12.7pptとなるように海水をさらに含有していた。培地を低温殺菌法によって滅菌した。
オーランチオキトリウム・リマシナムを、pHを6.8に調節したジュース培地で増殖させた。(藻類の播種および培養の両方を使用目的とした)合計15個の培地を、以下の各ジュース濃度となるように調製した:(1)10%濃度の有機認証済のビートジュース、(2)25%濃度の有機認証済のビートジュース、(3)50%濃度の有機認証済のビートジュース、(4)10%濃度の有機認証済のりんごジュース、(5)25%濃度の有機認証済のりんごジュース、(6)50%濃度の有機認証済のりんごジュース、(7)10%濃度のトマトジュース、(8)25%濃度のトマトジュース、(9)50%濃度のトマトジュース、(10)10%濃度のココナッツォーター、(11)25%濃度のココナッツォーター、(12)50%濃度のココナッツォーター、(13)10%濃度の有機認証済のにんじんジュース、(14)25%濃度の有機認証済のにんじんジュース、および(15)50%濃度の有機認証済のにんじんジュース。各培地は、塩分濃度が12.7pptとなるのに十分な海水をさらに含有していた。培地を低温殺菌法によって滅菌した。
オーランチオキトリウム・リマシナムを、ブドウ糖を添加したジュース培地で増殖させた。(藻類の播種および培養の両方を使用目的とした)合計5種の培地を、以下の各ジュース濃度となるように調製した:(1)25%濃度の有機認証済のビートジュース、(2)50%濃度のトマトジュース、(3)50%濃度の有機認証済のにんじんジュース、(4)50%濃度の有機認証済のりんごジュース、および(5)50%濃度のココナッツォーター。各培地は、6%濃度になるようにブドウ糖を添加した。培地を低温殺菌法によって滅菌した。
オーランチオキトリウム・リマシナムを、さまざまなジュース培地中で増殖させた。(藻類の播種および培養の両方を使用目的とした)合計15種の培地を、以下の各ジュース濃度となるように調製した:(1)10%濃度の有機認証済のビートジュース、(2)25%濃度の有機認証済のビートジュース、(3)50%濃度の有機認証済のビートジュース、(4)10%濃度の有機認証済のりんごジュース、(5)25%濃度の有機認証済のりんごジュース、(6)50%濃度の有機認証済のりんごジュース、(7)10%濃度のトマトジュース、(8)25%濃度のトマトジュース、(9)50%濃度のトマトジュース、(10)10%濃度のココナッツォーター、(11)25%濃度のココナッツォーター、(12)50%濃度のココナッツォーター、(13)10%濃度の有機認証済のにんじんジュース、(14)25%濃度の有機認証済のにんじんジュース、および(15)50%濃度の有機認証済のにんじんジュース。各培地は、塩分濃度が12.7pptとなるように海水をさらに含有し、濃度6%となるように有機認証済のブドウ糖が添加されている。培地を低温殺菌法によって滅菌した。
オーランチオキトリウム・リマシナムを、さまざまなジュース培地中で増殖させた。(藻類の播種および培養の両方を使用目的とした)合計15種の培地を、以下の各ジュース濃度となるように調製した:(1)10%濃度の有機認証済のビートジュース、(2)25%濃度の有機認証済のビートジュース、(3)50%濃度の有機認証済のビートジュース、(4)10%濃度の有機認証済のりんごジュース、(5)25%濃度の有機認証済のりんごジュース、(6)50%濃度の有機認証済のりんごジュース、(7)10%濃度のトマトジュース、(8)25%濃度のトマトジュース、(9)50%濃度のトマトジュース、(10)10%濃度のココナッツォーター、(11)25%濃度のココナッツォーター、(12)50%濃度のココナッツォーター、(13)10%濃度の有機認証済のにんじんジュース、(14)25%濃度の有機認証済のにんじんジュース、および(15)50%濃度の有機認証済のにんじんジュース。各培地は、塩分濃度が12.7pptとなるように海水をさらに含有し、濃度6%となるように有機認証済のブドウ糖が添加されている。培地を低温殺菌法によって滅菌した。
オーランチオキトリウム・リマシナムを、さまざまなジュース培地中で増殖させた。(藻類の播種および培養の両方を使用目的とした)合計15種の培地を、以下の各ジュース濃度となるように調製した:(1)10%濃度の有機認証済のビートジュース、(2)25%濃度の有機認証済のビートジュース、(3)50%濃度の有機認証済のビートジュース、(4)10%濃度の有機認証済のりんごジュース、(5)25%濃度の有機認証済のりんごジュース、(6)50%濃度の有機認証済のりんごジュース、(7)10%濃度のトマトジュース、(8)25%濃度のトマトジュース、(9)50%濃度のトマトジュース、(10)10%濃度のココナッツォーター、(11)25%濃度のココナッツォーター、(12)50%濃度のココナッツォーター、(13)10%濃度の有機認証済のにんじんジュース、(14)25%濃度の有機認証済のにんじんジュース、および(15)50%濃度の有機認証済のにんじんジュース。各培地は、塩分濃度が12.7pptとなるように海水をさらに含有し、濃度6%となるように有機認証済のブドウ糖を添加されている。培地を低温殺菌法によって滅菌した。
ナンノクロロプシス・オクラタを、ビートジュース培地で増殖させた。1×106個のナンノクロロプシス・オクラタ細胞を、以下の各培地に播種し、培養した:(1)F/2培地(市販の標準合成培地)、(2)(pHを7.6に調節し、海水で塩分濃度を25pptに調節した)自然発酵ビートジュース、(3)(pHを7.6に調節し、海水で塩分濃度を25pptに調節した)30mMブドウ糖を添加した自然発酵ビートジュース、および(4)(pHを7.6に調節し、海水で塩分濃度を25pptに調節した)30mMエタノールを添加した自然発酵ビートジュース。ビートジュース培地を低温殺菌法によって滅菌した。細胞を22℃にて160rpmで振盪させながら5日間インキュベートした。
スキゾキトリウム・リマシナムを、ビート糖蜜培地で増殖させた。1×106個のスキゾキトリウム・リマシナム細胞を、以下の各培地中に播種し、培養した:(1)6%ブドウ糖を添加した発酵ビートジュース(「BJ」)、(2)6%ブドウ糖を添加した、有機認証済の8.4%ビート糖蜜(「BM」)、および(3)酵素インベルターゼで加水分解した、有機認証済の8.4%ビート糖蜜(「BM−H」)。培地を低温殺菌法または濾過法によって滅菌した。窒素含有量を等しくするために、ビートジュースおよびビート糖蜜の処方を調整した。細胞を22℃にて200rpmで振盪させながら5日間インキュベートした。
10%ビート糖蜜、20%ビート糖蜜および100%ビートジュースそれぞれを含有する溶液(低温殺菌(95℃)、pH3.7)を、食品等級有機酸を使用してpH4.5に調節し、パン酵母(Sigma−Chemical Co.)由来のインベルターゼで無菌条件下にて処理した。得られた溶液をK−SUFRGキット(Megazyme、アイルランド)を用いて分析し、酵素処理の前と後における各処方中のショ糖、果糖およびグルコースの濃度を決定した。サンプルに標準グルコース、標準果糖および標準ショ糖を添加して濃度を上げた。溶液を55℃で24時間インキュベートした。結果を表10に示す。
スキゾキトリウム・リマシナムを、ビート糖蜜培地で増殖させた。1×106個のスキゾキトリウム・リマシナム細胞を、以下の各培地中に播種し、培養した:(1)市販の基礎培地(3%ブドウ糖、1%酵母エキスおよび海水)、(2)6%ブドウ糖を添加した発酵ビートジュース、(3)6%ブドウ糖を添加した8.4%ビート糖蜜、および(4)酵素インベルターゼで加水分解した8.4%ビート糖蜜。培地を低温殺菌法または濾過法によって滅菌した。窒素含有量を等しくするために、ビートジュースおよびビート糖蜜の処方を調整した。細胞を22℃にて200rpmで振盪させながら5日間インキュベートした。各培地で120時間増殖させた後に、同等数の細胞(2×108個の細胞)を含有する培養を一定量回収した。培養サンプルを凍結乾燥し、標準方法を用いたガスクロマトグラフィーにより、DHA含有量を求めた。
第一級アミノ基の窒素(L−アルギニンおよびグルタミン酸を含む)、尿素およびアンモニアなどの生物学的に利用可能な窒素の総量を、K−PANOPA、K−LargeおよびK−Glut生化学キット(Megazyme、アイルランド)を使用して測定した。ビート糖蜜を試験した。サンプルを10%濃度(vol/vol)に希釈してから試験した。測定値は100%ジュースの値に換算した。結果を表15にまとめる。
スキゾキトリウム・リマシナムを、トマトジュースベース培地に播種した。等容積の培養液を遠心分離にかけ、培地をデカントした。得られた藻類ペレットを、それぞれ個別にビートジュース、にんじんジュース、ブルーベリージュースまたは水中に9:1の比率で懸濁させた。各混合物のサンプル約0.8gを試験して開始時のDHA濃度を求めた。新たにサンプル約0.8gを等量に分配し、それぞれを1.5mL管に密封し、次いで最長6日間、60℃で加熱した。FAMEのガスクロマトグラフィーによってDHAレベルを定量した。結果を表16にまとめる。
スキゾキトリウム・リマシナムを、基礎培地に播種した。等容積の培養液を遠心分離にかけ、培地をデカントした。得られた藻類ペレットを、1:1の比率で水中に均一化し、以下の混合物中にそれぞれ1:1の比率で懸濁させた:(i)にんじんジュース、(ii)ビートジュース、(iii)ビートジュース+緑茶エキス、(iv)ブルーベリージュース、(v)水、(vi)水+6%ショ糖(遮光)、または(vii)水+6%ショ糖+緑茶エキス。緑茶エキスは、酸化防止性ポリフェノールフラボノイドであるエピガロカテキン没食子酸塩(EGCG)源として添加した。緑茶エキスを添加した水およびビートジュースは、1000ppmのEGCGを含有していた。約0.1グラムのサンプルを等量に分配し、1.5mL管に密封して、9日間にわたり、3日毎にDHA含有量を求めた。サンプルは60℃に維持した。新たに約1グラムのサンプルを等量に分配し、1.5mL管に密封して、12日間にわたり、3日毎に混合物の酸化防止活性を求めた。サンプルを60℃に維持した。FAMEのガスクロマトグラフィーによってDHAレベルを定量化し、一方で混合物の酸化防止剤含有量を、ラジカル捕捉活性をパーセンテージで表すDPPH(Sigma−Aldrich)分析法によって決定した。結果を図10(a)〜(b)および図11(a)〜(b)にまとめる。
スキゾキトリウム・リマシナムを、基礎培地に播種した。等容積の培養液を遠心分離にかけ、培地をデカントした。得られた藻類ペレットを、1:1の比率で水中に均一化し、以下の混合物中にそれぞれ1:1の比率で懸濁させた:(i)にんじんジュース、(ii)ビートジュース、(iii)ブルーベリージュース、(iv)水。約1グラムのサンプルを、ラジカル捕捉活性をパーセンテージで表すDPPH(Sigma−Aldrich)分析法を用いて酸化防止活性について試験した。これらの結果を表17にまとめる。
Claims (89)
- 培養培地中で藻類を増殖させ、前記培地から藻類バイオマスまたは藻類細胞培養液を回収することを含む、藻類バイオマスまたは藻類細胞培養物の製造方法であって、
前記培地が、1種または複数種の果実もしくは野菜または果実と野菜の任意の組合せより得られたジュース、酸素源、および窒素を含み、
前記培地が、滅菌されており、
前記窒素が、天然由来の窒素からなる、方法。 - 前記培地に食塩源が添加されている、請求項1に記載の方法。
- 前記食塩源が海水である、請求項2に記載の方法。
- 前記海水の塩分濃度が約10ppt〜約35pptの範囲である、請求項3に記載の方法。
- 前記海水の塩分濃度が約12.7pptである、請求項4に記載の方法。
- 前記培地に糖が添加されている、請求項1〜5のいずれか一項に記載の方法。
- 前記糖が、ブドウ糖または果糖である、請求項6に記載の方法。
- 前記糖が有機である、請求項7に記載の方法。
- 前記培地が、ジュース、酸素源、窒素、食塩源および糖からなる、請求項1に記載の方法。
- 前記培地が、ジュース、酸素源、窒素および食塩源からなる、請求項1に記載の方法。
- 前記培地が、ジュース、酸素源、窒素および糖からなる、請求項1に記載の方法。
- 前記糖が、果糖およびブドウ糖からなる群より選択されるものである、請求項9または11に記載の方法。
- 前記糖が有機認証済である、請求項12に記載の方法。
- 前記食塩源が海水である、請求項9または10に記載の方法。
- 前記海水の塩分濃度が約10ppt〜約35pptの範囲である、請求項14に記載の方法。
- 前記海水の塩分濃度が約12.7pptである、請求項15に記載の方法。
- 培地中で藻類を増殖させ、前記藻類から脂質化合物または組成物を抽出することを含む、1種または複数種の脂質化合物またはそれらの組成物の製造方法であって、
前記培地が、1種または複数種の果実もしくは野菜または果実と野菜の任意の組合せより得られたジュース、酸素源および窒素を含み、
前記培地が、滅菌されており、
前記培地中の窒素が、天然由来の窒素からなる、方法。 - 前記培地に食塩源が添加されている、請求項17に記載の方法。
- 前記食塩源が海水である、請求項18に記載の方法。
- 前記海水の塩分濃度が約10ppt〜約35pptの範囲である、請求項19に記載の方法。
- 前記海水の塩分濃度が約12.7pptである、請求項20に記載の方法。
- 前記培地に糖が添加されている、請求項17〜21のいずれか一項に記載の方法。
- 前記糖が、ブドウ糖または果糖である、請求項22に記載の方法。
- 前記糖が有機である、請求項23に記載の方法。
- 前記培地が、ジュース、酸素源、窒素、食塩源および糖からなる、請求項17に記載の方法。
- 前記培地が、ジュース、酸素源、窒素および食塩源からなる、請求項17に記載の方法。
- 前記培地が、ジュース、酸素源、窒素および糖からなる、請求項17に記載の方法。
- 前記糖が、果糖およびブドウ糖からなる群より選択されるものである、請求項25または27に記載の方法。
- 前記糖が有機認証済である、請求項28に記載の方法。
- 前記培地に食塩源が添加されている、請求項25または26に記載の方法。
- 前記食塩源が海水である、請求項30に記載の方法。
- 前記海水の塩分濃度が約10ppt〜約35pptの範囲である、請求項31に記載の方法。
- 前記海水の塩分濃度が約12.7pptである、請求項32に記載の方法。
- 前記化合物が脂肪酸である、請求項17〜33のいずれか一項に記載の方法。
- 前記脂肪酸が多価不飽和脂肪酸である、請求項34に記載の方法。
- 前記多価不飽和脂肪酸が、DHA、DPA、ピノレン酸およびEPAからなる群より選択されるものである、請求項35に記載の方法。
- 前記多価不飽和脂肪酸がDHAである、請求項36に記載の方法。
- 前記多価不飽和脂肪酸がEPAである、請求項36に記載の方法。
- 前記多価不飽和脂肪酸がピノレン酸である、請求項36に記載の方法。
- 前記化合物がカロチノイドまたは脂溶性ビタミンである、請求項17〜33のいずれか一項に記載の方法。
- 前記カロチノイドがβ−カロチンである、請求項40に記載の方法。
- 食品、化粧品、工業用組成物またはヒトもしくは動物用の医薬組成物の製造方法であって、
培養培地中で藻類を増殖させ、前記培地から藻類バイオマスまたは藻類細胞培養液を回収する工程であって、
前記培地が、1種または複数種の果実もしくは野菜または果実と野菜の任意の組合せに由来するジュース、酸素源、および窒素を含み、
前記培地が滅菌されており、
前記培地中の窒素が天然由来の窒素からなる、工程と、
前記培地から藻類バイオマスまたは藻類細胞培養液を回収する工程と、
前記食品、工業用組成物または医薬組成物を調製する工程
とを含む、方法。 - 藻類バイオマスまたは藻類細胞培養液から1種または複数種の脂質化合物またはそれらの組成物を抽出し、食品、工業用組成物または医薬組成物を調製する工程をさらに含む、請求項42に記載の方法。
- 前記脂質化合物が脂肪酸である、請求項43に記載の方法。
- 前記脂肪酸が多価不飽和脂肪酸である、請求項44に記載の方法。
- 前記多価不飽和脂肪酸が、DHA、DPA、EPAおよびピノレン酸からなる群より選択されるものである、請求項45に記載の方法。
- 前記多価不飽和脂肪酸がDHAである、請求項46に記載の方法。
- 前記多価不飽和脂肪酸がEPAである、請求項46に記載の方法。
- 前記多価不飽和脂肪酸がピノレン酸である、請求項46に記載の方法。
- 前記培地に食塩源が添加されている、請求項42〜49のいずれか一項に記載の方法。
- 前記食塩源が海水である、請求項50に記載の方法。
- 前記海水の塩分濃度が約10ppt〜約35pptの範囲である、請求項51に記載の方法。
- 前記海水の塩分濃度が約12.7pptである、請求項52に記載の方法。
- 前記培地に糖が添加されている、請求項42〜53のいずれか一項に記載の方法。
- 前記糖が、ブドウ糖または果糖である、請求項54に記載の方法。
- 前記糖が有機である、請求項55に記載の方法。
- 前記調製する工程は食品を調整する工程である、請求項42〜56のいずれか一項に記載の方法。
- 前記食品が栄養補助食品である、請求項57に記載の方法。
- 前記藻類が、トラウストキトリウム属、クラミドモナス属、ニッチア属、ナンノクロロプシス属およびオーランチオキトリウム属から選択される属である、請求項1〜58のいずれか一項に記載の方法。
- 前記属がオーランチオキトリウム属である、請求項59に記載の方法。
- 前記属がクラミドモナス属である、請求項59に記載の方法。
- 前記属がナンノクロロプシス属である、請求項59に記載の方法。
- 前記ジュースが、トマトジュース、ビートジュース、にんじんジュース、りんごジュース、ブルーベリージュース、有機認証済のビート糖蜜およびココナッツジュースからなる群より選択されるものである、請求項1〜62のいずれか一項に記載の方法。
- 前記ジュースがビートジュースである、請求項63に記載の方法。
- 前記ジュースがトマトジュースである、請求項63に記載の方法。
- 前記ジュースがにんじんジュースである、請求項63に記載の方法。
- 前記培地中のジュースの%濃度が、約5%〜約70%の範囲内である、請求項1〜66のいずれか一項に記載の方法。
- 前記培地中のジュースの%濃度が、約10%〜約50%の範囲内である、請求項67に記載の方法。
- 前記培地中のジュースの%濃度が、約10%、約25%および約50%からなる群より選択される%濃度である、請求項68に記載の方法。
- 前記ジュースが発酵したものである、請求項1〜69のいずれか一項に記載の方法。
- 前記ジュースが低温殺菌済である、請求項1〜70のいずれか一項に記載の方法。
- 前記ジュースが有機認証済である、請求項1〜71のいずれか一項に記載の方法。
- 前記培地のpHが約3〜約7の範囲である、請求項1〜72のいずれか一項に記載の方法。
- 前記培地のpHが約6.8である、請求項73に記載の方法。
- 前記培地が、化学添加物および保存料を含まない、請求項1〜74のいずれか一項に記載の方法。
- 前記藻類バイオマスまたは藻類細胞培養液が、有機認証済である、請求項1〜16および請求項59〜75のいずれか一項に記載の方法。
- 前記脂質化合物または組成物が有機認証済である、請求項17〜41および請求項59〜75のいずれか一項に記載の方法。
- 前記食品が有機認証済である、請求項42〜75のいずれか一項に記載の方法。
- 前記栄養補助食品が有機認証済である、請求項58に記載の方法。
- 前記ジュースが、ビートジュースまたは有機認証済のビート糖蜜であり、前記ジュースが非GMO由来インベルターゼによって加水分解されている、請求項17に記載の方法。
- 前記培地にブドウ糖および有機認証済の酵母エキスが添加されている、請求項17に記載の方法。
- 前記ジュースが、脂質化合物または組成物の酸化速度を下げる、請求項17に記載の方法。
- 前記脂質化合物がDHAである、請求項82に記載の方法。
- 前記ジュースが、にんじんジュース、ブルーベリージュースおよびビートジュースからなる群より選択されるものである、請求項82に記載の方法。
- 脂質化合物または組成物の酸化速度を下げる方法であって、脂質化合物または組成物を培地中に懸濁させることを含み、前記培地が、1種または複数種の果実もしくは野菜に由来するジュースまたは果実と野菜の任意の組合せに由来するジュースを含む、方法。
- 前記ジュースが、ビートジュース、にんじんジュース、およびブルーベリージュースからなる群より選択されるものである、請求項85に記載の方法。
- 前記脂質化合物がDHAである、請求項85に記載の方法。
- 前記脂質化合物または組成物が、請求項1〜84のいずれか一項に記載の方法によって製造されるものである、請求項85に記載の方法。
- 前記脂質化合物または組成物が藻類由来である、請求項85に記載の方法。
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CA2919676A1 (en) | 2014-12-18 |
BR112015031263A2 (pt) | 2017-07-25 |
WO2014199220A1 (en) | 2014-12-18 |
CN105452442A (zh) | 2016-03-30 |
EP3030647A1 (en) | 2016-06-15 |
KR20230119731A (ko) | 2023-08-16 |
KR20210028279A (ko) | 2021-03-11 |
AU2014279815A1 (en) | 2016-02-11 |
JP7048209B2 (ja) | 2022-04-05 |
BR112015031263B1 (pt) | 2022-10-11 |
JP2016523529A (ja) | 2016-08-12 |
AU2021200394A1 (en) | 2021-03-18 |
AU2014279815B2 (en) | 2020-10-22 |
US20160122706A1 (en) | 2016-05-05 |
HK1226096A1 (zh) | 2017-09-22 |
EP3030647A4 (en) | 2017-05-03 |
KR20160019424A (ko) | 2016-02-19 |
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