JP5886763B2 - 生物活性物質を含む乾燥ガラス状組成物 - Google Patents
生物活性物質を含む乾燥ガラス状組成物 Download PDFInfo
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- JP5886763B2 JP5886763B2 JP2012551295A JP2012551295A JP5886763B2 JP 5886763 B2 JP5886763 B2 JP 5886763B2 JP 2012551295 A JP2012551295 A JP 2012551295A JP 2012551295 A JP2012551295 A JP 2012551295A JP 5886763 B2 JP5886763 B2 JP 5886763B2
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Description
本出願は、2010年1月28日に米国特許商標局に出願された米国仮出願第61/299,315号の優先権を主張するものであり、その内容をあらゆる目的のために本明細書に援用する。
凍結乾燥は、伝統的に、センシティブな生物学的物質(生きているまたは死んだ細菌およびウイルスならびにタンパク質など)を保存するもっとも一般的な方法であるが、噴霧乾燥、流動噴霧乾燥(fluidized spray drying)および乾燥保存などの他の方法は一般的に適していない。このような方法で用いる高い乾燥温度では、生物活性物質自体にかなりの損傷がもたらされる。さらに、これらは、生成物を安定させるために必要な特定の残留水分または水分活性にまで物質を十分に乾燥させることができず、したがって他の手段による更なる乾燥工程が必要となりうる。従来の凍結乾燥プロセスは、典型的には、生物活性物質を含む溶液を凍結させること、および凍結した状態のまま完全真空下で凍結生体材料を凍結乾燥させることを含む。凍結乾燥プロセスの温度は低いので、生物活性物質の分解反応は減少し、最終乾燥形態での活性喪失は最小限に抑えられる。多くの場合、凍結乾燥プロセスでは、ゆっくりした乾燥過程の間に氷の結晶が形成されるため、かなりの活性喪失および生物活性物質の損傷がもたらされる。さらに、凍結工程自体が、正しく実施されないと、生物活性物質を変性させるかまたは不活性化することがある。氷の結晶構造が形成されることによって引き起こされる損傷は、凍結保護剤を生物活性溶液に添加することにより、ある程度回避されうる(Morgan et al., 2006)。そのような保護剤は、凍結時に細胞膜およびタンパク質を保護するため、また貯蔵時の安定性を向上させるために配合物に加えられる、高溶解性化学物質である。生きた細菌およびウイルス用の一般的な安定剤は、細胞物質または生物活性物質と一緒に高い濃度で高糖類(high sugars)(スクロース、グリセロール、またはソルビトールなど)を含む(Morgan et al., 2006; Capela et al., 2006)。しかし、そのような保護剤は、細胞内に十分浸透して細胞内体積中の活性成分を保護することができず、そのため凍結乾燥物質を貯蔵すると不安定になりうる。このような理由で、膜質生体材料(ウイルス、細菌、および細胞など)は凍結乾燥プロセスにおいて十分に生き延びない。それゆえに、乾燥物質の十分な貯蔵安定性を達成しつつ乾燥による損失を最小限に抑える、最適な乾燥プロセスおよび配合物を開発するという重大な課題が残っている。
本明細書で使用される用語は特定の実施形態を説明するためだけのものであり、限定することを意図するものではないことを理解すべきである。
本発明は、粘稠な溶液中に含まれる生物活性物質、マトリックス形成剤およびガラス形成剤の組成物を含む。本発明の配合物は、物理的構造および機能の点で、急速凍結およびパージの有無にかかわらず乾燥させた非粘稠または濃縮配合物とは本質的に異なることが見いだされた。例えば、先行技術の配合物は、最初に沸騰によって「発泡」させて、効率的な乾燥を促進させる。発泡工程は一般に、溶液の激しい沸騰および煮沸が起こる結果となり、それは液体状態での乾燥の避けられない結果である。その結果として、バイアルまたは容器中の物質の充填容量(loading capacity)は非常にわずかなものとなりうる(例えば、最終的な発泡生成物の厚さが2mm未満である、米国特許第6,534,087号を参照)。
本発明による乾燥粉末組成物を調製するために溶液中で好ましい生物活性物質と混合する物質は、少なくとも1種のマトリックス形成剤および少なくとも2種類のガラス形成剤を含む。このような物質は、好ましい生物活性物質と混合すると、液体窒素中でビーズ、マイクロビーズ、糸状体または小滴を形成し、また、本発明の方法にしたがって非晶質ガラス状態で効率的に凍結乾燥または脱水して、前記生物活性物質を貯蔵および投与するための大量の安定した乾燥組成物を得ることができる(乾燥後の種々の配合物の目視観察および顕微鏡観察ならびに水分活性(Aw)については図1Aおよび1Bを参照)。マトリックス形成剤は、構造的安定性を配合物に付与し、向上した乾燥プロファイルおよび/または物理的および化学的に保護するという利点を生物活性物質にもたらす。マトリックス形成剤はまた、真空圧下で配合物の粘度を増大させかつ配合物の特性をより良好に制御できるようにし、さらに本発明の乾燥配合組成物の構造的強度を増大させる(その特定の配合物のガラス状構造および乾燥状態については、図1Bの写真4、4b、4cを参照)。マトリックス形成剤は、多糖とオリゴ糖との混合物を含む。好ましい多糖(特に生きた生物の場合)は水溶性ガムである。これは、穏やかな温度で粘稠ゲルを形成するという独特の特徴があるためである。ある特定濃度のガムも、効果的に配合物を安定化し、非晶質ガラス状構造の形成を促進し、真空下での乾燥プロファイルを向上させることが見いだされた(図1Aの写真3a、3b、3c、4、および図1Bの4cおよび図6を参照)。
生物活性物質を含む安定した乾燥配合物の調製方法は、(1)生物活性物質を溶液中でマトリックスおよびガラス形成剤と混合することにより配合物を調製すること、(2)配合物を急速凍結させて固体凍結粒子を形成させること、(3)凍結粒子を高真空圧に短時間さらして粒子をパージし、その構造を安定化すること、(4)配合物の凍結温度を上回る温度、好ましくは−10℃より高い温度で配合物に熱を加えながら、減圧下で凍結乾燥および/または水分蒸発させて水を除去すること、(5)完全真空および高温下で配合物の水分活性を0.3Aw未満までさらに減らすことを含む。
乾燥配合物は、ひとまとめにして使用するか、所望の形状およびサイズに切断するか、あるいは破砕および粉砕して自由流動性の粉末にすることができ、これにより、湿式または乾式の凝集化、粒状化、錠剤化、圧縮、ペレット化、食品または飼料製品中での混合、または任意の他の種類の送達プロセスなどの下流処理加工が容易になり得る。破砕、微粉砕、粉砕または粉末化の方法は、当該技術分野において周知である。例えば、ハンマーミル、エアミル(air mill)、インパクトミル、ジェットミル、ピンミル、Wileyミル、あるい類似の粉砕装置を使用できる。好ましい粉砕粒子の粒径は、約1000μm未満、好ましくは500μm未満である。
安定した乾燥プロバイオティック物質の調製
基本配合物
75gのトレハロース(Cargill Minneapolis, MN)および22gの高度加水分解カゼイン(Marcor, Carlstadt, NJ)を、3gのアルギン酸ナトリウム(ISP Corp., Wayne, NJ)と乾燥形態で均一に混合した。ラクトバチルス・アシドフィルス(Lactobacillus acidophilus)のフレッシュな濃縮物(100ml、少なくとも10%の固形分を含み、発酵回収物からの直接のもの)をブレンダー内に加え、35℃に維持した。ガムと糖とタンパク質加水分解物との乾燥混合物をプロバイオティック培養物にゆっくり加え、35℃で10分間混合した。次いで粘稠なスラリーを、孔底を有する容器に移し、窒素を含む浴に滴らせた。次いでビーズを液体窒素から取り除き、乾燥させるためにすぐに移した。
凍結ビーズを100g/sq ftの充填容量でトレー上に均一に広げ、すぐに凍結乾燥機(Model 25 SRC, Virtis, Gardiner, NY)内の棚に置いた。次いで1000mTORRの真空圧にし、固体の凍結ビーズを10分間パージした。次いで真空を2700mTORRに調整し、棚の温度を+30℃まで上昇させた。これらの温度および真空圧を3時間維持した。次いで完全真空(150〜200mTORR)において第2の乾燥工程を行い、棚の温度をさらに2時間30℃まで上昇させた。配合物を完全に乾燥させて、その水分活性をHygropalm Awl測定器(Rotonic Instrument Corp., Huntington, NY.)で測定すると、Aw=0.23であった。
プロバイオティック細菌であるラクトバシラス・ラムノサス(Lactobacillus rhamnosus)LGGを含む安定した乾燥組成物。
ラクトバシラス・ラムノサス(Lactobacillus rhamnosus)LGG(商品供給元からの500gの凍結濃縮物)を、ジャケット付二重遊星形ミキサー(DPM, lqt, Ross Engineering, Inc. Savannah, GA,)内において37℃で解凍した。2種類のガラス形成剤(トレハロース(387g、Cargill Minneapolis, MN)および高度加水分解カゼイン(83g、Marcor, Carlstadt, NJ))を、乾燥形態で均一に2種類のマトリックス形成剤(アルギン酸ナトリウム(15g、ISP Corp., Wayne, NJ)およびインスタントの(instant)イヌリン(25g,Cargill Minneapolis, MN))と混合した。乾燥混合物を解凍されたプロバイオティック細菌にゆっくり加え、40RPMおよび37℃で10分間、混合した。50〜200mlの水を加えて、スラリーの粘度を12,000cPに調整した。次いでスラリーを、孔底を有する容器に移し、液体窒素を含む容器中に滴らせた。次いでビーズを液体窒素から取り除き、アルミニウム箔の施された密封バッグに入れ、ディープフリーザー内において−80℃で数週間貯蔵した。
トレハロース(752g、Cargill Minneapolis, MN)、高度加水分解エンドウ豆タンパク質(167g、Marcor, Carlstadt, NJ)、アルギン酸ナトリウム(30g、ISP Corp., Wayne, NJ)およびインスタントのイヌリン50g(Cargill Minneapolis, MN)を乾燥形態で均一にブレンドした。乾燥混合物をジャケット付二重遊星形ミキサー(DPM, 1qt, Ross Engineering, Inc. Savannah, GA,)内において80℃で1000mlの熱い脱イオン水にゆっくり加え、40RPMで10分間、混合した。混合物の温度を37℃まで下げて、商品供給元から得た100gの乾燥粉末のラクトバシラス・ラムノサス(Lactobacillus rhamnosus)LGGをゆっくり加え、20分間混合を継続した。次いでスラリーを2mmの孔の針に通して押し出して、液体窒素を含む浴に入れた。次いで糸状体/ビーズを液体窒素から取り除き、アルミニウム箔の施された密封バッグに入れ、ディープフリーザー内において−80℃で数週間貯蔵した。乾燥させるため、凍結した糸状体/ビーズを100から500g/sq ftまでの範囲の充填容量でトレー上に均一に広げ、トレーを凍結乾燥機(Model 25 SRC, Virtis, Gardiner, NY)内の棚に置き、実施例2で記載したようにして乾燥させた。すべての配合物は満足すべきことにトレー内にとどまり、すべての充填レベルで飛散も発泡も観察されなかった。配合物はより大きな充填容量でも完全に乾燥した。また水分活性を測定するとすべての試料で0.26Aw以下であった。
プロバイオティック細菌であるビフィドバクテリウム・ラクティス(Bifidobacterium lactis)(Bb12)を含むヒドロゲル配合物の調製:
ビフィドバクテリウム・ラクティス(Bifidobacterium lactis)(Bb12)の濃縮プロバイオティックスラリーを、実施例1にしたがって調製する。その基本配合物に、0.5gの二塩基性リン酸カルシウムを加え、その後、0.5gのグルコノラクトンを加える。スラリーは、室温でその後の2時間にわたって固まるようにさせて固体のヒドロゲルを形成させる。市販のスライサー/シュレッダーを用いて、堅いゲルをスライスして細長い糸状体にする。細い糸状体を液体窒素で急速凍結させ、トレー上に700g/sq ftの充填容量で載せ、乾燥させるために実施例2で記載したようにして凍結乾燥機内に入れる。配合物の水分活性(Aw)は0.05であった(HygroPalm Awl, Rotonic Huntington, NY)で測定)。標準ハンマー粉砕装置を用いて乾燥配合物をさらに粉砕して微粉にし、50〜250ミクロンの篩で分けた。
プロバイオティック細菌であるラクトバチルス・アシドフィルス(Lactobacillus acidophilus)を含み、アレルゲンを含まない組成物
トレハロース(752g、Cargill Minneapolis, MN)、高度加水分解エンドウ豆タンパク質(167g、Marcor, Carlstadt, NJ)、アルギン酸ナトリウム(30g、ISP Corp., Wayne, NJ)およびインスタントのイヌリン50g(Cargill Minneapolis, MN)を乾燥形態で均一にブレンドした。乾燥混合物をジャケット付二重遊星形ミキサー(DPM, 1qt, Ross Engineering, Inc. Savannah,GA,)内において80℃で1000mlの熱い脱イオン水にゆっくり加えることにより殺菌し、なめらかで透明なスラリーが形成されるまで40RPMで10分間、混合した。混合物の温度を37℃まで下げて、商品供給元から得たラクトバチルス・アシドフィルス(Lactobacillus acidophilus)を含む1000gの凍結ビーズをゆっくり加え、10分間混合を継続した。次いでスラリーを2mmの孔の針に通して押し出して、液体窒素を含む浴に入れた。次いで糸状体/ビーズを液体窒素から取り除き、アルミニウム箔の施された密封バッグに入れ、ディープフリーザー内において−80℃で数週間貯蔵した。乾燥させるため、凍結した糸状体/ビーズを1000g/sq ftの充填容量でトレー上に均一に広げ、トレーを凍結乾燥機(Model 25 SRC, Virtis, Gardiner, NY)内の棚に置き、実施例2で記載したようにして乾燥させた。乾燥組成物中のプロバイオティック細菌の初期CFUカウントは10.53 logs/gであった。また40℃および33%RHという加速貯蔵条件下で42日間貯蔵した後の生存率低下は、0.69 log CFU/gであった。
本発明の乾燥配合物を含む乳児用調製粉乳:
ラクトバシラス(Lactobacillus)GG(Valio Corp, Finland)を含む安定した乾燥配合物を実施例2にしたがって調製し、その後、2種類の粒径群(50μm超と150μm未満)に篩で分けた。99.9gのNutramigen(Mead Johnson; Evansville, IL)を50μm〜150μmの粒径範囲の乾燥配合物粒子0.1gと混ぜることによって、乳児用調製粉乳を調製した。最終生成物は、100gの乳児用調製粉乳当たり約108cfuのラクトバシラス(Lactobacillus)GGを含んでいる。
本発明の安定した乾燥配合物を含むプロバイオティック栄養補助品:
ラクトバチルス・アシドフィルス(Lactobacillus acidophilus)を含む安定した乾燥組成物を配合して、経口剤形(錠剤、カプレット、またはカプセル剤など)にする。圧縮剤(デキストロース)99.9gとサイズ範囲が50μm〜150μmの乾燥配合物粒子0.1gとを含む風味を付けたオレンジ色錠剤を、1/2インチの丸い標準凹型工具を使用して回転機での直接圧縮によって調製する。最終生成物は、約108cfu/単位用量を含む。錠剤の硬度は8〜10kpの範囲であり、崩壊時間はおよそ20秒である。圧縮錠剤を180ccのHDPE瓶に詰め(それぞれに100錠)、制御された温度/湿度(40℃/33%RH)にさらす。その生成物に対して、月1回の微生物学的安定性試験を、12ヶ月の期間にわたって、あるいは観察される試験カウントの減少が1×106/単位用量未満になるまで、実施する。
本発明の安定した乾燥配合物を含む機能性飲料
(重量%で)71%のスクロース、14%のマルトデキストリン、10%のイヌリン、2%のデキストロース、1%の無水クエン酸、0.3%のアラビアガム、0.3%の香料、0.3%のリン酸三カルシウムおよび0.1%の乾燥プロバイオティック配合物粒子(L.アシドフィルス(L. acidophilus))(50μm〜250μmの粒径範囲のもの)を含む乾燥混合物を調製する。最終生成物は、約109cfu/単位用量(30gの乾燥混合物)を含む。その生成物を、340mlの水の中で撹拌して、飲料用に小さなアルミニウム箔バッグに詰める(30g単位用量/バッグ)。飲料用乾燥混合物中のプロバイオティック細菌の安定性に対して、12ヶ月の期間にわたって、あるいは観察される試験カウントの減少が1×107/単位用量未満になるまで、月1回の微生物学的安定性試験を実施する。
プロバイオティックペットフードの調製:
市販の犬用ペレットペットフードを、水分活性が0.1になるまで熱対流炉中で乾燥させてから、実施例3で記載されているように調製された安定したプロバイオティック乾燥配合物でコーティングする。乾燥ペレットに、約5%の脂肪系防湿剤(fat-based moisture barrier)(40%の鶏脂、40%のカカオ脂、および20%の蜜ろうの混合物)を吹き付け、ドラムタンブラー中で乾燥粉末配合物(普通、ペットフード全体の0.1〜0.5%であり、108 CFU/gの服用量となる)と混合し、最後に脂肪系防湿剤のコーティングをさらに吹き付ける。コーティングの総量は、(ペットフードの)約15%である。コーティング時間は約30分間である。
数種類のプロバイオティック微生物を含む魚飼料の調製:
本発明による魚用ペレット飼料は、数種類のプロバイオティックスの混合物を用いて調製する。L.ラムノサス(L. rhamnosus)とL.アシドフィルス(L. acidophilus)とビフィドバクテリウム・ラクティス(Bifidobacterium lactis)との混合物を含む安定した乾燥プロバイオティック配合物を、実施例1に記載されているようにして調製する。市販のサケ用出発飼料(Zeigler Bros., Gardners, PA)を、最初に水分活性が0.1になるまで熱対流炉中で乾燥させ、その後、ドラムタンブラー中においてプロバイオティックス配合物でコーティングする。ペレット(1000g)に、最初に約5重量%の脂肪系防湿剤(40%の魚油、40%のカカオ脂および20%の蜜ろうの混合物)を吹き付け、その後、1gの安定した乾燥プロバイオティック配合物(107cfu/g(飼料)の服用量となる)と混合し、最後に脂肪系防湿剤のさらなるコーティングを吹き付ける。コーティングの総量は、(魚飼料の)約10%である。
酵素を含む安定した乾燥粉末:
40重量パーセントのサビナーゼ(Savinase)(Novozymes, Denmark)を含むヒドロゲル配合物を、実施例4で記載した配合物600gおよびサビナーゼ400gを水溶液1000g中で混合することにより調製する。細かくきざんだヒドロゲル配合物を液体窒素で急速凍結させ、真空オーブン内において50℃の配合物乾燥温度で乾燥させる。乾燥配合物の投入量および貯蔵安定性を測定するため、乾燥試料を正確に秤量して(<100mg)微小遠心管に入れる。200μlのジメチルスルホキシド(DMSO)を加える。配合物をボルテックスによってDMSO緩衝液に溶かす。この試料に、0.05N NaOH、0.5%のSDSおよび0.075Mクエン酸(三ナトリウム塩)を含む0.8mlの溶液を加える。管を45℃で10分間超音波処理し、その後、5,000rpmで10分間短く遠心分離する。透明なDMSO/NaOH/SDS/シトレート溶液のアリコットをマイクロプレートのウェルに入れ、ブラッドフォード(Bradford)試験法を用いてタンパク質含有量を分析する。この安定した酵素配合物の貯蔵安定性は、本発明の配合物を用いない乾燥酵素よりも著しく大きい。
ビタミンAを含む安定した乾燥粉末:
インスタントのイヌリン320g、マルトデキストリンDE−l(Tate&Lyle, London, UK)320g、カルボキシルメチルセルロースナトリウム(Ashland Aqualon Functional Ingredients, Wilmington, DE)50g、アスコルビン酸ナトリウム10gおよび結晶ビタミンA(BASF Corp., Florham Park, N.J)300gを水1000g中で混合することにより、30重量パーセントのビタミンAを含む配合物を調製する。湿った配合物を、Mobile-Minor噴霧乾燥機(GEA Process Engineering Inc., Columbia MD)で入口温度および出口温度がそれぞれ180℃および80℃において噴霧乾燥させるか、あるいは液体窒素で急速凍結させてから、1000g/sq ftの充填容量でトレー上に広げ、実施例2に記載したようにして乾燥させる。ビタミンAの組成物は、40℃および75%RHで3ヶ月間安定している(>80%)。
生物学的利用能の向上した保護配合物中のカロテンの調製:
貯蔵時または生物に食べさせた後に飼料中の他の成分によって酸化されるであろうカロテンの生物学的利用能を保護および向上させる配合物を、本発明の配合および方法にしたがって調製する。6gの水溶性キトサン(LSK BioPartners, Inc. Salt Lake City, Utah)を含む配合物を水200gに溶かす。この溶液に、天然アスタキサンチン(Naturose(商標), Cyanotech Corp., Kailua-Kona, HI))90gを加え、そのスラリーを5%のトリポリリン酸ナトリウムを含む浴に噴霧または押し出す。ヒドロゲル化微小粒子または糸状体を室温で4時間にわたって硬化させる。粒子を架橋浴から取り出し、水で洗い、スクロース90gと高度加水分解カゼイン10gとのドライブレンドと混合する。糖/タンパク質を担持した粒子を急速凍結させ、ただちに500g/sq ftでトレーに載せ、凍結乾燥機で水分活性が0.3未満に減少するまで凍結乾燥させた。乾燥配合物をさらに粉砕して所望の粒度分布になるようにし、包装する。
侵入性種餌(Invasive Species Bait)の調製
特に標的とした侵入性種のためのペレット餌を本発明にしたがって調製する。実施例1に記載した、殺虫剤を含む200gの配合物を調製し、200gmの水に加える。この溶液に、ロテノン90gmおよびリン酸水素カルシウム0.5gm、その後に0.5gmのグルコノラクトンを加える。スラリーを室温で2時間にわたって硬化させる。堅いゲルを、スライサー/シュレッダーによって細長い糸状体にスライスする。その細い糸状体をトレー上に充填し、凍結乾燥機に入れる。棚の温度を−30℃に設定し、配合物を凍結させてから完全真空にし、さらに一晩乾燥させるために棚の温度を+60℃に上昇させる。乾燥配合物は、標的とする特定種のための餌の大きさ仕様に合った適切な粒度分布になるまで粉砕する。
水不溶性配合物中の保護殺虫剤の調製:
貯蔵時または施した後にその環境で配合物中の他の成分によって分解するであろう殺虫剤の保護粒状配合物を、本発明の配合および方法で調製する。6gのペクチンおよび102gのスクロースを含む配合物を、水200gに加える。この溶液に、センシティブな殺虫剤の乾燥配合物90gおよびリン酸水素カルシウム1.5gと塩化カルシウム0.5gとを含む混合物を加え、その後に0.85gのグルコノラクトンを加える。そのスラリーを室温で4時間にわたって硬化させ、その後スライサー/シュレッダーによってスライスして細長い糸状体にする。細い糸状体をトレー上に充填し、凍結乾燥機で乾燥させて水分活性が0.1になるようにする。乾燥配合物をさらに粉砕して所望の粒度分布になるようにし、包装する。
保護植物プロバイオティック配合物の調製:
根圏細菌(Rhizobacteria)などの生物農薬を、実施例4にしたがって乾燥組成物の形で調製する。根圏細菌(Rhizobacteria)乾燥組成物の効果は、純粋隔離群条件下でのレタスの成長に基づいて評価する。用量として1つの植物当たり100mgの根圏細菌(Rhizobacteria)乾燥組成物を砂と一緒に広口瓶に接種し、事前に発芽させた(24h)レタスの苗木を植える。栄養素の用量として5mlの殺菌されたホーグランド(Hoagland)溶液を広口瓶中の植物に施す。光周期が12時間である28℃に維持されたグロースチャンバー内に広口瓶を無作為に並べる。接種後、7日間の間隔ごとの間に、植物とそれに付着する土を注意深く広口瓶から取り出す。殺菌したリン酸緩衝液(pH7.0)で根を洗い、根の長さの測定を記録する。
以下の文献および本明細書で引用されているすべての文献を、あらゆる目的のために本明細書に援用する。
6,190,701 Composition and method for stable injectable liquids, March 1999, Roser et al.
6,964,771 Method for stably incorporating substances within dry, foamed glass matrices, September 1997, Roser et al.
5,766,520 Preservation by formulation formation, June 1998,
Bronshtein
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7,153,472 Preservation and formulation of bioactive materials for storage and delivery in hydrophobic carriers, December, 2006, Bronshtein
20080229609 Preservation by Vaporization, June 2005, Bronshtein
6,306,345 Industrial scale barrier technology for preservation of sensitive biological materials at ambient temperatures, October 2001, Bronshtein et al.
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Claims (18)
- 沸騰又は発泡をさせることなく乾燥ガラス状組成物の調製方法であって、
(a)生物活性物質、少なくとも1種のマトリックス形成剤および少なくとも2種類のガラス形成剤を水性溶媒中で混ぜ合わせて粘稠なスラリーを生じさせることであって、
前記生物活性物質が、細胞、ウイルス、細菌、酵母、タンパク質、ペプチド、ホルモン、ワクチン、薬物、抗生物質、ビタミン、カロチノイド、ミネラル、殺菌剤、殺真菌剤、除草剤、殺虫剤または殺精子剤を含み、
前記少なくとも1種のマトリックス形成剤のそれぞれが、酢酸フタル酸セルロース(CAP)、カルボキシ−メチル−セルロース、ペクチン、アルギン酸ナトリウム、アルギン酸塩、ヒドロキシルプロピルメチルセルロース(HPMC)、メチルセルロース、カラギナン、グアーガム、アラビアガム、キサンタンガム、ローカストビーンガム、キトサンおよびキトサン誘導体、デンプンおよび加工デンプン、シクロデキストリン、イヌリン、マルトデキストリン、およびデキストランからなる群から選択され、
前記少なくとも2種類のガラス形成剤が、タンパク質、炭水化物、アミノ酸、メチルアミン、ポリオール、プロピレングリコール、ポリエチレングリコール、界面活性剤、およびリン脂質からなる群から選択される、ことと;
(b)前記スラリーを液体窒素中で急速凍結させて、ビーズ、小滴、または糸状体の形態の固体の凍結した粒子を生じさせることと;
(c)前記凍結粒子の凍結温度を上回る温度で真空下において蒸発させることにより前記凍結粒子を第1乾燥させることであって、圧力は2000mTORRより高く、前記粒子の温度が−10℃より高く、これにより第1乾燥配合物が形成される、ことと;
(d)完全真空および20℃以上の温度で、前記第1乾燥配合物の水分活性をAw0.3以下に下げるのに十分な時間、前記第1乾燥配合物を第2乾燥させ、これにより前記乾燥ガラス状組成物が調製されることと、
を含む、方法。 - 前記第2乾燥工程の温度が20℃〜70℃である、
請求項1記載の方法。 - 前記第2乾燥工程の間に前記温度又は真空度が増加される、
請求項1記載の方法。 - 前記組成物を切断、破砕、粉砕、または粉末化して自由流動性の粉末にすることをさらに含む、
請求項1に記載の方法。 - 前記粉末の粒径が1000μm未満である、請求項4に記載の方法。
- 前記組成物を、乳児用調製粉乳、機能性飲料、およびペットフードからなる群から選択される成分と混合することをさらに含む、
請求項1に記載の方法。 - 請求項1に記載の方法に従って調製された組成物。
- 請求項2に記載の方法に従って調製された組成物。
- 前記第1乾燥工程における圧力が2000mTORR〜10000mTORRである、
請求項1に記載の方法。 - 前記第1乾燥工程における圧力が2000mTORR〜4000mTORRである、
請求項1に記載の方法。 - 前記第1乾燥工程における温度が−5℃〜5℃である、
請求項1に記載の方法。 - 生物活性物質、少なくとも1種のマトリックス形成剤、および少なくとも2種類のガラス形成剤を含み、請求項1に記載の方法により調製された、乾燥ガラス状組成物。
- 前記組成物が、30重量パーセント〜70重量パーセントの範囲の全固形分を含む、請求項12に記載の組成物。
- 前記少なくとも1種のマトリックス形成剤が、1重量パーセント〜20重量パーセントの範囲の量で前記組成物中に存在する、請求項12に記載の組成物。
- 前記少なくとも2種類のガラス形成剤が溶液に溶け、水と接触したときに増粘または重合せず、乾燥時に結晶を生じない、請求項12に記載の組成物。
- 前記少なくとも2種類のガラス形成剤が、1重量パーセント〜80重量パーセントの範囲の量で前記組成物中に存在する、請求項12に記載の組成物。
- 前記少なくとも1種のマトリックス形成剤が、アルギン酸塩およびイヌリンからなり、前記少なくとも2種類のガラス形成剤が、トレハロースおよびタンパク質加水分解物からなる、請求項1に記載の方法。
- 前記少なくとも1種のマトリックス形成剤が、アルギン酸塩およびイヌリンからなり、前記少なくとも2種類のガラス形成剤が、トレハロースおよびタンパク質加水分解物からなる、請求項12に記載の組成物。
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ES2639397T3 (es) | 2017-10-26 |
CN102725393B (zh) | 2015-12-02 |
CA2785815A1 (en) | 2011-08-04 |
US9731020B2 (en) | 2017-08-15 |
CN102725393A (zh) | 2012-10-10 |
US8834951B2 (en) | 2014-09-16 |
EP2529004B1 (en) | 2017-06-07 |
RU2535869C2 (ru) | 2014-12-20 |
US20120322663A1 (en) | 2012-12-20 |
JP2013517801A (ja) | 2013-05-20 |
BR112012018839A2 (pt) | 2015-09-15 |
EP2529004A2 (en) | 2012-12-05 |
MX336076B (es) | 2016-01-07 |
WO2011094469A2 (en) | 2011-08-04 |
WO2011094469A3 (en) | 2011-12-29 |
DK2529004T3 (en) | 2017-09-25 |
SG182317A1 (en) | 2012-08-30 |
MX2012008795A (es) | 2012-08-17 |
EP2529004A4 (en) | 2013-10-23 |
US20150031544A1 (en) | 2015-01-29 |
NZ601017A (en) | 2014-07-25 |
PL2529004T3 (pl) | 2017-12-29 |
RU2012134269A (ru) | 2014-03-10 |
CA2785815C (en) | 2018-04-24 |
AR080073A1 (es) | 2012-03-14 |
AR125029A2 (es) | 2023-05-31 |
BR112012018839B1 (pt) | 2020-04-14 |
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