JP4972741B2 - How to use shochu residue - Google Patents
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Description
廃棄していた焼酎カスを、微細藻類の増殖促進効果等へ利用する方法に関する。 The present invention relates to a method for utilizing the discarded shochu residue for promoting the growth of microalgae.
特許文献１には、本発明者らによる焼酎カスの処理方法およびそれにより得られる有効液の水産および畜産への応用が記載されている。
特許文献１の利用方法では、充分な付加価値が得られず実用化まで至っていない。
In the utilization method of
そこで、本発明は、焼酎カスの蒸留段階を細分化して、有効成分の濃度を高めると共に、それ以外の不要画分は廃棄しやすい形態になるように工夫することを課題とする。更に得られた有効成分をより付加価値の高い例えば機能性食品分野に利用することにより、焼酎カスの処理コストの削減に寄与することを課題とする。 Therefore, an object of the present invention is to subdivide the distillation stage of shochu residue to increase the concentration of the active ingredient and to devise a form that makes it easy to discard other unnecessary fractions. Furthermore, it makes it a subject to contribute to reduction of the processing cost of shochu residue by utilizing the obtained active ingredient in the functional food field with higher added value.
本願発明者は、鋭意研究の結果、焼酎カスから取り出された有効成分を用いて付加価値の高い分野への利用を開発し、上記課題を解決した。
すなわち、本発明は、焼酎カスを攪拌・混練（羽根型、スクリュウ型、ブラベンダー型）し、減圧蒸留しながら個液分離する処理方法において、１段階では、焼酎カスに何も加えず低圧低温（６５〜８０度Ｃ、２００〜４００トール）で蒸留し、ＢＯＤおよびアンモニアを高濃度に含む（本工程で得られる総量の３０〜５０％に相当する）画分を抽出する（Ａ液）。次に、２段階では、焼酎カスに不揮発性酸を加えｐＨ２まで下げたのち、低圧低温（６５〜８０度Ｃ、２００〜４００トール）で蒸留し、酢酸とプロピオン酸を主成分とする有機酸を高濃度に含む（本工程で得られる総量の５０〜８０％に相当する）画分を抽出する（Ｂ液）。更に、３段階では、焼酎カスにアルカリを加え、ｐＨを６〜７に中和して（６５〜８０度Ｃ、２００〜４００トール）で蒸留し、ＢＯＤが２０ｐｐｍ以下の放流可能な水として抽出する（Ｃ液）。そして、２段階で得られたＢ液を有効成分として、機能性食品となる微細藻類の増殖促進剤に用いる方法である。
As a result of earnest research, the inventor of the present application has developed the use of high-value-added fields using active ingredients extracted from shochu residue and solved the above problems.
That is, the present invention is a treatment method in which shochu residue is stirred and kneaded (blade type, screw type, Brabender type) and separated into individual liquids while distillation under reduced pressure. (65 to 80 degrees C, 200 to 400 Torr), a fraction containing BOD and ammonia at a high concentration (corresponding to 30 to 50% of the total amount obtained in this step) is extracted (Liquid A). Next, in the second stage, after adding a non-volatile acid to the shochu residue and lowering it to
ここで、本発明にいう不揮発性酸とは、無機酸および／または有機酸であり、無機酸として硫酸、リン酸であり、有機酸として酢酸とプロピオン酸よりも分子量の高い有機酸である。 Here, the non-volatile acid referred to in the present invention is an inorganic acid and / or an organic acid, sulfuric acid or phosphoric acid as the inorganic acid, and organic acid having a higher molecular weight than acetic acid and propionic acid as the organic acid.
又、本発明にいうアルカリとは、水酸化マグネシウム、酸化マグネシウムである。 The alkali referred to in the present invention is magnesium hydroxide and magnesium oxide.
次に、本発明にいう微細藻類とはスピルリナ・プラテンシス（Ｓｐｉｒｕｌｉｎａ
ｐｌａｔｅｎｓｉｓ）のことである
Next, the microalgae referred to in the present invention is Spirulina platensis (Spirulina).
platenesis)
［焼酎カスの処理方法］焼酎製造過程で生成された焼酎カスを使用する。この焼酎カスは芋、麦、米、黒糖などを発酵させ常圧蒸留した残りのものである。この焼酎カスを、容器に入れ、攪拌および／または混練しながら以下の３段階にわけて機械・装置を用いる。 [Treatment method of shochu residue] Shochu residue generated in the process of producing shochu is used. This shochu residue is the remainder of fermented straw, wheat, rice, brown sugar, etc., and distilled at atmospheric pressure. The shochu residue is put into a container, and the machine / device is used in the following three stages while stirring and / or kneading.
第１段階として低圧低温（６５〜８０度Ｃ、２００〜４００トール）で蒸留し、２５〜３５Ｗｔ％の処理水を得た。この１段階で得られる処理液をA液とする。２段階では、焼酎カスに不揮発性酸を加えｐＨ２まで下げたのち、低圧低温（６５〜８０度Ｃ、２００〜４００トール）で蒸留し、２５〜３５Ｗｔ％の処理水を得た。この２段階で得られる処理液をＢ液とする。 As a first stage, distillation was performed at low pressure and low temperature (65 to 80 ° C., 200 to 400 Torr) to obtain 25 to 35 Wt% treated water. The treatment liquid obtained in this one stage is designated as liquid A. In the second stage, a non-volatile acid was added to the shochu residue to lower the pH to 2, and then distilled at low pressure and low temperature (65 to 80 ° C., 200 to 400 Torr) to obtain 25 to 35 Wt% treated water. The treatment liquid obtained in these two stages is designated as B liquid.
第３段階として底部に水酸化マグネシウムを入れてｐＨを６〜７に調整した。この場合、水酸化マグネシウムの代わりに、酸化マグネシウムや水酸化カルシウムなど他のアルカリでも良い。ただし、底部に残った焼酎カスは家畜の飼料にするので、飼料として不都合なアルカリは避ける事が好ましい。この焼酎カスをｐＨ６〜７の状態で、６５〜８０度Ｃ、２００〜４００トールで蒸留し、２５〜３５Ｗｔ％の処理水を得た。この第３段階で得られる処理液をＣ液とする。
As the third stage, magnesium hydroxide was added to the bottom to adjust the pH to 6-7. In this case, other alkalis such as magnesium oxide and calcium hydroxide may be used instead of magnesium hydroxide. However, since the shochu residue remaining at the bottom is used as livestock feed, it is preferable to avoid alkali that is inconvenient as feed. The shochu residue was distilled at 65 to 80 ° C. and 200 to 400 Torr at
最終的に底部に残った焼酎カスの蒸留残物の含水率は約２０Ｗｔ％である。Ａ液はアンモニア濃度が高いのが特徴である。Ｂ液はｐＨが３以下で酢酸とプロピオン酸の濃度が高いのが特徴である。Ｃ液はＡ液、Ｂ液に比べるとＢＯＤと窒素濃度が極めて低いのでそのままか、あるいは水道水で多少希釈して排水しても差し支えない。また、蒸留残分は特有の悪臭が取り除かれるので、家畜の飼料として用いる事が可能である。 The water content of the distillation residue of the shochu residue finally remaining at the bottom is about 20 Wt%. Liquid A is characterized by a high ammonia concentration. Liquid B is characterized by a pH of 3 or less and a high concentration of acetic acid and propionic acid. The C liquid has a much lower BOD and nitrogen concentration than the A and B liquids, so it can be left as it is or can be slightly diluted with tap water. In addition, the distillation residue can be used as livestock feed because it removes the peculiar odor.
焼酎カスの処理方法：実験には、宮内酒造の焼酎製造過程で生成された焼酎カスを使用した。この焼酎カスは芋を発酵させ常圧蒸留した残りのものである。使用した焼酎カスの成分は表１に示す。この焼酎カスを、以下の３段階に分けて低圧低温蒸留した。具体的には、ロータリーエバポレーターの底部に１Ｌの焼酎カスを入れ、１段階として６０〜８０度Ｃ、２００〜４００トールで蒸留し２００ｍｌ（２０Ｗｔ％）の処理水を得た。この１段階で得られる処理液をＡ液とした。次に底部に希硫酸を入れてｐＨを２に調整した。この場合、希硫酸の変わりに他の燐酸などの酸でも良い。ただし、揮発性の塩酸や硝酸は使用しない。この焼酎カスをｐＨ２の状態で、６５〜８０度Ｃ、２００〜４００トールで蒸留し３００ｍｌ（３０Ｗｔ％）の処理水を得た。この２段階で得られる処理液をＢ液とした。
Shochu residue processing method: In the experiment, shochu residue generated during the manufacturing process of Miyauchi Sake Brewery was used. This shochu residue is the remainder of fermented koji and atmospheric distillation. The components of the shochu residue used are shown in Table 1. This shochu residue was divided into the following three stages and subjected to low-pressure low-temperature distillation. Specifically, 1 L of shochu residue was put at the bottom of the rotary evaporator, and distilled at 60 to 80 degrees C and 200 to 400 torr as one step to obtain 200 ml (20 Wt%) of treated water. The treatment liquid obtained in this one stage was designated as A liquid. Next, dilute sulfuric acid was added to the bottom to adjust the pH to 2. In this case, other acids such as phosphoric acid may be used instead of dilute sulfuric acid. However, do not use volatile hydrochloric acid or nitric acid. This shochu residue was distilled at 65 to 80 ° C. and 200 to 400 Torr at
次に底部に水酸化マグネシウムを入れてｐＨを６に調整した。この場合、水酸化マグネシウムの代わりに、酸化マグネシウムや水酸化カルシウムなど他のアルカリでも良い。ただし、底部に残った焼酎カスは家畜の飼料にするので、飼料として不都合なアルカリは避ける。この焼酎カスをｐＨ６の状態で、６５〜８０度Ｃ、２００〜４００トールで蒸留し３００ｍｌ（３０Ｗｔ％）の処理水を得た。この３段階で得られる処理液をＣ液とした。最終的に底部に残った焼酎カスの含水率は約２０Ｗｔ％であった。
Next, magnesium hydroxide was added to the bottom to adjust the pH to 6. In this case, other alkalis such as magnesium oxide and calcium hydroxide may be used instead of magnesium hydroxide. However, since the shochu residue left at the bottom is used as livestock feed, avoid alkali that is inconvenient as feed. This shochu residue was distilled at 65 to 80 ° C. and 200 to 400 Torr at
得られたＡ液、Ｂ液、Ｃ液の成分分析の結果を表２に示す。表２に示すとおり、Ａ液はアンモニア濃度の高いことが特徴である。Ｂ液はｐＨが３以下で酢酸とプロピオン酸の濃度の高いことが特徴である。Ｃ液はＡ液、Ｂ液に比べるとＢＯＤと窒素濃度が極めて低いのが特徴である。Ｂ液は有機酸が豊富に存在するので以下の有効利用の検討を行った。Ｃ液はＢＯＤと窒素濃度が極めて低いので、そのままか、あるいは水道水で多少希釈して排水しても差し支えない。また、残った焼酎カスは特有の悪臭が取り除かれるので、家畜の飼料として運搬や使用に支障をきたす心配が無い。 Table 2 shows the results of component analysis of the obtained liquid A, liquid B and liquid C. As shown in Table 2, the liquid A is characterized by a high ammonia concentration. Solution B is characterized by a pH of 3 or less and a high concentration of acetic acid and propionic acid. The C liquid is characterized by extremely low BOD and nitrogen concentration compared to the A and B liquids. Since B liquid is rich in organic acids, the following effective utilization was examined. Since C liquid has very low BOD and nitrogen concentration, it can be drained as it is or with some dilution with tap water. In addition, since the remaining shochu residue removes the peculiar odor, there is no fear of hindering transportation and use as livestock feed.
微細藻類のＳｐｉｒｕｌｉｎａ ｐｌａｔｅｎｓｉｓ（NIES-39）は国立環境研究所微生物系統保存施設から購入した。培養に用いた培養液は国立環境研究所微生物系統保存施設が指定しているSOT培地を用いた。
培養には、まず上記の培養液を三角フラスコに入れ、滅菌及び放冷した後、クリーンベンチ内で無菌的に接種した。次いで人工気象機（SANYO GROWTH CABINET MLR350T）を用い温度30℃、照度100μmol photns/m2/s1、12時間明12時間暗のサイクルで培養した。Ｓｐｉｒｕｌｉｎａ ｐｌａｔｅｎｓｉｓは細胞が凝集又は沈殿してしまうため、滅菌したマグネットをフラスコに入れ、採取の際にスターラー（Pasolina STIRRER TR-300）で5分間攪拌した。
The microalga Spirulina platensis (NIES-39) was purchased from the National Institute for Environmental Studies, Microbial System Storage Facility. The culture medium used for the culture was the SOT medium designated by the National Institute for Environmental Studies microbial strain preservation facility.
For the culture, the above culture solution was first put into an Erlenmeyer flask, sterilized and allowed to cool, and then aseptically inoculated in a clean bench. Subsequently, using an artificial weather machine (SANYO GROWTH CABINET MLR350T), the cells were cultured at a temperature of 30 ° C., an illuminance of 100 μmol photns / m 2 / s 1 , and a cycle of 12 hours light and 12 hours dark. Since Spirulina platensis causes cells to aggregate or precipitate, a sterilized magnet was placed in the flask and stirred with a stirrer (Pasolina STIRRER TR-300) for 5 minutes.
Ｓｐｉｒｕｌｉｎａ ｐｌａｔｅｎｓｉｓの増殖促進効果の測定は以下のように行った。上記の培養液に対し、オートクレーブで滅菌したB液を無菌的に5%(v/v)、10%(v/v)、20％(v/v)の濃度になるように調整し、全量を200mlとした。乾燥重量、クロロフィル量、フィコビリンタンパク質量、タンパク質量を計測することでＳｐｉｒｕｌｉｎａ ｐｌａｔｅｎｓｉｓの増殖促進効果を決定した。各項目は0日目を最初とし、3日おきに18日目まで測定した。
The proliferation promoting effect of Spirulina platensis was measured as follows. Adjust the amount of B solution sterilized by autoclaving to 5% (v / v), 10% (v / v), 20% (v / v) aseptically to the above culture solution. Was 200 ml. The growth promoting effect of Spirulina platensis was determined by measuring the dry weight, the amount of chlorophyll, the amount of phycobilin protein, and the amount of protein. Each item was measured from
乾燥重量はスピルリナ藻体をグラスフィルター（Whatman社製,GF/C）でろ過し、乾燥させた後計測した。このグラスフィルターはあらかじめオーブンで乾燥させ秤量したものを用いた。スピルリナ藻体を5ml採取し、上記のフィルターでろ過した。蒸留水で2度フィルターを洗浄し、オーブン(Yamato Dry oven DX402)で80℃、4時間乾燥させた。乾燥したフィルターの重量から最初のフィルターの重量を差し引き乾燥重量とした。 The dry weight was measured after filtering Spirulina alga bodies with a glass filter (Whatman, GF / C) and drying. This glass filter was previously dried in an oven and weighed. 5 ml of Spirulina alga was collected and filtered with the above filter. The filter was washed twice with distilled water and dried in an oven (Yamato Dry oven DX402) at 80 ° C. for 4 hours. The weight of the first filter was subtracted from the weight of the dried filter to obtain the dry weight.
クロロフィルの定量にはメタノールで抽出した藻体色素を分光高度計で計測した。方法を以下に示す。
藻体1mlを、遠心分離機（TOMY MX-300）を用い15,000g、15分、温度4℃の条件で遠心分離し、上澄みを取り除いた後、100%メタノールを1ml加え−20℃で凍結させた。色素は藻体の緑色が無くなるまでメタノールを用いて抽出し、再び15,000g、5分、温度4℃の条件で遠心分離を行い、緑色の上澄みを採取した。上澄みに含まれるクロロフィルの吸光度は分光光度計 (BECKMAN, DU530)を用いて計測した。以下に示す数式に従い濃度を決定した。
クロロフィル量（μg/ml）=25.5×A650＋4.0×A665
尚、Aの値は750nmの値を引いたものを用いた。
For the determination of chlorophyll, the algal pigment extracted with methanol was measured with a spectrophotometer. The method is shown below.
Centrifuge 1ml of algal cells using a centrifuge (TOMY MX-300) at 15,000g for 15 minutes at a temperature of 4 ° C. Remove the supernatant, add 1ml of 100% methanol and freeze at -20 ° C. It was. The pigment was extracted with methanol until the green color of the algal bodies disappeared, and centrifuged again at 15,000 g for 5 minutes at a temperature of 4 ° C., and the green supernatant was collected. The absorbance of chlorophyll contained in the supernatant was measured using a spectrophotometer (BECKMAN, DU530). The concentration was determined according to the following formula.
Chlorophyll amount (μg / ml) = 25.5 × A 650 + 4.0 × A 665
A value obtained by subtracting the value of 750 nm was used.
フィコビリンタンパク質の抽出及び測定はAnamikaらの方法に従った。採取した藻体1mlを15,000g、30分、温度4℃で遠心分離した後、培養液成分を洗い流すため蒸留水で2度洗浄をした後、−20℃で凍結した。解凍後1mlのリン酸バッファー溶液（0.1M、pH6.50、0.1M水酸化ナトリウム含有）中で懸濁させ、細胞を完全に破壊するために60秒ソニケーションを行い、−20℃で再び凍結した。次に、十分にフィコビリンタンパク質を抽出させるため暗条件のもと室温で解凍した。15,000g、30分、温度4℃で遠心分離を行い、フィコビリンタンパク質を含んだ透明な上澄みを採取した。
上澄みに含まれるフィコビリンタンパク質の吸光度は分光光度計 (BECKMAN社製, DU530)を用いて計測した。C-PCの測定のために620nm、652nmの波長を計測し、以下に示す数式に従い濃度を決定した。
C-PC(mg/ml)=〔A620−0.474(A652)〕/5.34
タンパク質の定量にはBradford法を用いた。
Extraction and measurement of phycobilin protein followed the method of Anamika et al. 1 ml of the collected alga bodies were centrifuged at 15,000 g for 30 minutes at a temperature of 4 ° C., washed twice with distilled water to wash away the culture solution components, and then frozen at −20 ° C. After thawing, suspend in 1 ml phosphate buffer solution (containing 0.1M, pH6.50, 0.1M sodium hydroxide), perform sonication for 60 seconds to completely destroy the cells, and freeze again at -20 ° C. did. Next, in order to fully extract phycobilin protein, it thawed at room temperature under dark conditions. Centrifugation was performed at 15,000 g for 30 minutes at a temperature of 4 ° C., and a clear supernatant containing phycobilin protein was collected.
The absorbance of the phycobilin protein contained in the supernatant was measured using a spectrophotometer (BECKMAN, DU530). For the measurement of C-PC, wavelengths of 620 nm and 652 nm were measured, and the concentration was determined according to the following formula.
C-PC (mg / ml) = (A 620 -0.474 (A 652 )) / 5.34
The Bradford method was used for protein quantification.
藻体1mlに対し5%(50μl)量の30%水酸化ナトリウム溶液を加え細胞壁を溶解させた。温浴中で煮沸の後、ボルテックスを行った。この操作は2度繰り返した。細胞が懸濁している場合はソニケーションを行い、細胞を完全に破砕した。
タンパク質の吸光度はDU530(BECKMAN)を用い、595nmで計測した。染色液としてBIO-RAD社のプロテインアッセイ溶液を用いた。尚、標準物質として牛血清アルブミンを使用した。上記の標準物質で検量線を作成し、藻体の懸濁液から得られた吸光度を代入しタンパク質の値とした。尚、操作方法はBIO-RAD社のプロテインアッセイ取り扱い説明書に従った。
A 5% (50 μl) amount of 30% sodium hydroxide solution was added to 1 ml of algal cells to dissolve the cell wall. After boiling in a warm bath, vortexing was performed. This operation was repeated twice. When the cells were suspended, sonication was performed to completely disrupt the cells.
The absorbance of the protein was measured at 595 nm using DU530 (BECKMAN). BIO-RAD protein assay solution was used as a staining solution. In addition, bovine serum albumin was used as a standard substance. A calibration curve was created with the above standard substance, and the absorbance obtained from the algal suspension was substituted to obtain the protein value. In addition, the operation method followed the protein assay instruction manual of BIO-RAD.
図１から図4に示すように10%(v/v)又は20%(v/v)量のＢ液をＳｐｉｒｕｌｉｎａ
ｐｌａｔｅｎｓｉｓの培養液に混ぜた結果、乾燥重量やクロロフィル等全ての項目に対し極めて高い増殖効果を得られる事が分かった。
As shown in FIG. 1 to FIG. 4, 10% (v / v) or 20% (v / v) amount of B solution
As a result of mixing with the culture solution of platensis, it was found that an extremely high proliferation effect could be obtained for all items such as dry weight and chlorophyll.
以上のことから、焼酎カスの処理過程で生じるＢ液は、Ｓｐｉｒｕｌｉｎａ ｐｌａ
ｔｅｎｓｉｓの増殖に対し高い促進効果を有するため、焼酎カスの有効利用が可能とな
って、産業の発展に資すること大である。
From the above, the B liquid produced in the process of shochu residue is Spirulina pla.
Since it has a high promoting effect on the growth of tensis, it is possible to effectively use the shochu residue and contribute to the development of the industry.
Claims (1)
Nothing added to the shochu residue, distilled at low pressure and low temperature (65 to 80 degrees C, 200 to 400 torr), and high concentration of BOD and ammonia (corresponding to 30 to 50% of the total amount obtained in this step) First step of extracting fraction (liquid A), adding non-volatile acid to shochu residue and lowering to pH 2, then distilling at low pressure and low temperature (65-80 degrees C, 200-400 torr), acetic acid and propion A second step of extracting (liquid B) a fraction containing an organic acid mainly composed of an acid at a high concentration (corresponding to 50 to 80% of the total amount obtained in this step); Is distilled at a low pressure and low temperature (65 to 80 degrees C, 200 to 400 torr), and extracted as water that can be discharged with a BOD of 20 ppm or less (liquid C). In the method for treating rubbing shochu, Tsu is used as a main component obtained B liquid is flop Spirulina platensis (Spirulina platensis) Obtaining the growth-promoting agent.
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