WO2010089864A1 - 動物プランクトン餌料用セレン含有単細胞微細藻類とそれを使用したセレン含有動物プランクトンの培養方法 - Google Patents
動物プランクトン餌料用セレン含有単細胞微細藻類とそれを使用したセレン含有動物プランクトンの培養方法 Download PDFInfo
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- WO2010089864A1 WO2010089864A1 PCT/JP2009/051896 JP2009051896W WO2010089864A1 WO 2010089864 A1 WO2010089864 A1 WO 2010089864A1 JP 2009051896 W JP2009051896 W JP 2009051896W WO 2010089864 A1 WO2010089864 A1 WO 2010089864A1
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- selenium
- feed
- microalgae
- zooplankton
- rotifer
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- BUGBHKTXTAQXES-UHFFFAOYSA-N Selenium Chemical compound [Se] BUGBHKTXTAQXES-UHFFFAOYSA-N 0.000 title claims abstract description 97
- 229910052711 selenium Inorganic materials 0.000 title claims abstract description 96
- 239000011669 selenium Substances 0.000 title claims abstract description 96
- 238000000034 method Methods 0.000 title claims abstract description 16
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- 241001465754 Metazoa Species 0.000 title abstract description 7
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- 241000700141 Rotifera Species 0.000 claims description 61
- 241000195649 Chlorella <Chlorellales> Species 0.000 claims description 29
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- 238000004519 manufacturing process Methods 0.000 claims description 12
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- MBMBGCFOFBJSGT-KUBAVDMBSA-N all-cis-docosa-4,7,10,13,16,19-hexaenoic acid Chemical compound CC\C=C/C\C=C/C\C=C/C\C=C/C\C=C/C\C=C/CCC(O)=O MBMBGCFOFBJSGT-KUBAVDMBSA-N 0.000 description 2
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- TVBSSDNEJWXWFP-UHFFFAOYSA-N nitric acid perchloric acid Chemical compound O[N+]([O-])=O.OCl(=O)(=O)=O TVBSSDNEJWXWFP-UHFFFAOYSA-N 0.000 description 1
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Images
Classifications
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K20/00—Accessory food factors for animal feeding-stuffs
- A23K20/20—Inorganic substances, e.g. oligoelements
- A23K20/30—Oligoelements
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K61/00—Culture of aquatic animals
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K61/00—Culture of aquatic animals
- A01K61/20—Culture of aquatic animals of zooplankton, e.g. water fleas or Rotatoria
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K50/00—Feeding-stuffs specially adapted for particular animals
- A23K50/80—Feeding-stuffs specially adapted for particular animals for aquatic animals, e.g. fish, crustaceans or molluscs
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A40/00—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production
- Y02A40/80—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production in fisheries management
- Y02A40/81—Aquaculture, e.g. of fish
Definitions
- the present invention relates to a means for preventing selenium deficiency of larvae and juveniles during breeding after hatching in fish seedling production. Specifically, the present invention relates to a zooplankton culture feed used as a feed to be fed to larvae, that is, a selenium-containing unicellular microalgae, and a selenium-containing zooplankton culture method using the selenium-containing unicellular microalgae.
- Non-Patent Document 1 Currently, began to freshwater chlorella essential polyunsaturated fatty acids is enhanced for essential vitamins B 12 and hatched larvae fish growth for growth of rotifers are mainly used. However, rotifers cultured in these are insufficient in terms of whether all the nutritional requirements of hatched larvae are satisfied (see, for example, Non-Patent Document 1).
- Selenium is known to be an essential trace component contained in various enzymes that work in the body of animals.
- glutathione peroxidase containing selenium is important because it acts as a catalyst for glutathione that works to eliminate cellular peroxide and reactive oxygen species.
- the zooplankton co-poda and artemia contain selenium as a trace mineral in the body, the present inventors cannot detect selenium at all in the same zooplankton rotifer or daphnia, or are extremely low. We discovered that selenium deficiency may have occurred in marine larvae during the period when they were fed.
- a uniplanar microalgae for feed comprising selenium in a uniplanar microalgae in a zooplankton diet.
- a single-cell microalgae for feed having a selenium content of 6.0 to 1500 ⁇ g / 100 g per dry alga weight.
- the single-cell microalgae for feeding are freshwater chlorella and Nannochloropsis.
- single-cell microalgae for feed, in which the target zooplankton is an rotifer and daphnia, which is indispensable as an initial feed for fish seedling production.
- FIG. 1 is a diagram showing the effect of selenium-containing raw chlorella and selenium-containing chlorella powder on rotifer culture.
- FIG. 2 is a diagram showing the effect of raw chlorella with different selenium-containing concentrations on the growth of rotifers.
- FIG. 3 is a graph showing the relationship between the selenium content in selenium-containing raw chlorella and the selenium content enhanced by cultured rotifers.
- Typical examples of zooplankton capable of mass culture include rotifers and daphnids.
- Brachionus is cultivated around the world as a suitable live food to be given to larval and juvenile fish immediately after hatching in the production of seedlings for fish farming.
- the body length is 150-250 ⁇ m, and it is in the shape of a pot. It constantly filters the floating microalgae in the water while moving the cilia, and uses it as a nutrient source to live in brackish waters. Therefore, it has the habit of gathering towards the light because it seeks microalgae for photosynthesis.
- Nutritional components include 60% (w / w) protein per dry matter, 10% (w / w) total fat, 15% (w / w) carbohydrates, and 8% (w / w) ash content. It is characterized by high nutritional value. This rotifer can use the raw chlorella which became industrially mass-produced as the feed, and the produced live rotifer is used as a suitable biological feed of hatching larvae.
- Chlorella used in the present invention can be used regardless of species or strains as long as it belongs to the genus Chlorella, such as Bulgaris species, Solokiniana species, and Queselli species.
- Nannochloropsis can also be used regardless of species or strain as long as it belongs to the genus Nannochloropsis, such as Oculata and Salina.
- the medium used for culturing the single cell microalga can be used regardless of the type as long as the medium can grow the single cell microalgae.
- glucose 80 g / L, potassium phosphate 2.0 g / L, magnesium sulfate 2.0 g / L, Fe ⁇ EDTA 4.8 mg / L, trace mineral A 5 ( ⁇ 10) 1.08 ml / L, vitamin A medium of B 12 2.4 mg / L and urea 6.0 g / L can be used.
- Nannochloropsis potassium nitrate 0.5 g / L, sodium hydrogen phosphate 0.1 g / L, Fe.EDTA 4.15 mg / L, trace mineral A 5 ( ⁇ 10) 0.1 ml / L was dissolved in seawater.
- a culture medium can be used (for example, refer nonpatent literature 3).
- sodium selenite Na 2 SeO 3
- sodium selenite is used by adding to a medium for microalgae growth.
- the amount of sodium selenite added to the medium for microalgae is preferably within the range effective for rotifers and larvae without affecting the growth of microalgae, and can be strengthened by adding in the range of 18 to 1800 ⁇ g / L. good. More preferably, sodium selenite may be added in the range of 60 to 600 ⁇ g / L for strengthening.
- microalgae there are no restrictions on the culture conditions for microalgae. For example, when the temperature is in the range of 5 ° C. to 40 ° C., the culture solution is aerated and stirred, and irradiated with light as necessary, and microalgae can be grown by culturing for 0.5 to several days.
- Chlorella with a selenium content of 6.0-1500 ⁇ g / 100 g (per algae dry matter weight) is obtained by inoculating and culturing a seed strain using the above-mentioned medium for growing microalgae containing selenium at an appropriate concentration.
- Nannochloropsis can be obtained. Even more preferably, 20 to 600 ⁇ g / 100 g (per dry alga body weight) of chlorella and Nannochloropsis can be obtained.
- zooplankton is cultured using these selenium-containing raw microalgae as feed.
- zooplankton culture method There are no restrictions on the zooplankton culture method.
- rotifers are inoculated into seawater at a density of 10/500 to 500 / ml, this rotifer culture is kept at 10 ° C. to 35 ° C. and aerated to give microalgae enriched with selenium, starting from one day.
- rotifers can be grown from 300 individuals / ml to 2000 individuals / ml. It is preferable to use living cells rather than dead cells for microalgae.
- microalgae it is also possible to use a combination of microalgae, baker's yeast and other feeds.
- the method for feeding microalgae can be given at the start of the culture, but it is preferable to measure the density of the rotifer every day and give the microalgae in an amount that the rotifer can feed on a daily basis.
- the selenium content of cultured rotifers increases as the selenium content in the microalgae used as a feed increases, and the selenium content in the rotifers can be controlled by controlling the selenium content in the microalgae. is there.
- the selenium content of the rotifer is preferably in the range of 10 to 500 ⁇ g / 100 g (per dry matter weight) from the viewpoint of the effect on larvae and juveniles. Further, considering the production efficiency of rotifer, the range is preferably 20 to 200 ⁇ g / 100 g (per dry matter weight).
- single-cell microalgae containing selenium as a feed during zooplankton culture made it possible to produce zooplankton containing an appropriate amount of selenium.
- the selenium-containing zooplankton produced in this way is used as a feed for larvae and larvae, the survival rate and vitality of larvae and larvae can be increased as compared to the conventional case.
- the growth of zooplankton is also promoted, so that rotifers can be efficiently produced.
- Example 1 Production of selenium-containing rotifer using selenium-containing raw chlorella as a feed
- the selenium-containing raw chlorella was cultured using a 10 L jar fermenter.
- the chlorella strain (Chlorella vulgaris) slant owned by our company was inoculated aseptically into the Sakaguchi flask of dark culture basal medium, and precultured at 28 ° C. After the utilization of glucose, it was transplanted to a 10 L jar fermenter, and selenium-enhanced chlorella was aseptically cultured.
- the culture conditions were medium volume 6 L, temperature 36 ° C., aeration 3 L / min, pH 7.0, stirring speed 300 rpm / min.
- Medium composition is glucose 80 g / L, potassium phosphate 2.0 g / L, magnesium sulfate 2.0 g / L, Fe ⁇ EDTA 4.8 mg / L, trace mineral A 5 ( ⁇ 10) 1.08 ml / L, vitamin B 12 Sodium selenite was added to basal media of 2.4 mg / L and urea 6.0 g / L at concentrations of 5.4 ⁇ g / L, 18 ⁇ g / L, 54 ⁇ g / L, 180 ⁇ g / L and 540 ⁇ g / L, respectively. The culture was started using the sample.
- a culture test was carried out by feeding a total of four types of test feed of powder to a rotifer (S type). As test conditions, 50 L of artificial seawater having a 1.5% (w / w) salt concentration was put in each 100 L panlite water tank, the water temperature was 28 ° C., the aeration was 45 L / min, and the starting rotifer density was 80 individuals / ml.
- the amount of food to be fed was cultured for 1 week on the basis of 0.5 to 0.7 ⁇ g / rotifer individual / day in terms of dry alga mass. The results are shown in FIG.
- the selenium-containing raw chlorella group had a higher growth rate of the rotifer than the conventional method.
- there was a clear difference in the form of feed between the raw state and the powdered state and almost no growth of rotifers was observed in the powdered test plots. This is thought to be due to the fact that the raw chlorella of the feed was freeze-dried, the cells were broken, and nutrient components such as protein were eluted in the rotifer's breeding water, resulting in an oligotrophic state. This was supported by the ongoing contamination of the breeding water in both districts fed with powdered feed.
- the rotifer (S type) culture test was performed using the test foods with different selenium concentrations prepared by the above method (selenium content 0 ⁇ g, 6 ⁇ g, 19 ⁇ g, 58 ⁇ g, 191 ⁇ g, 570 ⁇ g / 100 g per dry weight). did.
- 40 L of artificial seawater having a 1.5% (w / w) salt concentration was put in each 50 L panlite water tank, the water temperature was 28 ° C., the aeration was 45 L / min, and the starting rotifer density was 80 individuals / ml.
- the amount of food to be fed was cultured for 4 days on the basis of 0.5 to 0.7 ⁇ g / rotifer individual / day in terms of dry alga mass.
- the growth of rotifer was excellent when the selenium concentration in the feed was in the range of 19 to 191 ⁇ g / 100 g (per dry alga body weight). This result shows that selenium enrichment of the feed is effective for the growth of rotifer.
- the selenium content of the rotifer was measured after the test, and the result is shown in FIG. From the results of FIG. 3, the selenium content in rotifers cultured with raw chlorella (selenium concentration: below detection limit) produced by the same method as the conventional method was below the detection limit ( ⁇ 5 ⁇ g / 100 g). In contrast, the selenium content in rotifers cultured with selenium-containing raw chlorella (selenium concentration of 6 to 570 ⁇ g / 100 g) increases with an increase in the selenium concentration in the raw chlorella, up to 200 ⁇ g / 100 g (per dry matter). ) Live rotifer was able to be produced.
- Example 2 Production of selenium-containing daphnia using selenium-containing raw Nannochloropsis as a feed
- Selenium-containing Nannochloropsis was cultured using a 30 L flat culture tank. Aseptically inoculate a 100 ml small flat flask containing a light culture basal medium from our company's Nannochloropsis slant, and stir with 80 ml of light culture basal medium, 5% (v / v) CO 2 aerated. Pre-culture was performed for 2 weeks at an illuminance of 2000 lux and a water temperature of 20 ° C.
- the culture conditions were a medium volume of 25 L, 5% CO 2 aeration and stirring at 3 L / min, an illuminance of 2500 lux, and a water temperature of 20 ° C.
- the composition of the medium was selenite in 0.5 g / L potassium nitrate, 0.1 g / L sodium hydrogen phosphate, 4.15 mg / L Fe.EDTA, 0.1 ml / L trace mineral A 5 ( ⁇ 10)
- the culture was started using sodium 50 ⁇ g / L dissolved in diluted seawater (1% w / w salt).
- the product was harvested and washed three times with pure water to obtain 20 g of live Nannochloropsis algae (converted to dry matter) having a selenium content of 480 ⁇ g / 100 g (per dry matter).
- Example 3 Breeding test of flounder larvae using selenium-containing rotifer A flounder (Paralichthys olivacens) larvae breeding test was conducted using selenium-containing rotifers produced using selenium-containing raw chlorella as a feed.
- the breeding test conditions are as follows: 500 L filtered seawater is introduced into each 600 L circular polycarbonate aquarium, water conversion rate is 100% / day, water temperature is 17-20 ° C., aeration is 500 ml / min. 0.05 mm * 1 ) 1.1 ⁇ 10 4 were accommodated.
- Example 3 The test results of Example 3 and Comparative Example 1 are shown in Table 1.
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Abstract
Description
及びそのセレン含有量が、藻体乾燥物重量当たり6.0~1500μg/100gである餌料用単細胞微細藻類。
及びその単細胞微細藻類が、淡水産クロレラ及びナンノクロロプシスである餌料用単細胞微細藻類。
及び対象となる動物プランクトンが、魚類種苗生産の初期餌料として不可欠な、ワムシ類(Rotifer)及びミジンコ類である餌料用単細胞微細藻類。
及びセレン含有単細胞微細藻類を動物プランクトンの餌料として使用し、生産された動物プランクトンのセレン含有量が乾燥物重量当たり10~500μg/100gである動物プランクトンの培養方法。
動物プランクトンの培養方法に制限はない。例えば、ワムシを海水に10個体/mlから500個体/mlの密度で接種し、このワムシ培養液を10℃から35℃に保って通気を行い、セレンを強化した微細藻類を与えて1日から10日程度培養することによって、ワムシを300個体/mlから2000個体/mlの密度まで増殖させることができる。微細藻類は死細胞よりも生きた細胞を用いる方が好ましい。また、微細藻類とパン酵母やその他の餌料を併用して与えることも可能である。微細藻類の給餌方法は、培養開始時に一度に与えることもできるが、毎日ワムシ密度を測定し、ワムシが1日に摂餌できる量の微細藻類を毎日与える方が好ましい。
セレン含有生クロレラを餌料としたセレン含有ワムシの生産
10Lジャーファーメンターを用いて、セレン含有生クロレラの培養を行った。当社保有のクロレラ株(Chlorella vulgaris)スラントから暗培養基礎培地の坂口フラスコに無菌的に植菌して、28℃で前培養を行った。グルコースの資化終了後に10Lジャーファーメンターに植え継ぎ、セレン強化クロレラの無菌培養を行った。培養条件は、培地液量6L、温度36℃、通気3L/分、pH7.0、撹拌速度300rpm/分とした。培地組成は、グルコース80g/L、リン酸カリウム2.0g/L,硫酸マグネシウム2.0g/L、Fe・EDTA4.8mg/L、微量ミネラルA5(×10)1.08ml/L、ビタミンB12 2.4mg/L、尿素6.0g/Lの基礎培地に、亜セレン酸ナトリウムを5.4μg/L、18μg/L、54μg/L、180μg/L、540μg/L、の濃度で各々加えたものを用いて、培養を開始した。増殖中に高度不飽和脂肪酸(ドコサヘキサエン酸60%濃度)1.5g/Lを各々添加し、グルコースの資化終了後に純水で3回水洗を行い、セレン含有量6~570μg/100g(乾燥物重量当たり)の範囲で生クロレラ藻体を各々200g(乾燥物換算)得た。
図3の結果より、従来法と同様の方法で生産された生クロレラ(セレン濃度:検出限界以下)で培養したワムシ中のセレン含量は、検出限界(<5μg/100g)以下であった。それに対して、セレン含有生クロレラ(セレン濃度 6~570μg/100g)で培養したワムシ中のセレン含量は、生クロレラ中のセレン濃度の増加に伴って増加し、最大で200μg/100g(乾燥物当り)の生きたワムシを生産できた。
セレン含有生ナンノクロロプシスを餌料としたセレン含有ミジンコの生産
30L容扁平培養槽を用いて、セレン含有ナンノクロロプシスの培養を行った。当社保有のナンノクロロプシス(Nannochloropsis)のスラントから明培養基礎培地入りの100ml容小型扁平フラスコに無菌的に植菌して、80mlの明培養基礎培地、5%(v/v)CO2通気攪拌、照度2000ルクス、水温20℃で2週間前培養を行った。藻体濃度が2g/L(乾燥物当たり)に達した時点で30L扁平培養槽に植え継ぎ、セレン強化培養を行った。培養条件は、培地液量25L、5%CO2通気撹拌3L/分、照度2500ルクス、水温20℃温度とした。培地組成は、硝酸カリウム0.5g/L、リン酸水素ナトリウム0.1g/L、Fe・EDTA4.15mg/L、微量ミネラルA5(×10)0.1ml/Lの基礎培地に、亜セレン酸ナトリウム50μg/Lを希釈海水(塩分1%w/w)に溶解させたものを用いて、培養を開始した。4週間培養を行った後に収穫して、純水で3回水洗を行い、セレン含量480μg/100g(乾燥物当り)の生きたナンノクロロプシス藻体20g(乾燥物換算)を得た。
セレン含有ワムシを使ったヒラメ仔魚の飼育試験
セレン含有生クロレラを餌料として生産したセレン含有ワムシを用いて、ヒラメ(Paralichthys olivacens)仔魚の飼育試験を実施した。飼育の試験条件は各600L容円型ポリカーボネイト水槽に、500Lのろ過海水を投入して、換水率100%/日、水温17~20℃、通気500ml/分とし、孵化仔魚(体長2.1±0.05mm*1)を1.1×104尾収容した。試験餌料として、セレン含有ワムシ(セレン含量100μg/100g乾燥重量当たり)を用いて飼育を開始し、飼育水中のワムシ密度が6~12個体/mlとなるように1日2回給餌を行い、合計2水槽(a、b)で17日間飼育試験を行った。その結果、試験期間中のヒラメ仔魚の摂餌は旺盛で、順調な成育がみられた。
従来法による生クロレラで培養したワムシを使ったヒラメ仔魚の飼育試験
従来の生クロレラを餌料として生産したワムシ(セレン濃度<5μg/100g:検出限界以下)を用いて、ヒラメ仔魚の飼育試験を実施した。飼育の環境条件及び給餌条件は、上記と同様とし、合計2水槽(c、d)で17日間飼育試験を行った。その結果、実施例3の場合とほぼ同様の成育がみられたが、飼育期間中の斃死個体数は増加した。
*2:網による30秒間空中露出後の生残率(n=50)
表1の各17日間のヒラメ仔魚飼育試験結果より、従来法で生産したワムシ給餌区に比べてセレン含有ワムシ給餌区は、体長に大きな差はみられなかったものの、生残率及び活力において優れていた。これは、ワムシ給餌期のヒラメ孵化仔魚にとってワムシ餌料のセレン強化が、その生残率や活力において大きく影響を与えることを示す。
Claims (5)
- 動物プランクトンの餌料において、単細胞微細藻類にセレンを含有させることを特徴とする餌料用単細胞微細藻類。
- セレン含有量が、藻体乾燥物重量当たり6.0~1500μg/100gであることを特徴とする請求項1記載の餌料用単細胞微細藻類。
- 単細胞微細藻類が、淡水産クロレラ及びナンノクロロプシスであることを特徴とする請求項1記載の餌料用単細胞微細藻類。
- 動物プランクトンが、魚類種苗生産の初期餌料として不可欠な、ワムシ類及びミジンコ類であることを特徴とする請求項1記載の餌料用単細胞微細藻類。
- 請求項1記載のセレン含有単細胞微細藻類を動物プランクトンの餌料として使用し、生産された動物プランクトンのセレン含有量が乾燥物重量当たり10~500μg/100gであることを特徴とする動物プランクトンの培養方法。
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CN110684666A (zh) * | 2019-09-17 | 2020-01-14 | 浙江海洋大学 | 一种含硒桡足类的培养方法 |
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