JPS621712B2 - - Google Patents
Info
- Publication number
- JPS621712B2 JPS621712B2 JP16144580A JP16144580A JPS621712B2 JP S621712 B2 JPS621712 B2 JP S621712B2 JP 16144580 A JP16144580 A JP 16144580A JP 16144580 A JP16144580 A JP 16144580A JP S621712 B2 JPS621712 B2 JP S621712B2
- Authority
- JP
- Japan
- Prior art keywords
- gaseous substance
- oxygen
- gas
- air
- culture
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired
Links
- 239000000126 substance Substances 0.000 claims abstract description 45
- 239000007789 gas Substances 0.000 claims abstract description 26
- 235000015097 nutrients Nutrition 0.000 claims abstract description 15
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 claims abstract description 14
- 239000001301 oxygen Substances 0.000 claims abstract description 14
- 229910052760 oxygen Inorganic materials 0.000 claims abstract description 14
- 238000005192 partition Methods 0.000 claims abstract description 12
- 244000005700 microbiome Species 0.000 claims abstract description 6
- 238000012258 culturing Methods 0.000 claims description 5
- 230000001580 bacterial effect Effects 0.000 claims description 3
- 230000002503 metabolic effect Effects 0.000 claims description 2
- 238000004090 dissolution Methods 0.000 claims 3
- 238000012136 culture method Methods 0.000 claims 1
- CURLTUGMZLYLDI-UHFFFAOYSA-N Carbon dioxide Chemical compound O=C=O CURLTUGMZLYLDI-UHFFFAOYSA-N 0.000 abstract description 18
- 229910002092 carbon dioxide Inorganic materials 0.000 abstract description 9
- 239000001569 carbon dioxide Substances 0.000 abstract description 8
- VNWKTOKETHGBQD-UHFFFAOYSA-N methane Chemical compound C VNWKTOKETHGBQD-UHFFFAOYSA-N 0.000 abstract description 6
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 abstract description 5
- 238000004880 explosion Methods 0.000 abstract description 5
- 239000006227 byproduct Substances 0.000 abstract description 4
- 239000001963 growth medium Substances 0.000 abstract description 3
- 238000011084 recovery Methods 0.000 abstract description 2
- 238000003756 stirring Methods 0.000 abstract description 2
- JCXJVPUVTGWSNB-UHFFFAOYSA-N nitrogen dioxide Inorganic materials O=[N]=O JCXJVPUVTGWSNB-UHFFFAOYSA-N 0.000 abstract 1
- 238000010926 purge Methods 0.000 abstract 1
- 238000000926 separation method Methods 0.000 description 9
- 239000007788 liquid Substances 0.000 description 8
- 238000000034 method Methods 0.000 description 7
- ATUOYWHBWRKTHZ-UHFFFAOYSA-N Propane Chemical compound CCC ATUOYWHBWRKTHZ-UHFFFAOYSA-N 0.000 description 2
- 238000007664 blowing Methods 0.000 description 2
- 238000007796 conventional method Methods 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- 229910052757 nitrogen Inorganic materials 0.000 description 2
- UGFAIRIUMAVXCW-UHFFFAOYSA-N Carbon monoxide Chemical compound [O+]#[C-] UGFAIRIUMAVXCW-UHFFFAOYSA-N 0.000 description 1
- 229910002091 carbon monoxide Inorganic materials 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 239000001257 hydrogen Substances 0.000 description 1
- 229910052739 hydrogen Inorganic materials 0.000 description 1
- 125000004435 hydrogen atom Chemical class [H]* 0.000 description 1
- 239000012528 membrane Substances 0.000 description 1
- 239000000047 product Substances 0.000 description 1
- 239000001294 propane Substances 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 238000001179 sorption measurement Methods 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
- C12M23/00—Constructional details, e.g. recesses, hinges
- C12M23/34—Internal compartments or partitions
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
- C12M27/00—Means for mixing, agitating or circulating fluids in the vessel
- C12M27/18—Flow directing inserts
- C12M27/24—Draft tube
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
- C12M29/00—Means for introduction, extraction or recirculation of materials, e.g. pumps
- C12M29/06—Nozzles; Sprayers; Spargers; Diffusers
- C12M29/08—Air lift
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
- C12M29/00—Means for introduction, extraction or recirculation of materials, e.g. pumps
- C12M29/24—Recirculation of gas
Landscapes
- Health & Medical Sciences (AREA)
- Wood Science & Technology (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Organic Chemistry (AREA)
- Chemical & Material Sciences (AREA)
- Zoology (AREA)
- Biomedical Technology (AREA)
- Sustainable Development (AREA)
- Microbiology (AREA)
- Biochemistry (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Biotechnology (AREA)
- Clinical Laboratory Science (AREA)
- Apparatus Associated With Microorganisms And Enzymes (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
Description
【発明の詳細な説明】
本発明はガス状物質を栄養源として好気性微生
物を培養する方法およびその装置に関する。DETAILED DESCRIPTION OF THE INVENTION The present invention relates to a method and apparatus for culturing aerobic microorganisms using gaseous substances as a nutrient source.
従来、ガス状物質、例えばメタン、プロパン、
プタン、一酸化炭素および水素等を栄養源とし
て、好気性微生物を培養し、培養液中に代謝生産
物もしくは菌体を生成蓄積させることは、種々の
方法で行なわれているが、その１例を第１図のフ
ローシートにより説明する。 Traditionally, gaseous substances such as methane, propane,
There are various methods for culturing aerobic microorganisms using nitrogen, carbon monoxide, hydrogen, etc. as nutrients, and producing and accumulating metabolic products or bacterial cells in the culture solution. will be explained using the flow sheet shown in FIG.
培養槽１内での好気性培養に際し、栄養源であ
るガス状物質７、酸素源である空気１０は各々圧
縮機１４，１５で圧縮され、それぞれライン９，
１１を経て培養槽１底部から供給され、培養槽１
内では栄養源ガス状物質７と酸素源空気１０の混
合ガス系で培養が行なわれる。未利用ガス状物
質、未利用空気、副生二酸化炭素等の混合ガスは
培養槽１の上方部よりライン４を経て排出される
が、この混合ガス中から未利用ガス状物質を気体
分離工程１２で分離、回収し、この回収ガスはラ
イン６を経て混合タンク１３で新鮮なガス状物質
７と混合され、ライン８を経由して培養に循環使
用される。また連続培養の場合、原料液はライン
２から供給され、ライン３から排出される。 During aerobic culture in the culture tank 1, the gaseous substance 7 as a nutrient source and the air 10 as an oxygen source are compressed by compressors 14 and 15, respectively, and are passed through lines 9 and 15, respectively.
11 from the bottom of the culture tank 1.
Culture is carried out in a mixed gas system of a gaseous substance 7 as a nutrient source and air 10 as an oxygen source. A mixed gas such as unused gaseous substances, unused air, and by-product carbon dioxide is discharged from the upper part of the culture tank 1 through a line 4. From this mixed gas, unused gaseous substances are separated into a gas separation step 12. The recovered gas is mixed with fresh gaseous substance 7 in a mixing tank 13 via line 6, and recycled for culture via line 8. Further, in the case of continuous culture, the raw material liquid is supplied from line 2 and discharged from line 3.
上記気体分離工程１２での気体分離法として
は、深冷分離法、膜分離法、吸着分離法等が用い
られるが、回収率はせいぜい70〜80％であり、更
に設備費もかなり高価である。また栄養源のガス
状物質はほとんど可燃性であり、空気との混合ガ
スは爆発のおそれがあり、この危険を回避するた
めに高価な設備費を要していた。 As the gas separation method in the above gas separation step 12, cryogenic separation method, membrane separation method, adsorption separation method, etc. are used, but the recovery rate is at most 70 to 80%, and the equipment cost is also quite expensive. . In addition, most of the gaseous substances that are the nutrient source are flammable, and there is a risk of explosion when mixed with air, requiring expensive equipment costs to avoid this danger.
本発明はガス状物質を栄養源として好気性微生
物を培養するに際し、気体分離工程などに高価な
装置を使用せず、簡単な方法でガス状物質を有効
利用でき、しかも爆発などの危険のない方法およ
び装置を提供するもので、培養槽内部を複数個の
室に区切り、栄養源ガス状物質、酸素源である空
気を別々の室に供給し培養液に溶解させ、未利用
のガス状物質、未利用空気は各々別個に排気して
ガス状物質と空気の混合による爆発を回避し、未
利用ガス状物質および必要に応じ未利用空気を回
収、循環使用すると共に、ガス状物質および空気
を溶解した培養液は充分混合して培養を効率よく
行なうものである。 When culturing aerobic microorganisms using gaseous substances as a nutrient source, the present invention enables effective use of gaseous substances in a simple manner without using expensive equipment for the gas separation process, and without the risk of explosion. The method and apparatus are provided by dividing the inside of a culture tank into multiple chambers, supplying gaseous substances as a nutrient source and air as an oxygen source to separate chambers, dissolving them in the culture solution, and removing unused gaseous substances. , each unused air is evacuated separately to avoid an explosion due to the mixture of gaseous substances and air, and the unused gaseous substances and unused air are collected and recycled as necessary, and the gaseous substances and air are The dissolved culture solution should be thoroughly mixed to carry out the culture efficiently.
本発明方法を第２図のフローシートにより詳細
に説明する。 The method of the present invention will be explained in detail with reference to the flow sheet shown in FIG.
培養槽１は液中ガス吹き込み型で、全体の液混
合を向上させるためドラフトチユーブ付エアリフ
ト槽が好ましく、ドラフトチユーブ１７等で循環
流を生じさせる構造がよい。この槽１の空間部を
少なくとも１つの、垂直面である仕切板１８によ
り分割している。 The culture tank 1 is of a submerged gas blowing type, and is preferably an air lift tank with a draft tube to improve overall liquid mixing, and preferably has a structure in which a draft tube 17 or the like generates a circulating flow. The space of this tank 1 is divided by at least one vertical partition plate 18.
栄養源のガス状物質７を圧縮機１４で圧縮しラ
イン９を経て、ドラフトチユーブの下端部より上
方に、かつドラフトチユーブの外側に供給する。
ガス状物質はドラフトチユーブ１７外側部を空気
と混合することなく上昇し、培養液に一部溶解
し、残存する未利用のガス状物質および副生CO2
はライン４を経てCO2除去装置２１に入つてCO2
を除き、更に蓄積されるCO2をライン１９より取
り出し、残りのガスはライン２０より圧縮機１４
に入つて新鮮なガス状物質７と合流し、培養槽に
循環供給する。 A gaseous substance 7 as a nutrient source is compressed by a compressor 14 and supplied through a line 9 above the lower end of the draft tube and to the outside of the draft tube.
The gaseous substances rise through the outside of the draft tube 17 without being mixed with air, are partially dissolved in the culture medium, and remain as unused gaseous substances and by-product CO2.
passes through line 4 and enters the CO 2 removal device 21 where CO 2
The remaining gas is taken out from the line 19 and the remaining gas is sent to the compressor 14 from the line 20.
The liquid enters the tank, joins with fresh gaseous substance 7, and is circulated and supplied to the culture tank.
一方、空気１０は圧縮機１５で圧縮しライン１
１を経て、ドラフトチユーブの下端部より上方
に、かつドラフトチユーブの外側に供給し、この
空気はドラフトチユーブ外側部をガス状物質と混
合することなく上昇し、その間培養液に一部とり
こまれ、未利用の空気はライン１６より系外に取
り出される。このとき上記ガス状物質と同様、未
利用空気を循環使用してもよい。 On the other hand, air 10 is compressed by compressor 15 and line 1
1, the air is supplied above the lower end of the draft tube and to the outside of the draft tube, and this air rises up the outside of the draft tube without mixing with gaseous substances, during which time it is partially taken up by the culture medium, Unused air is taken out of the system through line 16. At this time, unused air may be circulated and used like the gaseous substance described above.
ここで第３図の培養槽の図から判るように、仕
切板１８は、ドラフトチユーブ１７内では仕切板
１８の下端部はドラフトチユーブ１７の上端部よ
り下側に位置し、培養槽１の空間部分を２室に完
全に分割している。ドラフトチユーブ１７外では
仕切板１８の下端部はガス吹き込み位置より下側
に位置している。ドラフトチユーブ外側でのガス
状物質７と空気１０の混合防止の働きを、ドラフ
トチユーブおよび仕切板１８のドラフトチユーブ
外側部が行つている。したがつてライン４からの
排出ガスはほとんど酸素、窒素を含まないガス状
物質７と副生した二酸化炭素であり、二酸化炭素
除去装置２１を設け、その二酸化炭素を除き、更
に蓄積される二酸化炭素をライン１９よりパージ
する程度で、第１図の従来法に示した気体分離工
程１２を使用しなくてよい。 As can be seen from the diagram of the culture tank in FIG. The part is completely divided into two chambers. Outside the draft tube 17, the lower end of the partition plate 18 is located below the gas blowing position. The draft tube and the partition plate 18 on the outside of the draft tube serve to prevent the gaseous substance 7 and air 10 from mixing outside the draft tube. Therefore, the exhaust gas from the line 4 is a gaseous substance 7 containing almost no oxygen or nitrogen, and by-product carbon dioxide. A carbon dioxide removal device 21 is provided to remove the carbon dioxide and remove the further accumulated carbon dioxide. It is not necessary to use the gas separation step 12 shown in the conventional method of FIG. 1, as long as the gas is purged from the line 19.
第４図ａは１枚の仕切板１８を用いて槽１およ
びドラフトチユーブ１７を仕切板１８で２分割し
た場合の断面図であり、例えばＡの部分に下部よ
りガス状物質７を供給すれば、Ｂの部分に下部よ
り空気１０を供給する。通常、断面積Ａ：Ｂ＝
１：１である。第４図ｂは４分割の場合で、ガス
状物質７の供給部Ａ、空気１０の供給部Ｂを交互
に設定できる。更に第４図Ｃの場合は、ガス状物
質、空気の量が異なつて、例えばガス状物質＜空
気の場合、ガス状物質を狭い水平断面を有するＡ
の部分に供給し、空気１０を広い水平断面を有す
るＢの部分に供給するもので、培養条件により、
ガス空塔速度が一定になるような断面積Ａ，Ｂの
比の設定も可能である。 FIG. 4a is a cross-sectional view when the tank 1 and the draft tube 17 are divided into two parts using one partition plate 18. For example, if the gaseous substance 7 is supplied to the part A from the bottom, , B are supplied with air 10 from the bottom. Usually, cross-sectional area A:B=
The ratio is 1:1. FIG. 4b shows the case of four divisions, and the supply section A for the gaseous substance 7 and the supply section B for the air 10 can be set alternately. Furthermore, in the case of FIG. 4C, the amounts of the gaseous substance and air are different.
, and air 10 is supplied to part B, which has a wide horizontal cross section.Depending on the culture conditions,
It is also possible to set the ratio of cross-sectional areas A and B such that the superficial gas velocity remains constant.
以上、示した構造により培養槽内でガス状物
質、空気の分割供給、分割排出ができるが、一
方、槽内の液は十分混合して菌体とガス状物質、
空気の接触を十分に行なう必要があり、これはド
ラフトチユーブによるエアリフト効果がきいてく
るが、ドラフトチユーブ１７内の仕切板１８の下
端部に第５図ａ，ｂに示すようなリブ２０を設け
て旋回流を起こし、槽内液の混合を均一にした
り、ドラフトチユーブ内に撹拌羽根等を設け下降
流を生じさせて一層の均一化をはかることが好ま
しい。更に培養槽１は多段に多孔板等を設置して
気液接触効果を増大させることができる。 The structure shown above allows gaseous substances and air to be dividedly supplied and discharged in the culture tank, but on the other hand, the liquid in the tank is sufficiently mixed and the bacterial cells and gaseous substances are separated.
It is necessary to ensure sufficient air contact, and this is achieved by the air lift effect of the draft tube, but by providing a rib 20 at the lower end of the partition plate 18 in the draft tube 17 as shown in FIGS. 5a and 5b. It is preferable to create a swirling flow to uniformly mix the liquid in the tank, or to provide a stirring blade or the like in the draft tube to create a downward flow to achieve further uniformity. Furthermore, the culture tank 1 can be equipped with perforated plates or the like in multiple stages to increase the gas-liquid contact effect.
以上のように本発明では気体分離工程を設け
ず、栄養源のガス状物質を簡単に、しかも効率よ
く回収、循環使用できると共に、上記ガス状物質
と空気は別々に供給、排出するので、ガス状物質
と空気との混合による爆発の危険を除くことがで
き、しかも液は全体によく混合され好気性培養を
行うことができるものである。 As described above, the present invention does not require a gas separation process, and gaseous substances as a nutrient source can be easily and efficiently recovered and recycled, and the gaseous substances and air are supplied and discharged separately, so that gaseous substances can be easily and efficiently recovered and recycled. It is possible to eliminate the risk of explosion due to mixing of the liquid with air, and the liquid is well mixed throughout, making it possible to carry out aerobic culture.
第１図および第２図はガス状物質を栄養源とし
た好気性微生物の培養方法のフローシートを示
し、第１図は従来法、第２図は本発明方法に関す
るものであり、第３図は本発明の一実施態様の培
養装置の構造を示す一部切欠図であり、第４図は
仕切板の設置例を示す断面図であり、第５図は仕
切板の下端部にリブを設けた場合を示す図であ
る。
Figures 1 and 2 show flow sheets of a method for culturing aerobic microorganisms using gaseous substances as a nutrient source, with Figure 1 relating to the conventional method, Figure 2 relating to the method of the present invention, and Figure 3 relating to the method of the present invention. is a partially cutaway view showing the structure of a culture device according to an embodiment of the present invention, FIG. 4 is a sectional view showing an example of installing a partition plate, and FIG. 5 is a rib provided at the lower end of the partition plate. FIG.
Claims (1)
養し、代謝生産物および菌体を生産せしめる際
に、単一の培養槽内に少なくとも１つ以上の仕切
板を設けて、栄養源のガス状物質と酸素源とを
別々に培養液に供給、溶解せしめ、未利用のガス
状物質および未利用の酸素源は別々に取り出し、
このうち少なくともガス状物質については回収し
て循環使用すると共に、栄養源の溶解した培養液
と酸素の溶解した培養液とを混合することを特徴
とする培養方法。 ２ ガス状物質供給管およびガス状物質排出管を
有するガス状物質溶解室、仕切板により同ガス状
物質溶解室とは気密に隔離され、かつ含酸素気体
供給管および含酸素気体排出管を有する含酸素気
体溶解室、上記両溶解室と連通し、ガス状物質溶
解培養液と含酸素気体溶解培養液とを混合する混
合培養室、および、上記ガス状物質排出管を上記
ガス状物質供給管に連通させるガス状物質循環管
とから構成される培養装置。[Claims] 1. When culturing aerobic microorganisms using gaseous substances as a nutrient source to produce metabolic products and bacterial cells, at least one or more partition plates are provided in a single culture tank. , a gaseous substance as a nutrient source and an oxygen source are separately supplied and dissolved in the culture solution, and the unused gaseous substance and the unused oxygen source are separately taken out;
A culture method characterized in that at least gaseous substances among these substances are recovered and recycled for use, and a culture solution in which a nutrient source is dissolved and a culture solution in which oxygen is dissolved are mixed. 2. A gaseous substance dissolving chamber having a gaseous substance supply pipe and a gaseous substance discharge pipe, airtightly isolated from the gaseous substance dissolution chamber by a partition plate, and having an oxygen-containing gas supply pipe and an oxygen-containing gas discharge pipe. an oxygen-containing gas dissolution chamber, a mixing culture chamber that communicates with both of the above-mentioned dissolution chambers and mixes the gaseous substance-dissolved culture solution and the oxygen-containing gas-dissolved culture solution, and the gaseous substance discharge pipe connected to the above-mentioned gaseous substance supply pipe. A culture device consisting of a gaseous substance circulation pipe that communicates with the
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP16144580A JPS5786290A (en) | 1980-11-18 | 1980-11-18 | Cultivating method and apparatus |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP16144580A JPS5786290A (en) | 1980-11-18 | 1980-11-18 | Cultivating method and apparatus |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| JPS5786290A JPS5786290A (en) | 1982-05-29 |
| JPS621712B2 true JPS621712B2 (en) | 1987-01-14 |
Family
ID=15735242
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP16144580A Granted JPS5786290A (en) | 1980-11-18 | 1980-11-18 | Cultivating method and apparatus |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPS5786290A (en) |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPWO2010116946A1 (en) * | 2009-04-06 | 2012-10-18 | 株式会社エフティエルインターナショナル | Microalgae culture equipment |
-
1980
- 1980-11-18 JP JP16144580A patent/JPS5786290A/en active Granted
Also Published As
| Publication number | Publication date |
|---|---|
| JPS5786290A (en) | 1982-05-29 |
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