bioseparation in production of lipase using fusarium sp
TRANSCRIPT
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BIOSEPARATION
a) Identify the types of desired product (Intra, Extra or Whoe !e)" Appy the rues of
thu#$ for the process of $io%separation
What is our desire product&
At least 99.7% of pure Lipase enzyme
Where 'e can et it&
It is produced extracellularly by Fusarium sp. Extracellular is not as complicated as intracellular
product where the product are in mix with medium and the microoranism species.
What are our i#purities&
!. "edia
#onstituent Amount
$ipotassium phosphate !"anesium ulfate &.'
Iron ulfate &.!
Asparaine !.'
(east extract )*a*+, !
-lucose &
/olysorbate &01ween &2 3ml
4heat 5ran 97'.6
$istilled water !L
Autoclae at !&8# for '&min 0p :.'2
1able !.!; #omposition of mycelium medium 0""2 for ! liter.
. Funal mycelia
Rue * Separate the #ost pentifu i#purities first$ownstream processes for extracellular and intracellular product are different where< for the case
intracellular product formation< cell harestin is the first step since the product are formed
inside the cell while the outside of the cells are the most plentiful impurities. 5ut for case of
lipase enzyme produced extracellularly by fusarium sp.< the lipase enzymes are produced outside
of the cell. o< this rule is nelected.
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Rue +* or extraceuar product for#ation, rue of thu#$ starts fro# second eneric
heuristic" Re#o-e the easiest to re#o-ed first
Fusarium sp. is the easiest to remoe first. Leain the funal to later step and remoin them
will be waste of treatin unwanted mass which can lead to increase in operatin cost. =emoinmycelia of Fusarium sp. is more difficult than rowin Fusarium sp. since it>s a funus that
deelops in its eetatie form< eneratin hyphae. In areement with the second eneric
heuristic< re#o-e the easiest to re#o-e i#purities first < fusarium sp. mycelia remoal is the
first step of downstream processin of extracellular products. 1his step can be accomplished by
usin rotary acuum filtration since it>s the most suitable for mycelia separation.
Rue .* !hoose those processes that 'i expoit the differences in the physicoche#ica
properties of the product and i#purities in the #ost efficient #annerIn laboratory studies< olid tate Fermentation are enerally carried out in Erlenmeyer flas?s<
bea?ers< petri dishes< roux bottles< @ars and lass tubes. For this laboratory scale fermentation< the
enzyme and solid fermented matter can be easily separated based on the different in physical
state. 1he fermented matter is in solid state and the enzyme is in moderately iscous liuid state
on the surface on the fermented matter. ence the separation is @ust as simple as scoopin out the
enzyme by spoon.In industrial separation process< scoopin each tray are inefficient. ence other basic of
separation are need to be considered for the separation. 5ased on the chemical properties< lipase
enzyme has reatly different solubility in Ammonium ulfate compared to the fermented matter
thus solubility would be a ood basic of separation to separate lipase enzyme from solid
fermented matter. 1he chosen separation process that can separate lipase from fermented broth
based on solubility is precipitation.
/recipitation< which is the process of comin out of solution as a solid< is an important method in
the isolation of enzymes and protein that usually comes early in the purification process. 1he
primary adantaes of precipitation are that it is relatiely inexpensie< can be carried out with
simple euipment< can be done continuously< and leads to a form of the enzyme or protein that is
often stable in lonBterm storae. 1he oal of precipitation is often concentration to reduce
olume< althouh sinificant purification can sometimes be achieed.alt 0Ammonium sulfate2 precipitation is used to recoer lipase enzyme from the fermented
matter. alt concentration plays a role in the rate of reaction. At low concentrations< the presence
of salt stabilizes the arious chared roups on a protein molecule< thus attractin lipase enzyme
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into the solution and enhancin the solubility of lipase enzyme. 1he solubility of lipase enzyme
depends on salt concentration in the solution. oweer< as the salt concentration is increased< a
point of maximum lipase enzyme solubility is usually reached. Further increase in the salt
concentration implies that there is less and less water to solubilize lipase enzyme. Finally< lipase
enzyme starts to precipitate when there are not sufficient water molecules to interact with lipase
enzyme molecules. 1his phenomenon of lipase enzyme precipitation in the presence of excess
salt is ?nown as saltinBout. 1he precipitate then is dissoled in trsB#l buffer.
Rue /* 0a1e the #ost difficut and expensi-e separations ast
#hromatoraphy is typically done later in a process in areement with the third eneric heuristic
#a1e the #ost difficut and expensi-e separations ast. 4ith the preious separation steps< a
lare fraction of contaminants are remoed< which reduces the olume of material that needs to
be treated further. In fact< a '&B!&& fold olumetric reduction is uite common for hih alue
bioloical products< resultin in a protein content of !B'% w) in the feed stream to
chromatoraphic units.
+ne type of molecule called tween & miht not be completely remoed in precipitation step.
ence unnecessary proteins and 1ween & were remoed by CBsepharose ion exchane
chromatoraphy. 1he reason why 1ween & must be remoed first before final purification was
remoed was that 1ween & affected the next /henylBsepharose ion exchane chromatoraphy
because it decreased the hydrophobic nature of proteins. As final purification< the lipase was
further purified by /henylBsepharose ion exchane chromatoraphy.
Rue 2* Seect and se3uence processes that use different separation dri-in forces"
1he downstream process was seuenced in bloc? diaram below was based on the eneralized
bloc? diaram of downstream processin from 5ioseparation ceince and Enineerin 5oo?.
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$iaram; bloc? diaram for downstream process of lipase enzyme production based on the
eneralized bloc? diaram of downstream processin from 5ioseparation ceince and
Enineerin 5oo?.
Rotary vacuumfltration
Fungal mycelia
removal
Precipitation usingOrganic solvent
Product
Q-Sepharose on-
!hromatography to remove
Phenyl-Sepharose on-
!hromatography as fnal
'ehydration or
Solvent removal y
dr in
Final Purifcation
ris-*!l
Preci itate
Solution
'ehydration
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$)Industria Appication
Appications of ipases
Lipases are widely used in the processin of fats and oils< deterents and dereasin formulation<
food processin< the synthesis of fine chemicals and pharmaceuticals< paper manufacture< and
production of cosmetics< and pharmaceuticals. Lipase can be used to accelerate the deradationof fatty waste and polyurethane. "ost of the industrial microbial lipases are deried from funi
and bacteria.
Industry that
uses ipase
4escription
$eterent
industry
Lipases are added to deterents such as household and industrial laundry and
in household dishwashers< where their function is in the remoal of fatty
residues and cleanin cloed drains. 1he cleanin power of lipase deterents
increases mar?edly.
Enzymes can reduce the enironmental load of deterent products as the
chemicals used in conentional deterents are reducedD they are
bioderadable< nonBtoxic and leae no harmful residues. 5esides lipases<
other enzymes are widely used in household cleanin products and in
launderin.$ecompose fatty material. Lipase is capable of remoin fatty stains such as
fats< butter< salad oil< sauces and the touh stains on collars and cuffs.
Food industry Fats and oils are important constituents of foods. 1he nutritional and sensory
alue and the physical properties of a trilyceride are reatly influenced by
factors such as the position of the fatty acid in the lycerol bac?bone< the
chain lenth of the fatty acid< and its deree of unsaturation. Lipases allow us
to modify the properties of lipids by alterin the location of fatty acid chains
in the lyceride and replacin one or more of the fatty acids with new ones.
1his way< a relatiely inexpensie and less desirable lipid can be modified to
a hiher alue fat. #ocoa butter< a hihBalue fat< contains palmitic and stearic
acids and has a meltin point of approximately ,7 °
#. "eltin of cocoa
butter in the mouth produces a desirable coolin sensation in products such as
chocolate. LipaseBbased technoloy inolin mixed hydrolysis and synthesis
reactions is used commercially to uprade some of the less desirable fats to
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cocoa butter substitutes.
/ulp and paper
industry
/itch control is an important aspect in pulp and paper manufacture< and the
first example where microbial biotechnoloy proided successful solutions in
this industrial sector. 1rilycerides cause deposits in softwood mechanical pulpin< and both microbial and enzymatic products hae been
commercialized to be applied on wood and pulp< respectiely. 1he former are
based on colorless strains of sapstain funi. 1he latter are improed lipases<
includin thermostable ariants from directed eolution. 1hese enzymes are
amon the addities of choice in pulpin of hihBresinBcontent softwoods.
+ranic ynthesis 1he use of enzymes for oranic synthesis has become an interestin area for
oranic and bioBoranic chemists. ince many enzymes hae beendemonstrated to possess actiity aainst nonBnatural substrates in oranic
media they hae become widely used to carry out synthetic transformations.
ydrolases are the most freuently used enzymes due to their broad substrate
spectrum and considerable stability. Additionally< many of them are
commercially aailable and they wor? under mild reaction conditions and
without the necessity for cofactors. Amon the hydrolases< lipases are
considered the most popular and useful enzymes for asymmetric synthesis.
Applications for lipases include ?inetic resolution of racemic alcohols< acids<
ester or amines as well as the desymmetrization of prochiral compounds.
1hey are alsosuccesfully employed in reioselectie esterification or
transesterification of polyfunctional compounds< for instance in the
chemoenzymatic synthesis of nucleoside deriaties. =ecently< nonB
conentional processes< such as aldol reactions or "icheal addition hae been
archied usin lipases
5ioconersion inaueous media
Enzymes in oranic media without a free aueous phase are ?nown to displayuseful unusual properties< and this has firmly established nonBaueous
enzyme systems for synthesis and bioBtransformations. Lipases hae been
widely inestiated for arious nonBaueous bioBtransformations.
resolution of tereoBselectiity of lipases has been used to resole arious racemic oranic
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racemic acids and
alcohols
acid mixtures in immiscible biphasic systems. =acemic alcohols can also be
resoled into enantiomerically pure forms by lipaseBcatalyzed transB
esterification.
Ester synthesis Lipases hae been successfully used as catalyst for synthesis of esters. 1heesters produced from shortBchain fatty acids hae applications as flaorin
aents in food industry. "ethyl and ethyl esters of lonBchain acids hae been
used to enrich diesel fuels.
+leo chemical
industry
se of lipases in oleochemical processin saes enery and minimizes
thermal deradation durin alcoholysis< acidolysis< hydrolysis< and
lycerolysis. Althouh lipases are desined by nature for the hydrolytic
cleaae of the ester bonds of triacyllycerol< lipases can catalyze the reersereaction 0ester synthesis2 in a lowwater enironment. ydrolysis and
esterification can occur simultaneously in a process ?nown as
interesterification. $ependin on the substrates< lipases can catalyze
acidolysis 0where an acyl moiety is displaced between an acyl lycerol and a
carboxylic acid2< alcoholysis 0where an acyl moiety is displaced between an
acyl lycerol and an alcohol2< and transesterification 0where two acyl moieties
are exchaned between two acyllycerols2.
1able !.; application of lipase enzyme
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c) Propose a RIPP sche#e for desired products
The RIPP sche#e Stae O$5ecti-es Typica unit 6nit Operation used
=ecoery
0separation of
insoluble2
=emoe cells debris )other particulate.=educe olume
=otary acuum Filter
Isolation of product
=emoe material hae properties widely
different from those desired in product.
=educe olume
/recipitation
/urification =emoe remainin impurities which typically
similar to desired product in chemical
functionality G physical properties.
Ion Exchane #hromatoraphy<
/olishin =emoe liuid. $ryin<
#onsideration that need to be ta?en durin deelopin a bioseparation process;
!. The nature of startin #ateria" 1he startin material is wheat bran. Its oriinal state is
in the solid state. 4heat bran is rich in carbohydrate which are ery suitable as substrate
for fusarium sp. additional nutrients are added for a better rowth of fusarium sp.
. The initia ocation of the taret product* product are formed at extracellular
,. The -ou#e or fo'%rate of the startin #ateria" 1he olume of startin material is
'&&&m,)l. 1he flow rate of startin material is let to flow until maximum allowable
olume for tray bioreactor is reached.3. The reati-e a$undance of the product in the startin #ateria < lipase enzyme is
absence in the startin material. 1he relatie abundance of the product will start to
increase in the fermentation trays when the medium is inoculated.
'. The suscepti$iity to deradation e"" its p7 sta$iity, sensiti-ity to hih shear rates
or exposure to oranic so-ents. Lipase derades at 7&o#< p stability at p6.6. ince
funus suitable enironment is solid state< stirrin no stirrer or impeller are installed.
:. The desired physica for# of the fina product. Final product is desired to be in powdered form.
7. The 3uaity re3uire#ents. 99% pure lipase enzyme.
8" Process costin and econo#ics" economic
A =I// scheme is commonly used in bioseparation. 1his stratey inoles use of low resolution
techniues first for recoery and isolation followed by hih resolution techniues for purification
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and polishin. 1he hihBthrouhput< lowBresolution techniues are first used to sinificantly
reduce the olume and oerall concentration of the material bein processed. 1he partially
purified products are then further processed by hihBresolution lowBthrouhput techniues to
obtain pure and polished finished products.
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$iaram ,.; /rocess Flow diaram based on =I//
=otary acuum filtration which remoes the funal mycelia as the first step in downstream is the
recoery process. /recipitation usin oranic solent 0ammonium sulfate2 is product isolation
process. 1hese recoery and isolation process are hih throuhput< low resolution techniues.
/urification by chromatoraphy and polishin by dryin are low throuhput< hih resolution
techniue.
ray ioreactor
Fungal myceliaremoval y Rotaryvacuum fltrationRecover
Precipitation yOrganic solvent
*igh
throughput+ lo" solation
Q-Sepharose ion-
!hromatography to remove
PurifcatioPhenyl-Sepharose ion-
!hromatography as fnal
,o"
throughput+
'ehydration orSolvent removal y
dr in
Polishing
,ipase en&yme in po"der %orm are paced
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c) !acuate the reco-ery (e" concentrations, purity and 9 or efficiency) for each of the
unit operation in $io%separation process"
Rotary itration :acuu#
#rude lipase mass flow rate ; :.6?)batch
1otal mass flow rate ; ::,.6' ?)batch1otal olume flow rate ; ::97.&33L)batch
#oncentration of lipase enzyme H6.82kg/batch
6697.044 L /batch H !.& x !&B, ?)L
/urity of lipase enzyme H6.82kg /batch
6632.285 kg /batch x !&&% H &.!&,%
Precipitation
#rude lipase mass flow rate ; :.36: ?)batch1otal mass flow rate ; :&.33, ?)batch
1otal olume flow rate ; !39:.&'9L)batch
#oncentration of crude lipase enzyme H6.486 kg/batch
1496.059 L/batch H 3.,,' x !&B, ?)L
/urity of lipase enzyme H6.486kg /batch
1496.059 kg /batch x !&&% H &.3,,'%
;%sepharose !hro#atoraphy
#rude lipase mass flow rate ; 3.!'!?)batch
1otal mass flow rate ; !33:.:63?)batch
1otal olume flow rate ; 9:7.76!L)batch
#oncentration of lipase enzyme H4.151 kg/batch
967.781 L/batch H 3.9 x !&B, ?)L
/urity of lipase enzyme H4.151 kg/batch
1446.684 kg/batch x !&&% H &., %
Pheny%Sepharose !hro#atoraphy
/ure lipase mass flow rate ; &.6,&?)batch
1otal mass flow rate ; !33,.,:,?)batch
1otal olume flow rate ; 9'3.3:L)batch
#oncentration of lipase enzymeH0.830 kg/batch
954.426 L /batch H 6.:9: x !&B3 ?)L
/urity of lipase enzyme H0.830kg /batch
1443.363 kg /batch x !&&% H &.&'7' %
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4ryin
/ure lipase mass flow rate ; &.::3?)batch
1otal mass flow rate ; &.::3?)batch1otal olume flow rate ; &.:79L)batch
#oncentration of lipase enzymeH
0.664 kg /batch
0.679 L/batch H &.96?)L
/urity of lipase enzyme H0.664 kg /batch
0.664 kg /batch x !&&% H !&&%
Purity su##ary ta$e
Rotary itration
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d) 4esin one unit operation ony in the do'nstrea# process (e" utrafitration,
sedi#entation, centrifuation, chro#atoraphy, etc)"
Rotary :acuu# itration (R:)
!. $ischare $esin. $rum $esin
,. pecial ystem $esin
3. Filter #loth) eptum $esin'. 5asic =F +peration
4ischare 4esin
1he fie basic dischare types are;
!. craper
. Endless 5elt,. trin
3. =oll2" Precoat
Each is desined to be able to dischare specific types of formed ca?e solids. In essence< these
fie mechanisms enable the rotary acuum filter to efficiently handle mechanism such as filter
solidBliuid slurry and dischare the formed solids as a complete spectrum of process slurries.
ince media formulation for fermentation are desined as 6&% moisture for a ood rowth rate
of fusarium sp.< the fermentation broth are low solid concentration slurry. 1he problem faced is
durin rotary acuum filtration. Accordin to au< -. 0!9992< precoat dischare is used if slurry
with ery low solid concentration slurry is used that resulted in difficult ca?e formation or if the
slurry is difficult to filter to produce ca?e formation. ence< filter with precoat dischare are
applied since it is the most suitable this case.
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4ru# 4esin
Any =F utilizin a scraper< endless belt< strin or roll dischare must hae a drum with 0!2
filtrate pipes and a 02 alebody with bride bloc?s. A filter with a precoat dischare can use 0!2
a drum with filtrate pipes< 02 a drum with a alebody< 0,2 a aleless drum or 032 a drum
without filtrate pipes. For this reason< precoat dischare filters hae a wide array of desins<
specialty features and aryin reuirements for successful operation.
For all the dischare desin< alebody is a reuirement< accept for precoat dischare. alebody
is a deice which controls the radial position of application for form and dry zone acuum
blowbac? pressure if reuired< and entin to the arious surfaces of the drum as the drum
rotates throuh its cycle. 1he alebody is the connection between the filter which is at the drum
and the acuum system typically the acuum receier. ince the drum don>t hae ent and bride
bloc?s< alebody are not installed for a reater performance. /recoat specific drum desins
cannot control the acuum leel at arious radial positions on the drumD the entire drum is at the
same acuum leel throuhout the entire drum cycle drum reolutionJ All liuid and air are
contained within the filtrate pipes.
$iaram ,.,; aleless drum schematic
Filtrate pipe *o alebody
*o radial position control of acuum
Specia Syste# 4esins
1here are ' types of common system desin for =F
!. #a?e 4ash
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. ?noc1%out%recei-er,. 1iltin alt
3. ydraulic Aitator
/recoat dischare applications are typically hih foam enerators. ince foam does not easily
separate out from the air or liuid stream comin out of the filter drum< there is a hih tendency
for the foam to be swept throuh the acuum receier< into the acuum pump and out of the
filtration system with the acuum pump seal water. 1his can cause enironmental problems<
acuum pump operation problems and a loss of filtered product. 5y addin a second receier
with a diameter sufficient to reduce the air flow elocity to ! ft)sec 0or less2< most foam can be
dropped out of the air stream. Foam carryBoer can also be eliminated by reducin the filter
operatin acuum leel. oweer< this will reduce the filter throuhput and increase operatin
costs< especially with a precoat dischare filter. ence amon the other systems< Kno?B+utB
=eceier is the most suitable for hiher efficiency.
acuum =eceier 1he purpose of the acuum receier is to 0!2 separate the two phase mixture comin out of the
filter< the air and liuid 0filtrate2. If foam is present< the receier must also be capable of
preentin carry oer of foam to the acuum pump. 1he essel diameter is the critical dimension
for effectin the separation of the two phasesD essel heiht is to accommodate sures in flow.
acuum /ump #apacity
For the precoatin mode< the pump must delier at least .' to ,.' #F" per suare foot of filter
area. $urin the process mode< .& to ,.& #F" per suare foot is satisfactory. It is usually
satisfactory to employ a sinle acuum pump for these small filters. /umps should be capable of
achiein 6 acuum and sized for the reuired #F" capacity at & operatin leel.
Filter .id/ae-up
Knoc?Bout or
secondary
acuumreceier
0acuum
llustrated1Precoat
PrecoatSlurry
Rotary0acuum
Filter
/rimary or
main acuumreceier
$iaram
,.3;Knoc?Bout
receier desi nFiltrate
!hec o
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Filtrate /ump #apacity
"ost precoat filter applications will hae a process rate of filtration considerably lower than that
of the precoatin mode.
Filteraid
/recoat lurry "ix
5asic rule for desin precoat slurry mix;
!2 1he mix tan? must be lare enouh to hold the entire chare of filteraid for a full precoat
ca?e at an appropriate slurry concentration 0a satisfactory desin for small filters2D or
2 1he reuired amount of filteraid can be added so as to maintain the desired slurry
concentration durin a ,& minute time span.
,2 *ote that the desired slurry concentration 0wt;wt basis2 is typically '% B 6%. 1he %
concentration should bei2 #onstant throuhout the precoatin mode or
ii2 $ecrease uniformly to Mzero< as with recirculation systems.
ince the basis from the bioreactor is in reater amount< bier filter is desined and the filteraid
is added to maintain the desired slurry durin a ,& minute span. 1he amount of filteraid chose to
add is '% from the slurry concentration and the concentration are set to be constant throuhout
the precipitation precoatin.
"a?eBup 4ater =ecirculationIf a recirculation line is used< ery little ma?eBup water will be needed durin precoatin.
iter !oth9Septu# 4esin
/recoat dischare filter has different reuirements than other dischare type because the septum
is not the filter mediumD the precoat ca?e is the filter medium.
Filteraids is a roup of inert materials that can be used in filtration pretreatment. 1here are two
ob@ecties related to the addition of filteraids. +ne is to form a layer of second medium which
protects the basic medium of the system. 1his is commonly referred to as Mprecoat. 1he second
ob@ectie of filteraids is to improe the flow rate by decreasin ca?e compressibility and
increasin ca?e permeability. 1his type of usae is termed as Madmix or Nbody feedN. Filtration
without filter aid< with precoat< and with precoat and body feed is shown in Fi. ! 0EaleB/itcher
"inerals< Inc.< !97&2.
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a2Filtration without filteraid b2Filtration with filteraid c2 Filtration with filteraid and admix
$iaram ,.'; Filteraid1he common filter aids are diatomaceous earth 0$E2< perlite< cellulose and others. $E is the
s?eleton of ancient diatoms. 1hey are mined from ancient seabed< processed< and classified to
ma?e different rade of filter aids. 1here are different rades of commercial $E. A finer rade
may be employed to increase the clarity of filtrate. 1he smaller the filter aid particle size< the
smaller the process particles can be remoed. oweer< the filtration rate is lower. 1here is
always a balance between initial filtrate clarity and filtration rate.
$iatomaceous earthince the filteraid is the actual filterin medium< careful attention must be paid to the sinle most
important selection criterion that is process solids penetration. For effectie performance< any
filteraid must limit the deree of solids penetration into the precoat ca?e to &.&& O &.&&'.
-reater penetration reuires too hih of a ?nife cut to remoe the Mspent filteraid resultin in
hih filteraid and disposal costs. #onersely< if the filter aid is too Mtiht< for example< too fine<
solids penetration will be minimized< but flow rate will also be forfeited. sin too tiht of a
filteraid rade not only forfeits aailable flow 0filtration2 rates and reduces filteraid efficiency< it
may not yield any improed filtrate clarity compared to an optimum rade 0in this case< more
open2. In li?e manner< there may not be a deradation of filtrate clarity if the filter aid rade is
too open< but excessie uantities of filteraid would be reuired for the same output 0flow rate2
compared to a tihter 0optimum2 rade.
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Knife cut analysis must always be based on ?nife adance rate per drum reolution. "ost precoat
dischare filters hae ?nife adance dries which are independent of the drum drie. 1his system
desin ma?es it necessary to ad@ust the ?nife adance rate wheneer the drum speed is chaned
0assumin that the oriinal ?nife cut was an optimum one2. If the drum speed is reduced< the
optimum cut will chane to an excessie cut. If the drum speed is increased< the optimum cut
will chane to an insufficient cut 0without a ?nife adance rate chane< the ?nife will adance at
a constant rate per time period< not per drum reolution2.
$iaram; Knife #ut analysis
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Basic R: Operation
at leel and drum speed are the two basic operatin parameters for any rotary acuum drum
filter. 1hese parameters are ad@usted dependently to each other to optimize the filtration
performance.
at Leelat leel determines the proportion of the filter cycle< such as one drum reolution< dedicated to
ca?e formation and ca?e dryin. In the absence of any other contradictin factors< at leel
should be ad@usted to maximum hiher at leel is eual to reater filtration. 1he two basic
reasons for reducin the operatin at leel are;
P ard to filter slurries which form thin< elatinous or slimy ca?esD orP lurries with ery hih suspended solids content which form ery thic? ca?es.
ummary of +peratin at Leel #ycles
ih at Leel
P "aximum filtration timeP "aximum solids formation per cycle
P "aximum ca?e thic?nessP "aximum ca?e moisture content
P ihest filter output
Low at Leel
P maximum ca?e dryin)washin time
P minimum solids formation per cycleP minimum ca?e thic?ness
P minimum ca?e moisture content
P lowest filter output
$iaram; $rum of =otary acuum $rum Filter As the drum rotate throuh the feed in the filter tan?< acuum is applied to dewater ca?e pic?ed
on the media. acuum cutoff occurs @ust prior to the ca?e dischare point.
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From the article of Enhancin the /erformance of =otary acuum $rum Filter 01. ia?umar<
&!!2< with the oerflow weir set to a maximum the Napparent submerenceN is normally ,,B
,'% so the slurry leels between &3.&& and &6.&& hrs. +nce a sector enters submerence acuum
is applied and a ca?e starts to form up to a point where the sector emeres from the slurry. 1he
portion of the cycle aailable for formation is the Neffectie submerenceN and its duration
depends on the number of sectors< the slurry leel in the tan?.
ince for hiher performance< hihest of possi$e -at e-e that is /2= is desined.
$rum speed
4ith a precoat dischare filter< hiher drum speeds also means lower filteraid efficiencies
0hence< hiher production costs2. $rum speed and at leel are usually ad@usted dependently in
order to optimize filter performance. 1he drum speed is let to be #inute9 re-oution of dru#"
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1o calculate the area of drum submered
=ate of cell broth H &&&L)hacuum /ressureH 7&?/a
#ycle time H:&s
#a?e formation time H !'siscosity of broth H .&cp
#a?e solid 0dry basis2 per olumeH !&)pecific ca?e resistance 9 x !&!& cm)
4e can use the nitrated form of the filtration euation< with = m H&
t)0)A2 H0QoRS)Tp20)A2
4e sole forA to obtainAHQoRSc
)Tpt
In applyin this euation it is helpful to focus on the area of the drum that is submered< which is
where the ca?e is bein formed and where filtrate is bein obtained. 1hus< A is the area of that
part of the drum that is submered. 4e can calculate the olume of filtrate that needs to becollected durin the ca?e formation time of !'s.
!'sH&&&L)h x !'s x !h),:&&s H6.,,L
4e use this olume of filtrate with time t H!'s in the euation for A to obtain
A H&.&)cm.s x 09 x !&!&cm)2 x !&)L x 06.,,L2 x 0cm.s?/a) !&3 2 x !&,cm,)L x0m)!&cm23
x 7&?/a x !'s
H&.'9'm3
A H &.77!m,
Area A> of the entire rotary acuum filter< can be calculated from the ca?e formation time and the
total cycle time as
A@ (
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e) 4iscussion
5ioseperation in this pro@ect is to recoer< isolate< purify and polish the lipase enzyme in the
downstream process to obtain hihest percentae of purity. First of all the content from the
fermentation broth is identified first. 5efore desinin the downstream process< the ' thumb
rules of bioseparation is well understood.' rule of thumbs;
=ule !; eparate the most plentiful impurities first
=ule ; For extracellular product formation< rule of thumb starts from second eneric heuristic.
=emoe the easiest to remoed first
=ule ,; "a?e the most difficult and expensie separations last=ule 3; #hoose those processes that will exploit the differences in the physicochemical
properties of the product and impurities in the most efficient manner
=ule '; elect and seuence processes that use different separation driin forces.
5y followin all the thumb rules aboe< appropriate euipment are selected to desin a hih
performance downstream process with lowest cost as possible. 1he entire important unit
operations chose is in correct seuenced by referrin to eneralize bloc? diaram in the rule of
thumbs.1hen< the by utilizin =I// 0=ecoery< Isolation< /urification< /olishin2 scheme< the seuence is
then arraned in more accordin order. =ecoery is the process is remoin cellsD the Isolation is
the process of remoin material has properties widely different from those desired in product.
/urification is remoin remainin impurities which typically similar to desired product in
chemical functionality G physical properties. /olishin is process of remoin liuid and
conert product to crystallized form. For recoery< rotary acuum drum filter< for isolation<
precipitation< for purification< ion exchane chromatoraphy and for polishin< is dryin.For the desin a unit operation< rotary acuum filter0=F2 is chose. $ischare desin< drum
desin< special system desin< filter cloth) septum desin basic =F operation is considered in
the desin. 1he chosen dischare desin is precoat dischare since it is the most suitable for hih
moisture 06&%2 solid state fermentation. 1he chosen drum desin is aleless drum schematic
which hae filtrate pipe no ale body and no radial position control of acuum. aleless drum
schematic is the more suitable for precoat dischare type. #hosen special system desin is
?noc?Bout receier. 1his system desin is desined especially for the drum uses precoat. /recoat
dischare applications are typically hih foam enerators. ince foam does not easily separate
out from the air or liuid stream comin out of the filter drum< there is a hih tendency for the
foam to be swept throuh the acuum receier< into the acuum pump and out of the filtration
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system with the acuum pump seal water. /recoat dischare filter has different reuirements than
other dischare type because the septum is not the filter mediumD the precoat ca?e is the filter
medium. 1he chosen filter aid is diatomaceous earth. In thhe basic =F operation< the important
parameters are at leel and drum speed. 1he reater the at leel the reater the filtration.
ence the hihest possible at leel is choosed that is ,'%. For drum speed< increase in drum
speed lower the filteraid efficiency. ence the drum speed is let to ! min per reolution of drum.
1he area of drum submeres calculated for !m $iameter and ! m lon is ,.&6m .