bioseparation in production of lipase using fusarium sp

8/16/2019 Bioseparation in production of lipase using fusarium sp

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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


9/23

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.


10/23

$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


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


1443.363 kg /batch x !&&% H &.&'7' %


12/23

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


13/23

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.


14/23

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


15/23

. ?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


16/23

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.


17/23

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.


18/23

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


19/23

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.


20/23

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#"


21/23

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@ (


22/23

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 .

bioseparation in production of lipase using fusarium sp

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