Separation of proteins by charged ultrafiltration membranes

Desalination,70 (1988) 191-205 Elsevier Science Publishers B.V., Amsterdam -

SEPARATION

S. NAKAO.

OF

PROTEINS

H. OSADA,

BY CHARGED

H. KURATA,

Department of Chemical University of Tokyo 7-3-l Hongo, Bunkyo-ku,

ULTRAFILTRATION

T. TSURU

Engineering, Tokyo

191 Printed in The Netherlands

and

S. KIMURA

Faculty

113,

MEMBRANES

of

Engineering,

Japan

SUWARY The separation of a protein mixture by charged ultrafiltration membranes was studied. A negatively charged polymer was obtained by sulfonation of polyand a positively charged polymer was synthesized by chloromethylation sulfone, of polysulfone and then by quaternization of the amino group. Then, the negatively and positively charged ultrafiltration membranes were cast from solutions of charged polymer/NMP(or DMF)/lithium nitrate. The molecular weight cut-off of the membranes were controlled by the changing casting conditions. Single protein solutions were ultrafiltrated at the isoelectric point and at another pH level by the use of charged membranes. At the isoelectric point, rejection of the protein was low, while it was high at the pH level which gave the protein the same sign of charge as that of the membrane. A protein mixture of myoglobin and cytochrome C was separated by the charged ultrafiltration membranes at the isoelectric point of one of the proteins. At the isoelectric point of cytochrome C, myoglobin has a negative charge. Thus myoglobin was rejected with a rejection of about 80% by the negatively charged membrane. At the same time, cytochrome C permeated completely through the membrane. Conversely, at the isoelectric point of myoglobin, cytochrome C has a positive charge and thus it was rejected with a rejection of about 20% by the positively charged membrane. The rejection of myoglobin here was almost zero.

Introduction Many used

kinds

of

ultrafiltration

commercially

in

separate

solutes

mixtures

of macromolecules

An

sign

membrane been

is thought as the by

the

confirmed

sulfonated The

to

OOll-9164/88/$03.50

be

in

layer

the

polysulfone charged

and

membrane gel

the

having

able

to

charge,

expel

formation

ultrafiltration membrane

ultrafiltration

and

charge

charged

the

and

less

membrane

experiments

membrane

can

they

is more

solutes

is

ovalbumin

developed

these

and

membranes

cannot

separate

interesting

applications,

charge,

it

been

Since

mechanism,

practical

the

that on

already

size.

a fixed

for of

so

sieving

molecular

membrane density

have

applications.

so-called

membrane

sign

membranes

industrial

of similar

ultrafiltration

variables:

membrane same

on

ultrafiltration

a noncharged three

based

various

pore and

fouled

size.

the

The

than it has

charged

colloids

having

than

noncharged

surface.

using

because

the

This

has

negatively

the

already charged

solutions(ref.1). separate

0 1988 Elsevier Science Publishers B.V.

charged

and

noncharged

192 solutes size.

by

an electric

The

permeation. proposed

applied

inorganic larger

pores

by

ranged

the

pH,

respectively.

isoelectric

could

as

is

also

known

the can

same

molecular

control

protein

permeation

obtained

the

to

amino or

acid

by

of

using

has

been

charged

and

based

of

substances Miyama

et

as

on the

salts

the

al.

made

a

permeation

It

or

lower

ammonium

Therefore

it

by

and

may

be

using

of

however, molecular

very size.

negatively possible

a charged

charged

and

constituents Separation

is,

of their

ones

at its

industries

solution

noncharged

positively

of

higher

proteins.

polyacrylonitrile-grafted-poly

controlled

the

at

higher

membrane

medical

blood

charged

or

(ref.5).

and

charged,

neutral

quaternary

attempted.

solution.

from

at

of

molecular

lower

charged

much

cut-off

whose

at

having

difference

positively the

the

to separate

been

had

electrically

effect

hormones,

weight

charge

food

rejected

membrane

acids,

is

membrane

such

only

are

acid

positive

important

charge,

the

molecular

charge

has

negative

amino

inorganic

antibodies,

a

though

permeates

the

fields,

pH

quaternized and

An

by

mixtures

they the

neutral

methacrylate,

separated

ultrafiltration

membrane. from

fractional

ultrafiltration

them

has even

a

increasingly

that

according

membrane

have and

controlled

also

amino

rejected

enzymes,

to separate

separate

the

separate

by

which

a negative

industrial

proteins,

filtration

namely

has

is

it becomes

well

charged

this

effect

ZOO(ref.4).

and

charged

substances

difficult

75 to

and

various

In

these

solutes charged

results

membrane,

membrane

Therefore

point

bioindustry, such

explaining

electric

salts,

point

positively

groups

both

negatively

experimental

of

This

from

isoelectric

It

means

than

its

A

model the

polysulfone

salts

lO.OOO(ref.3).

weights

pH.

to

though

is

dextrans(ref.2).

sulfonated

about

even

membrane

A quantitative

and

noncharged A

effect

glomerular

to

ultra-

ultrafiltration

N, N-dimethylaminoethyl

proteins,

by

changing

the

pH

of

a

solution(ref.6). The

purpose

of the

ultrafiltration were to

synthesized

permeate

a

present

membrane. as

membrane

membrane,

condition

in order

protein

solutions

were

level

with

the

mixed

the

charged

charged proteins

membrane

to obtain

separate

membranes

and

must

thus

to

have

at

was

isoelectric

obtained.

membranes

using

a charged

polysulfones

noncharged

pores.

In

point of

isoelectric

and

proteins

this

determined.

Separation the

by

charged

enable

large

membranes

at the

proteins

positively

In order

such

ultrafiltrated

ultrafiltration was

is to

negatively

materials.

the

casting

by

work

Both

study,

Then,

at another

a protein point

of

a

single pH

mixture one

of

attempted.

MEMBRANES Polymer

synthesis

Polysulfone(P dichloride

solution

1700,

Union

using

a

Carbide) 2/l

was

sulfur

sulfonated

at

trioxide/triethyl

30°C

in

an

phosphate

ethylene complex

193

C,H,O -@&O&@j)O&

-I- C,H.O-i=O.SOs

3

-

C2H50.S03

CH,ONa * (CHJ*CHOH

Fig.1

Procedure

(S03/TEP) then

as

of SPS

a sulfonating

neutralized

with

polysulfone(SPS) in our

was

previous

into using

shown

in Fig.2.

polysulfone(P chloromethyl

as a catalyst.

In the

methyl

The

ammonium

Union

first

and

scheme

as

groups(APS)

was

of

sulfonic sodium

the

acid

salt

synthesis,

synthesized

was step, at

the

chloromethyl

50°C

of APS

synthesis

was

reported

the

scheme

solution

polysulfone

with

to

introduced

agent then

triethylamine,

ZnCI,

*

and

reacted and

obtained.

DMF

Procedure

was

sulfonated

was

N t&H,),

Fig.2

which

group

CHCIJZHCI~

CI-

the

in a tetrachloroethane

CICH20CH3,

C2H5-$I+-CZH5

synthesized

of

according

a chloromethylation

chloromethyl solution

the

in Fig.1.

Carbide)

ether

resulting

N,N-dimethylformamide(DMF) quaternary

The

is shown

polysulfone

1700,

polysulfone

methoxide,

obtained.

charged

The

agent.

sodium

study(ref.1).

Positively (ref.7)

synthesis

zinc at

chloride 50°C

polysulfone

in a with

194 Casting

procedure

A solution l/8/0.2, plate

was

at

bath

room

at 4°C APS

from

of SPS/N-methyl-2

warmed

temperature.

with

solutions The

carried

out

membrane

on

were

of

Then

cast

in

ice-cold

with was

was

a flat

plate

no

put

into

ranged

itself

evaporation

was

a

gelation

room

order

to

out

with

the

water

obtained. temperature

from 60°C.

was

on a glass

was

period.

in

ratio

cast

at the

ratio

tried

weight

then

membrane

maintained

solution

also

and

sheet

weight

casting

water cloth

plate

and

whose

the

whose

temperature,

glass

on a glass

of

nonwoven

of the

the

period,

APS/DMF/LiN03,

temperature

a

pyrrolidone(NMP)/LiN03,

to a certain

no evaporation

membranes

l/9/0.2.

strength

or cooled

l/4/0.2 Gelation

Casting

increase

to was

an

APS

mechanical

membrane.

EXPERIMENTAL Apparatus

and materials

Ultrafiltration Fig.3.

The

module

Industries,

Ltd.)

cm.

The

channel

experiments used

was

having

height

were

carried

a thin-channel

an effective

of the

cell

flow

membrane

was

cell(UFP-2, width

of

apparatus Mitsui

7 cm

and

shown

in

Petrochemical a length

of

17

1.6 mm.

Flow meter Concentrate

Manometer 1;

f

pH controller

Permeate Feed

Thermostat Feed tank

Pump

Fig.3

Experimental

Dextrans 500

and

Blue

molecular

and

of various Dextran

weight

Three

kinds

are

molecular

of the

of proteins

listed

schematically

weights,

The

used:

molecular 1.

is Dextran

employed

in

T-10. order

T-40, to

T-70,

measure

Tthe

membranes.

were

in Table

that

were

2000(Pharmacia),

cut-off

myoglobin(Sigma).

proteins

apparatus,

bovine

weights

and

serum

albumin(BSA),

isoelectric

cytochrome

points(pI)

C

of these

195 Table

1

Molecular

Protein

weights

Molecular

BSA Myoglobin Cytochrome

C

Positively a charged Conditions

and

The

17,500

pH 7.0

12.400

pH 9.0

conditions

rate

feed

solution

6 l/min;

mg/l

in two-solute

(2)

A certain

about

pure

water

amount of

permeate

flux

and

sequence

of

steps

was

also

used

as

500,000.

experiments

temperature

were

15°C.

single-solute

followed:

feed

Concentration

experiments

and

of

about

the

50/50

pH of the

as follows:.

for

about

and

then

was

added

solution

was

concentration

hr

the

set

in

pure

to the

order

water

to

obtain

flux(PWF)

a

was

time-

measured.

system.

using

were

was

(2)-(4)

3

a NaOH

measured

repeated

at

or HCl

with

solution.

time.

different

pH

levels

of

the

solution.

(6) After

a series

chlorite The

or

performance rejection

liquid

AND

The

feed

analyzer

in

the

cleaned

using

a sodium

permeate

was

hypo-

solution.

solution the

chromatography(HPLC) from

was

Hakusui)

in the

carbon

calculated

and

the

single

solute

the

two-solute

in

concentrations

of the

system

feed

measured

and

by

system. and

the

high

Solute

permeate.

DISCUSSION

Characteristics

of the

ion

determined

lO(Henke1

concentration organic

was

the membrane

of experiments,

P3 ultrasil

solute

a total

by

membranes

exchange an

yielded

capacity(

acid-base nearly

meq/g-polymer.

In the

the

an

SPS

flux,

of solute

(5) The

methods

feed

in

was

compacted

(4) The

RESULTS

15 kPa;

procedure

was

value

using

was

ultrafiltration

100 mg/l

(3) The

feed

dextran(DEAED)(Sigma)

of DEAED

used

experiments.

membrane

independent

for

pressure

was

experimental

(1)

weight

proteins

point

pH 4.8

diethylaminoethyl

of the

procedure

flow

The

points

Isoelectric

weight

Molecular

standard

A

isoelectric

69,000

charged

solute.

and

having

IEC)

titration

equal

IEC.

The

ultrafiltration

IEC of 0.88

of

method

the

SPS

and

IEC of

by the

experiments,

meq/g-polymer

were

and an

APS

synthesized

elementary

SPS

ranged

the

membranes

used.

The

was

analysis. from

0.4

Both to

prepared

IEC of

the

1.3 from

APS

was

1.2 meq/g-polymer. The

effect

membranes mechanically

is

of

the

shown

in

stable

and

casting-solution Fig.4. had

The the

membrane highest

temperature cast

on

from

time-independent

an

the

PWF

80°C PWF,

of

the

solution while

SPS was

a 0°C

196

I

100

kPa

I

6

001 time

Fig.4

Effect of temperature membrane

casting

solution

independent PWF

was

were

cast

APS

PWF. of

from

high.

The

APS

membrane l/DMF the

solution

from

SPS

on

0.2 lacked

during

the

water

easily

very in

the

low,

flux

of a SPS

compacted.

even

though

ultrafiltration

into

the

step

on and

of a casting

a lower

The the

time-

initial

experiments

from

used

was

on

PWF

of

in a higher

solution

consisting

ultrafiltration

cloth, and

the

completely

of

nonwoven

In order

nonwoven

was

an

a sheet

tried.

solution in

a casting

in

on the

resulted

experiment

strength.

directly

or

solution

concentration

plate be

mechanical

solution

existing

gelation

not

membrane

casting

and

a glass

could

polyvinylpyrrolidone-DMF

vinyipyrrolidone

used

on pure

was

therefore

concentration

large,

obtained

sinking

was

which

membranes

polymer

of an APS

above-mentioned

membrane

membrane

very

9/LiN03

membrane

Casting

a

solution

solution.

of the was

of a casting

in

The

an 80°C

effect membranes

because

wt%

resulted

PWF of this

rather

The the

C hr I

the

dried

nonwoven removed

to

cloth before cloth from

cloth

prevent was

coated

casting. dissolved the

APS

using

the

the

casting

with

a

33

The

poly-

into

water

membrane

thus

had

high

obtained. As

illustrated

in Fig.5,

the

membrane

cast

on

the

cloth(M2)

very

197

400

I

1

I

0

m w

M2 .

-

“E e g 2DDP

APS/DMF/LINOJ

0 t

$ P 100

1

1 / 6 / 0.2

M2

1 / 9 / 0.2

13 kPa

A

0

Ml

*

.

*

Ml

A

I

I

I

1

2

3

4

time C hr 1

Fig.5

Pure water flux glass plate(M1)

Fig.6

SEM

photograph

of an APS

of cross

membrane

section

cast

on a nonwoven

of a SPS membrane

cloth(M2)

and

on a

198 and

stable

PWF,

on a glass

approximately

plate(M1)

ultrafiltration The shows

a typical

of

7

determined

shows

using

solution,

a

a

the

adsorption

of

which

Effect

of the membrane

weight

DEAED(Mw As

reduces

the to

the

Here

the

structure

charge

of

curve had

In the

membrane

of

The

in

reason

diameter

of

a

an

very

the

for

the

T-500

of

the

SPS

APS

MWCO

with

direction

this

shift

membrane

before

membrane

large

experiment

definitely

Dextran

,

might

pores.

an

of

a

BSA

of

low

be an

The

APS

ultrafiltration

and

on the Dextran

pores

4x106

of a SPS

rejection T-500

T-500(Mw were

1

C -

curve

Dextran

I

106

Weight

cut-off

membrane

membrane

with

dextrans

of polyelectrolytes

by a APS 500,000)

much

measured

larger

membrane

are

shown

was

almost

zero,

than

the

molecular

in Fig.8.

it

could sizes

be of

500,000). illustrated

in Fig.8

by the

positively

Effect

of pH on the

(i)BSA solution

0.2.

cast

solution.

of DEAED

rejection the

very

105

Molecular

that

membrane

6/LiN03

Fig.6.

membrane

, ,~,, /y(,

Fig.7

said

in

the

used.

ultrafiltration

in Fig.7.

rejection

,

The

an

shifted

Molecular

the

shown

of

l/DMF

were

cut-off(MWC0)

dextrans. after

a protein

Rejections

is

PWF

of APS

structure;

weight

of

104

Since

membrane

illustrated

no

I/

0

the

membranes

fingerlike

curve

as

BSA,

with

than

consisting

M2-type

molecular

series

MWCO

showed

experiment

SPS

However,

weight,

membrane

the

higher

is similar.

2,000,OOO.

molecular

the

asymmetric

obtained

Figure

a solution

experiments,

structure

membrane

about

from

13 times

The

because

charged

SPS

the

APS

protein

membrane

permeation

membrane BSA

positively

rejected

adsorption

charged

because

DEAED

of electrical

in the

single

BSA

completely

on

the

solute

membrane

at

was

strongly

rejected

repulsion. system any

reduced

pH

level membrane

of

the pore

199

1.0

1.0

0.8

0.8

0.6

0.6

0.4

0.4

0.2

0.2

Dextran

T-500

0 100

100

60

60

40

40

20

20

80 0

‘-

80 0

0

30 time

Fig.8

The

isoelectric pH

of

adsorbed the

30

[: min 1

permeate

point, the

BSA

negative

membrane

m v)

;II0

and

flux

pH 5, the

solution

molecules charge the

was

flux

flux

changed were

of

affected

the

started

then

from

pH

pH,

with 5 to

negatively

membrane.

pf charged

by the

decreased

as

time pH

to the

the

and

shown

of

after

and BSA

initial

noncharged

in Fig.9.

because

10.3

charged

Therefore,

to return

60

I: min I

time

Fluxes and rejections in ultrafiltration dextran solutions by an APS membrane

diameter.

The

60

thus was

value.

8o

e "E

60

5

40

P 5

20

_; 0 0

1

2

time

Fig.9

Effect of pH on flux by a SPS membrane

4

3

C hr

5

6

1

in ultrafiltration

of a BSA

solution

BSA two

were

Near

the

adsorption. hours. repulsed

removed

from

The by the

200 1.0

I

I

1

” I

I

I

II

t

u

0.5

z pe”

0 ?

60

“f

+------

5 E 0 P

pH5

.-----+-~H3.5

20

0 c >

0

0

7

1

2

3

Fig.10

Effect of pH on flux by an APS membrane the

In and

flux.

near

the

pH 3.8

case

be

of the

the 5.

APS

At

charged

adsorbed

pH

3.8, the

near

membrane,

BSA

the

the

such

however, BSA

both

rejected

that

isoelectric

flux

are

both

rejection

and

high

when

the

protein

had

It can

be seen

from

charged

proteins

charge. of

and

illustrated

neutral

and

that

obtained

with

BSA mentioned

than

those

charged membrane. and

of

were

above

C

APS

Therefore, C by

near

same

that

obtained

and

are

of the the

by the

effect

was

low

flux

at

positive

membrane

were

showed

of size

rather

low

at

positively

higher

the

membrane

and

charge

the

rejection

the

SPS

point, as

that

that

repulsion

the much

a

while

they

acts

rejection higher

were

membrane

between

the

rejection

was

the

larger

result

SPS membrane

smaller

cases,

the

from

of the

permeate

all

In

of

membrane

be supposed

through

and

membranes.

difference

pores

was

APS

isoelectric of

Electrical

membrane

of pH on rejection

SPS

with

It may

be concluded

SPS

rejection

flux

between

rejection

repulsion

the sign

permeating

it may

increase

only

the

effects

both

Figs.ll-14

membrane.

the

The

membrane.

molecule

rejected

that

molecules

for

low

the

that

The

both

permeate

acting

solution

point.

cytochrome

the APS

the

protein

cytochrome

membrane.

with

were

electrical

in Figs.ll-14 flux

and

at pH 3.8.

pores

BSA

by

pH affected

rejection

repulsion

of a BSA

membrane.

large

than

(ii)Myoglobin

high

BSA molecules

had

6

solution's

both

electrical

and

5

in ultrafiltration

the

that

5) and

by

neutral

rejection

shows

point(pH

membrane

and

APS

explained

Electrically and

the

IO clearly

isoelectric

can

charge

of

Figure

and

4 1. hr 1

time

pH

-j

40

are

smaller

more

strongly

pore

of

a

of charged than

that

than

obtained

on

a

charged myoglobin

by

the

APS

201

1.0

0

I

1

I

1

2

3

time

Fig.11

I: hr

I 4

I

Effect of pH on Flux and rejection solution with a SPS membrane

Fig.12

Effect of pH on flux solution with an APS

of a myoglobin

90

60 time

in ultrafiltration

C min

and rejection membrane

1

in ultrafiltration

of a myoglobin

202

1.0 l-l

I

I4

0.5

d

a” 0 ‘? i

60

p

40

I

1

I

I

pH9 ------+pHlO

+pHll-

0 &

20

c -:

0

I 0

I

I

1

1

I

1

2

3

4

5

time

Fig.13

C hr 1

Effect of pH on flux and rejection solution with a SPS membrane

in ultrafiltration

of a cytochrome

C

of a cytochrome

C

1.0 m I u 2 a”

0.5

0 7

60.

“E Q

40-

1

I

I

I

-:

-

Y

fJ I

0 0

I

1

I

I

Effect of pH on flux solution with an APS

I

I

90

60

30 time

Fig.14

1

pH 9.7

pH 6.7

z 6 c

I

I

120

[: min 1

and rejection membrane

in ultrafiltration

203

Separation of the protein mixture A mixture of myoglobin and cytochrome C was separated with the SPS and the APS membranes,

The r e s u l t s

o b t a i n e d are shown in Figs,15 and 16.

In the

experiment with the SPS membrane the solution pH was c o n t r o l l e d at pH 9.2, Thus, the cytochrome C molecules were almost e l e c t r i c a l l y neutral and the myoglobin molecules were negatively charged. I t is obvious from Fig, 15 that the negatively charged myoglobin was rejected because of the e l e c t r i c a l repulsion caused by the negative charge of the SPS membrane, while the neutral cytochrome C permeated the membrane without the e l e c t r i c a l e f f e c t , As shown in Fig. 15o under the condition of lower f l u x , that is a flow rate of 4 I/min and pressure of 5 kPa, the r e j e c t i o n of myoglobin was higher than that obtained at a flow rate of 6 I/min and pressure of 15 kPa, The smaller f l u x reduces the e f f e c t of the concentration p o l a r i z a t i o n , namely a lower concentrat i o n at a membrane surface which results in a higher r e j e c t i o n . On the other hand, the smaller flow rate increases the e f f e c t of the concentration p o l a r i z a t i o n which results in the lower r e j e c t i o n ,

I t could be supposed that,

in t h i s

experiment, the e f f e c t of the smaller f l u x was larger than that of the lower flow rate. Therefore, the net r e j e c t i o n observed was r e l a t i v e l y increased. From Figs.13 and 15, i t can be seen that there are some effects of mixing on the r e j e c t i o n of proteins, As shown in Fig. 13, the r e j e c t i o n of cytochrome C at pH 9 is about 50%, while in Fig.15 i t

is almost zero. The reason f o r t h i s is

not clear at present.

1,0

I

"

I

I

.

I

0.5

CytochromeC

o

.It

0

~

~

.t

~

~

I~

~ = =

60 ~

4O

7 o

2O

°"°." ' ' 4 ~ t 6 1 15/kPam i ~n!= 41/m!n~-!,

T'-' I

0

Fig.15

1

l

'

I

2 3 time [ hr ]

/

4

Separation of mixed myoglobin and cytochrome C by a SPS u l t r a f i l t r a t i o n membrane

204

m ’ L-I

0.4

g cc

0.2

Cytochrome

l

C

0

Myoglobin

0

0

30

60 time

Fig.16

Separation membrane

The

separation

satisfactory, was

near

the

charged

as

of

shown

effects

in Fig.16.

The

point

as

experiment

pH

shown

was

120

cytochrome

C by an APS

and

myoglobin

experiment

of myoglobin. 5.5

in

very

90 1

and

C

C at

However,

the

myoglobin

cytochrome

isoelectric

cytochrome

membrane. from

of mixed

C min

was

the

about

was

by

the

APS

membrane

performed

It was

rejected

Fiy.16,

low:

by

assumed the

rejection indicating

the

SPS

at

that

which

charged

there

not

positively

cytochrome

that

was

pH 5.5, the

positively of

20%,

ultrafiltration

C

APS

obtained

must

be

some

of mixing.

From

both

possible

to

results

separate

charged

ultrafiltration

proteins

are

very

obtained proteins

with

having

membrane,

different

even

though

and

APS

membranes,

isoelectric the

points

molecular

it by

seems

using

weights

of

a the

similar.

CONCLUSIONS In order points

using

charged

separate

ultrafiltration

polysulfone molecular heated

to

a charged

having weight

membranes quaternary

cut-offs

to 80 or 60°C.

proteins

based

ultrafiltration

could

were

made

ammonium be

on

the

membrane, from

difference both

sulfonated

groups(APS).

developed

of

by

their

negatively

isoelectric

and

positively

polysulfone(SPS) Membranes

casting

a

with

polymer

and large

solution

205 Both rejected

negatively charged

a charged

dextran

electrically

volume

because

charge A

sign

of

was

mixture

ultrafiltration of

one

of

which

neutral

Permeate point

and

of

while

repulsion

acting

such have

proteins flux

protein

equal

of

the

same

near a

protein

charged the

The

sign

solution

on

a

protein

as

was

the

C

was be

near

the

the

because the

of

of

the

protein

increased. by

a

charged

isoelectric

protein

and

isoelectric

when

separated

pH near

neutral

and

low.

greatly

strongly points

Rejection

was

However,

could

rejected

molecules

membrane.

low

flux

solution's

membranes

isoelectric

points

was

the

electrically

protein

their

membrane.

charge,

the

near

isoelectric

cytochrome

by setting

mixed.

between

and

ultrafiltration not

charge

their

adsorption

myoglobin

the

charged

as proteins

to the membrane

membrane

proteins

membrane,

positively

solutes,

permeated the

point the

electrical

membrane.

REFERENCES I. Jitsuhara and S. Kimura, Structure and properties of charged ultrafiltration membranes made of sulfonated polysulfone. J. Chem. Enq. _ _ Japan, 16(5), (1983) 389-393. W. M. Deen, B. Satvat and J. M. Jamieson, Theoretical model for glomerular filtration of charaed solutes, Am. J. Phvsiol., 238 (1980) F126-F139 I. Jitsuhara and S: Kimura, Rejection of-inorganic salts by charged ultrafiltration membranes made of sulfonated polysulfone, J. Chem. Eng. Japan, 16(5) (1983) 394-399 S. Kimura and A. Tamano, Separation of amino acids by charged ultrafiltration (Ed.), Membranes and membrane membranes, in: E. Driori and M. Nakagaki processes, Plenum Press, New York, 1986, pp.l91-197 S. Abe, S. Nakao and S. Kimura, Separation in two-component system by charged ultrafiltration membrane, Proc. of 7th Annual Meeting of the Membrane Society of Japan, Tokyo, Japan, May 24-25, 1985, p.19 H. Miyama, K. Tanaka, N. Fujii, H. Tanzawa and S. Nagata, Charged ultrafiltration membrane for permeation of proteins, Proc. of the 1987 Int. Congress on Membranes and Membrane Processes, Tokyo, Japan, June 8-12. 1987, pp.376-377 S. Abe, M. Date and T. Kawakita, Charged ultrafiltration membrane and its manufacturing process, Jpn. Kokai Tokkyo Koho (1987)