U.S. patent application number 10/285240 was filed with the patent office on 2003-05-15 for membrane adsorber device.
Invention is credited to Phillips, Michael W..
Application Number | 20030089664 10/285240 |
Document ID | / |
Family ID | 23312220 |
Filed Date | 2003-05-15 |
United States Patent
Application |
20030089664 |
Kind Code |
A1 |
Phillips, Michael W. |
May 15, 2003 |
Membrane adsorber device
Abstract
The present invention is an adsorber membrane and a device
containing one or more such membranes. Both the membrane and the
device have a Peclet number (Pe) of at least 100. The membrane and
the device are designed for the removal of trace contaminants in
protein containing streams such as exist in the biopharmaceutical
industry. A preferred membrane has tight pore size distribution and
high permeabilities that allow for high throughput separations. A
device of the present invention can contain a flat sheet membrane
such as a pleated filter, a tangential flow filter or a spiral
wound filter. Preferably, the device is formed in a stacked disk
arrangement where one or more layers of membrane are sealed to each
of the two large surface of the disk. One such device is formed of
a series of disks, each disk having eight layers of membranes
sealed to each of the two large surfaces of the disk. These disks
are placed within a sealed capsule having an inlet on one end and
an outlet on the other. The disks are sealed so that all fluid that
exits the outlet does so by having first passed through the
membranes on one side of a disk.
Inventors: |
Phillips, Michael W.;
(Tyngsborough, MA) |
Correspondence
Address: |
MILLIPORE CORPORATION
290 CONCORD ROAD
BILLERICA
MA
01821
US
|
Family ID: |
23312220 |
Appl. No.: |
10/285240 |
Filed: |
October 31, 2002 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
|
60335543 |
Nov 2, 2001 |
|
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|
Current U.S.
Class: |
210/660 ;
210/435; 210/456 |
Current CPC
Class: |
B01D 2313/90 20130101;
B01D 63/08 20130101; G01N 30/38 20130101; B01D 61/00 20130101; B01D
63/084 20130101; G01N 2030/527 20130101; B01D 2313/08 20130101;
B01D 61/18 20130101; G01N 1/405 20130101; C07K 1/34 20130101; G01N
1/34 20130101; G01N 2030/527 20130101 |
Class at
Publication: |
210/660 ;
210/435; 210/456 |
International
Class: |
B01D 061/00; B01D
029/00 |
Claims
What I claim:
1. A membrane adsorber device comprising a housing having an inlet
and an outlet and one or more layer of an adsorptive membrane
wherein the device has a Peclet number (Pe) of at least 100.
2. The device of claim 1 wherein the Pe is at least 150.
3. The device of claim 1 wherein the Pe is at least 200.
4. The device of claim 1 wherein the Pe is at least 500.
5. The device of claim 1 wherein the Pe is at least 2000.
6. The device of claim 1 wherein the Pe is from about 100 to about
4000.
7. The device of claim 1 wherein the one or more layers of
membranes are formed on one or more porous disks with their inner
and outer edges being sealed to the disks forming a space between
the membranes and the disk into which filtered liquid can flow, an
outlet from the disk for the filtered liquid.
8. The device of claim 1 wherein the one or more membranes have a
Pe of at least 100.
9. The device of claim 1 wherein the one or more membranes have a
Pe of at least 200.
10. The device of claim 1 wherein the one or more membranes have a
Pe of at least 500.
11. The device of claim 1 wherein the one or more membranes have a
Pe of at least 1000.
12. The device of claim 1 wherein the one or more membranes have a
Pe in the range of from about 100 to about 4000.
13. The device of claim 1 further comprising a flow distributor
located between the inlet and the one or more layers of
membranes.
14. A process for the removal of trace contaminants from an aqueous
protein containing stream comprising the steps of selecting a
membrane based adsorber device wherein the membrane and device each
have a Pe of at least 100 and flowing an aqueous protein containing
stream through said membranes of said device to remove any trace
contaminants.
15. The process of claim 14 wherein the trace contaminants are
selected from the group consisting of viruses, endotoxins, DNA, RNA
and mixtures thereof.
16. A process for determining the Peclet number of a membrane
absorber comprising the steps of: (a) equilibrating the membrane or
membrane adsorber device with an equilibration buffer at a known pH
and conductivity; (b) challenging the membrane with a known
concentration of a specific solute in the equilibration buffer; (c)
monitoring the breakthrough of the solute downstream of the
membrane as a function of value selected from the group consisting
of time, challenge volume and other suitable variable related to
total quantity of material challenged to membrane; (d) analyzing
the solute breakthrough curve to determine pertinent flow
characteristics of the membrane adsorber device; and (e) comparing
the results calculated in step (d) to results from known integral
devices.
17. The method of claim 16 wherein the analysis of (d) is by
calculating the sharpness of the breakthrough curve.
18. The method of claim 16 wherein the monitoring of step (c) is by
a detector.
19. The method of claim 16 wherein the analysis of (d) is by
monitoring the time of solute breakthrough.
20. The method of claim 16 wherein the analysis of step (d) is by
monitoring a variable related to total quantity of material
challenged to membrane adsorber at a point in time selected from
the group consisting of the initial onset of solute breakthrough
and a fraction of solute breakthrough.
21. The method of claim 16 wherein the analysis of step (d) is by
monitoring a variable related to total quantity of material
challenged to membrane adsorber at a specific fraction of solute
breakthrough.
22. The method of claim 16 wherein the analysis of step (d) is by
monitoring a variable related to total quantity of material
challenged to membrane adsorber at a specific fraction of solute
breakthrough wherein that fraction is from about 5% to about
50%.
23. The method of claim 16 wherein the analysis of step (d) is by
monitoring a variable related to total quantity of material
challenged to membrane adsorber at a specific fraction of solute
breakthrough wherein that fraction is from about 5% to about
20%.
24. The method of claim 16 wherein the analysis of step (d) is by
monitoring a variable related to total quantity of material
challenged to membrane adsorber at a specific fraction of solute
breakthrough wherein that fraction is from about 5% to about
10%.
25. The method of claim 16 wherein the analysis of step (d) is by
calculating a breakthrough curve sharpness and an initial onset of
solute breakthrough.
Description
[0001] The present invention relates to an adsorber membrane and a
device containing it for removing selected components from a liquid
stream. More particularly, it relates to a membrane based adsorber
device having a membrane and a device containing one or more such
membranes, each with a Peclet number (Pe) of greater than 100.
BACKGROUND OF THE INVENTION
[0002] The use of membrane chromatography or normal flow membrane
based adsorbers is well known; see U.S. Pat. No. 4,895,806 and
Membrane Chromatography: Preparation and Applications to Protein
Separation, Zeng, X, Biotechnol. Prog 1999, vol 15,
p.1003-1019.
[0003] All of these devices are basically formed of a housing
having an inlet and an outlet and one or more layers of an
adsorptive membrane located between the inlet and outlet such that
all liquid entering the inlet must flow through the one or more
membrane layers before reaching the outlet. The membranes are
typically rendered adsorptive by surface modification, in situ
copolymerization or grafting, direct formation from adsorptive
materials or by the inclusion of adsorptive particles (such as
chromatography media) in the membrane matrix during formation of
the membrane. In this way, one or more constituents of the liquid
stream are bound to the membrane surface and removed from the
stream. After completion of the filtration step, the bound material
is then eluted by adding a different solution or changing pH
conditions or by other well known methods in the art and either
disposed of or processed and used for whatever purpose.
[0004] Typically, the material removed is the protein of interest.
The remainder of the materials in the stream, such as viruses,
endotoxins, nucleic acids, host cell proteins and the like pass
through the device unhindered and are removed from the system.
[0005] Some have suggested removing the trace contaminants such as
viruses, endotoxins, nucleic acids, host cell proteins and the like
from the stream instead of removing the protein of interest.
Traditionally, this has been done through the use of chromatography
columns containing media with quaternary amine chemistry. This
approach has several advantages such as higher yields of the
product of interest. However, it has several disadvantages. For
one, the use of columns results in a significant underutilization
of the capacity of the column components, typically less than 1%.
Moreover, the process is time consuming often taking hours to
complete due in large part to long residence time required for the
stream to be in the presence of the chromatography media. Lastly,
the cost of the media, additional buffers, along with the QC and
validation costs associated with their use, significantly impact
the economics of using chromatography columns for this
application.
[0006] The potential of using membrane-based adsorbers in lieu of
chromatography media has been mentioned to overcome the above
problems. However, the current devices have their own set of
problems that need to be overcome. The problem with these devices
has been that they are not efficient and are therefore expensive to
make and operate. The mere adding of layers does not increase the
efficiency. Instead it merely adds to the expense of the
manufacture of the device and its operation. Some have tried
various flow distribution devices such as tapered end plates and
screens similar to what is traditionally used in chromatography
columns to improve efficiency. Yet the overall results have not
been satisfactory.
[0007] What is needed is a membrane adsorber device that is
efficient, utilizes its capacity, has high throughput and
preferably is disposable so as to eliminate the need for cleaning
and revalidation of the device before reuse. The present invention
provides such a membrane and device, especially for trace
contamination removal.
SUMMARY OF THE INVENTION
[0008] The present invention is an adsorber membrane and a device
containing one or more such membranes. Both the membrane and the
device have a Peclet number (Pe) of at least 100. The membrane and
the device are designed for the removal of trace contaminants in
protein containing streams such as exist for example in the
biopharmaceutical industry. A preferred membrane has tight pore
size distribution, uniform capture mechanism densities and
capacities (regardless of whether the capture mechanism is ligand
based or otherwise) and high permeabilities that allow for high
throughput separations.
[0009] A device of the present invention can contain a flat sheet
membrane such as a pleated filter, a tangential flow filter or a
spiral wound filter. Preferably, the device is formed in a stacked
disk arrangement where one or more layers of membrane are sealed to
each of the two large surface of the disk. One such device is
formed of a series of disks, each disk having eight layers of
membranes sealed to each of the two large surfaces of the disk.
These disks are placed within a sealed capsule having an inlet on
one end and an outlet on the other. The disks are sealed so that
all fluid that exits the outlet does so by having first passed
through the membranes on one side of a disk. Such a device is
linearly scalable.
IN THE DRAWINGS
[0010] FIG. 1 shows a graph of a series of Pe values.
[0011] FIG. 2 shows a cutaway view of a device of the present
invention according to a first embodiment of the present
invention.
[0012] FIG. 3 shows an exploded view of a portion of a device of
the present invention according to a first embodiment of the
present invention.
[0013] FIG. 4 shows an exploded view of a device of the present
invention according to a second embodiment of the present
invention.
[0014] FIG. 5 shows a planar view of a disk useful in one of the
embodiments of the present invention.
[0015] FIG. 6 shows a representative cross-section of a device
according to the embodiment of FIG. 3 and the fluid flow path
through it.
[0016] FIG. 7 shows a close view of the representative
cross-section of a device according to the embodiment of FIG. 3 and
the fluid flow path through it of FIG. 6.
[0017] FIG. 8 shows the Pe values obtained by an integral membrane
packet and one having one layer compromised.
[0018] FIG. 9 shows the graph of the Pe data for currently
available adsorber membranes.
[0019] FIG. 10 shows the bacteriophage LRV plotted as a function of
challenge linear velocity.
[0020] FIG. 11 shows the bacteriophage LRV plotted as a function of
number of membrane adsorber layers.
DETAILED SPECIFICATION OF THE INVENTION
[0021] The level of trace contaminants in feed stream can vary but
they are typically low, generally in the parts per million (ppm)
range or lower and the desire is to remove those contaminants such
as viruses, endotoxins, DNA and host cell proteins to
non-detectable levels. For example, in a typical feedstream, one
has virus levels at from 1 to 10 ppm, DNA at 100 picograms/mL,
endotoxins at 10 EU (endotoxins units)/mL and host cell proteins at
from about 10 to about 100 nanograms/mL. Removing these
contaminants effectively and efficiently is a difficult task.
[0022] It has been discovered that an efficient membrane based
adsorber device can be made for use in protein purification by
utilizing a membrane and device configuration, each of which has a
Peclet number (Pe) of at least 100. When the membrane and device
both have a Pe of 100 or greater, one achieves high retention and
efficiency with good flow and yield characteristics. Additionally,
one is able to make a device that is linearly scalable which is of
great benefit to the user.
[0023] The Peclet number (Pe) is derived from Peclet analysis and
relates to the generation of a breakthrough curve for contaminants.
Basically, a test material (that is representative of the trace
contaminant) is flowed through a selected adsorptive membrane or
device and the amount of the test material that is in the filtrate
is measured. The valves are plotted on a graph with volume on the
X-axis and breakthrough % on the Y-axis. For low Pe values, such as
those below 10, the breakthrough of test material appears nearly
immediately, well below the capacity of the membrane device, i.e.
exhibiting poor efficiency. For higher Pe values, such as above 100
and preferably above 1000, the breakthrough curve begins to
approach ideality and breakthrough corresponds to the capacity of
the device, i.e. high efficiency.
[0024] FIG. 1 shows the theoretical plot of Pe values described
above. The ideal plot is a vertical line. The closer the curve
becomes to vertical, the higher the Pe value.
[0025] For adsorptive applications, such as trace contaminant
removal in validated biopharmaceutical applications, breakthrough
is the critical issue, as contamination of the filtrate at any
appreciable level is not allowed. It has been found that the Pe
value is an important predictor of adsorptive performance of
membranes and devices containing them.
[0026] Additionally, with viral removal the Pe value appears to
correlate to the LRV of the membrane and device. LRV means log
reduction value and is represented by the ratio of two numbers. In
viral applications it is represented by the number of viral
particles that are contained on the upstream side of the filter to
the number of viral particles found in the filtrate. Therefore a
LRV of 4 means that the membrane was challenged with 10.sup.4
particles and only one was found in the filtrate. The log of this
ratio being 4. This means that the membrane is capable of removing
99.99% of all viral particles.
[0027] Ion exchange capacity is not an acceptable predictor of
performance in adsorptive devices for trace contaminant removal as
all devices have excess capacity relative to the volume of
contaminant to be removed. The issue is ensuring that the
contaminant, often present in the ppm range, is removed efficiently
and as completely as possible.
[0028] A method used by Applicant to determine the Peclet number of
a membrane (housed in a device optimally designed to minimize
upstream and downstream dead volumes while affording adequate fluid
distribution to effectively challenge entire membrane area) or
device containing one or more membranes is as follows:
[0029] (1) Equilibrating the membrane or membrane adsorber device
with an equilibration buffer at a known pH and conductivity.
[0030] (2) Challenging the membrane or membrane adsorber device
with a known concentration of a specific solute in the
equilibration buffer. The selection of the specific solute is
typically based upon its ability (or inability) to bind to the
adsorber by a specific mechanism and its ability to be readily
detected downstream (various chromophores for UV detection,
fluorescently labeled solutes, etc.). The selection of the
challenge solute concentration is typically based upon both the
thermodynamic principles associated with solute adsorption
(adsorption isotherm characteristics) and the specific objectives
of the test. For example, if the objective of the test were to
characterize the performance of a specific membrane adsorber
device, one would select buffer and solute conditions and
concentrations such that the solute binding characteristics to the
membrane adsorber were within the linear part of the solute
adsorption isotherm. However, if the objective of the test was to
characterize the flow characteristics through only the membrane
(thereby minimizing the effects of upstream dead volume) one would
typically select buffer and solute conditions and concentrations
such that the solute binding characteristics to the membrane
adsorber were within the non-linear part of the solute adsorption
isotherm.
[0031] (3) Using a suitable detector, monitor the breakthrough of
the solute downstream of the membrane adsorber as a function of
time, challenge volume, or other suitable variable related to total
quantity of material challenged to membrane adsorber.
[0032] (4) Analyze the solute breakthrough curve to determine
pertinent flow characteristics of the membrane adsorber device.
This analysis could include
[0033] (a) calculating the sharpness of the breakthrough curve. One
such means for calculating breakthrough curve sharpness is by
calculating an effective Peclet number (Pe), details of which are
described below. High Pe values are associated with uniform flow
through the membrane adsorber, uniform density and distribution of
the capture mechanism and effective distribution of flow to the
entire membrane adsorber surface. A device with a high Pe number
would most likely have good trace impurity retention
characteristics. Low Pe values are associated with poor flow
distribution properties associated with the membrane adsorber
device, excessively large flow dispersive characteristics of the
membrane adsorber, poor capture mechanism distribution and/or
densities or a combination of two or more of these. Low Pe values
may indicate that trace impurity retention characteristics are
compromised.
[0034] (b) monitoring the time (or other suitable variable related
to total quantity of material challenged to membrane adsorber) at
either initial onset of solute breakthrough or at a specific
fraction of solute breakthrough (e.g., 5% or 10%). Premature
breakthrough of the solute relative to some standard (e.g., 50%
breakthrough) may indicate the presence of defects that may
compromise trace impurity retention characteristics. The ability to
detect premature breakthrough is highly dependent upon the
breakthrough curve sharpness, as calculated above in step (a). For
example, the detection of defects in membrane adsorber devices that
exhibit very sharp breakthrough curves is much easier than in
devices in which the breakthrough curve is very diffuse. The
sharpness of breakthrough curves can be enhanced by utilizing
membranes with inherently high Pe numbers, designing membrane
adsorber devices with low dead volumes and good flow distribution
properties, by exploiting the thermodynamics of non-linear
adsorption, or by a combination of any of the above.
[0035] (c) calculating both the breakthrough curve sharpness and
initial onset of solute breakthrough (a combination of steps (a)
and (b)). In this manner, defects and/or flow distribution
properties (either membrane or device related) that may compromise
trace impurity retention characteristics could be detected.
[0036] (5) Comparing the results calculated in step (4) to results
from known integral devices, thereby determining either the
integrity of the membrane adsorber or membrane adsorber device or
the ability of such a device for removing trace impurities.
[0037] As stated above, one such means of determining the sharpness
of a breakthrough curve is by calculating an effective Peclet
number (Pe). Breakthrough curves are typically sigmoidal in shape
(s-shaped). Lapidus and Amundson (Lapidus, L. and N. R. Amundson,
"Mathematics of adsorption in beds. VI. The effect of longitudinal
diffusion in ion-exchange and chromatographic columns," J. Phys.
Chem., 56, 984 (1952).) developed a mathematical model that related
the shape of the breakthrough curve to various model parameters,
given by: 1 C A C 0 = 1 2 { 1 + erf [ ( Pe ) 1 2 ( V - V _ ) 2 ( V
V _ ) 1 2 ] } equation ( 1 )
[0038] where
[0039] C.sub.A is the effluent solute concentration
[0040] C.sub.0 is the inlet solute concentration
[0041] V is the challenge volume
[0042] V.sub.bar is the challenge volume at 50% breakthrough
(C.sub.A/C.sub.0=0.5)
[0043] Pe is the Peclet number
[0044] From a technical point of view, this equation was derived
for linear systems. However, this form of equation can be used to
interpret any breakthrough curve. Accordingly, an effective Pe can
be determined by simply fitting this equation to an experimentally
determined breakthrough curve. Various means for fitting
breakthrough curve data to this equation include (a) a
least-squares fit by which all the data is simultaneously used to
determine a best-fit (one which minimizes the least-squares error)
(b) a multipoint method by which a discreet number of points (2 or
3) are used (c) or any of several other methods. An example of
method (b) is to use equation (1) to make a generic plot of Pe
versus 2 V 90 - V 10 V 50 ,
[0045] where V.sub.10, V.sub.50, and V.sub.90 and the breakthrough
volumes corresponding to 10, 50, and 90% solute breakthrough,
respectively. Then, from the experimental breakthrough curve,
determine the values for V.sub.10, V.sub.50, and V.sub.90. Then,
from the generic Pe plot, determine the effective Pe number. It
should be noted, however, that this is only one means by which the
sharpness of the breakthrough curves can be quantified.
[0046] While the above process is the preferred means for obtaining
a Pe of a membrane or device, others methods can be developed that
would quantify the breakthrough curve sharpness and thereby provide
one with the same type of relevant information. It is meant by this
invention to encompass and include those methods within the
teachings of the present invention.
[0047] The membrane of the present device must have a Peclet number
that is sufficiently high to accomplish the level of contaminant
removal that is desired. Typically, the membrane(s) itself will
have a Pe of from about 100 to greater than 10,000. Preferably, it
is at least 100, more preferably at least 200, even more preferably
at least 500 or at least 1000 and most preferably at least 10,000
or greater.
[0048] It has been found that the device Pe will typically be equal
to or lower than the Pe of the membrane. To date, no device has
been found that is capable of having a Pe higher than the Pe of the
membrane. A variety of device properties such as poor flow
distribution, the method of membrane incorporation into the device
(pleating, stacked disk, other methods) and the like can adversely
affect the Pe number of the device. Therefore, the use of membranes
having a Pe higher than the desired device Pe is recommended.
[0049] The membrane may be a microporous or macroporous membrane
formed of a polymer selected from olefins such as polyethylene,
including ultrahigh molecular weight polyethylene, polypropylene,
EVA copolymers and alpha olefins, metallocene olefinic polymers,
PFA, MFA, PTFE, polycarbonate, vinyl copolymers such as PVC,
polyamides such as nylon, polyesters, cellulose, cellulose acetate,
regenerated cellulose, cellulose composites, polysulphones,
polyethersulphones, polyarylsulphones, polyphenylsulphones,
polyacrylonitrile, polyvinylidene fluoride (PVDF), and blends
thereof. Additionally, nonwoven and woven fabrics of the same
materials, such as Tyvek.RTM. paper available from E. I. DuPont de
Nemours and Company of Wilmington, Del. Likewise fibrous media such
as a cellulosic pad, MILLISTAK+.TM. filtration media available from
Millipore Corporation of Bedford, Mass. may be used. The membrane
selected depends upon the Pe, the desired filtration
characteristics, the particle type and size to be filtered and the
flow desired.
[0050] The membranes selected must be capable of adsorbing one or
more species from a desired stream of liquid. The membranes, such
as regenerated cellulose membranes, are inherently functional such
that no further treatment is required. However in most cases, the
selected membrane is either not functionalized or is insufficiently
functionalized such that additional treatment of the membrane is
required. There are several well-known methods of rendering
membrane functional. See Membrane Chromatography: Preparation and
Applications to Protein Separation, Zeng, X, Biotechnol. Prog 1999,
vol 15, p.1003-1019. The functional characteristic may be one or
more of the following: hydrophilicity, hydrophobicity, charge
(positive or negative), oleophilicity, oleophobicity, and ligand
chemistry.
[0051] The most common methods of rendering a membrane material
functional include: incorporating a functional material into a
membrane structure during formation; treating the surface of the
membrane with a functionalizing material and grafting or
polymerizing and crosslinking the functional material onto the
surface and the use of ligands bound to the membrane surface.
[0052] One can add the material as a solid such as ion exchange
resin or chromatography media into the membrane structure during
formation, see U.S. Pat. No. 5,531,899. Alternatively, one can add
a liquid component such as PVP into the batch material used to make
the membrane to provide the desired functionality.
[0053] Preferably, one uses a surface treatment of the preformed
membrane. By surface it is meant all surfaces of the membrane, the
upper and lower faces as well as the inner walls of the pores in
the membrane structure. This can be a monomer that is polymerized
and crosslinked in place such as is taught in U.S. Pat. No.
4,944,879 or it may be a polymer such as is taught in U.S. Pat.
Nos. 5,629,084 and 5,814,372. U.S. Pat. No. 5,137,633 teaches
adding two components a monomer for philicity and epichlorohydrin
adding a positive charge to the membrane.
[0054] Alternatively, one may graft a polymer onto the membrane
surface such as is taught by U.S. Pat. No. 4,340,482 although this
is not preferred due to the harsh treatment conditions imposed and
the deterioration of the membrane structure that occurs under such
conditions.
[0055] The above membranes as treated may be used as is or if
desired, ligands such as quaternary amines can be bound to their
surfaces to impart a different or increased selectivity. Typically
the membrane surface is first treated to render it hydrophilic and
then the ligands are attached to the membrane surface via a linker
arm. See U.S. Pat. Nos. 4,923,901, 5,547,760, 5,618,433 and
5,980,987.
[0056] Any of the above methods as well as any other method that
results in an adsorbing membrane can be used in the present
invention.
[0057] There should be sufficient membrane in the device to provide
the required Pe and capacity desired. Depending on the device
configuration, this typically means more than one layer of membrane
in a device. It has been found that there is a minimum number of
membrane layers that are required to achieve the desired Pe in a
given device format. The number required depends upon the membrane
and the device format selected. Typically, it has been found that
two to four layers are sufficient to give one the desired Pe. In a
preferred embodiment of the present invention as shown in Example
3, one layer provided a sufficiently high Pe number, with 3 layers
providing the maximum Pe obtainable with the selected membrane.
Additional layers can then be added to provide the desired
capacity.
[0058] Additionally, one may use two or more different membranes to
achieve even greater efficiency by selecting a membrane that is
most efficient for a particular contaminant.
[0059] Commercial products useful in this invention include a 0.65
micron nominal pore size membrane known as Durapore.RTM. membrane
available from Millipore Corporation of Bedford, Mass. This
membrane is modified to render it philic and to carry a positive
charge. This membrane has a Pe of at least 2000, preferably 4000
when tested by the Peclet test described below in an eight layer
format. Other membranes that are useful in the present invention
include CHEMPURE 1 membranes available from Millipore Corporation
of Bedford, Mass. and EMPOR membranes available from 3M of
Minneapolis, Minn., both of which are membranes that incorporate
particulate chromatography media into the membrane structure;
regenerated cellulose membranes such as the charged PL series of
membranes available from Millipore Corporation of Bedford, Mass.,
ICDM membrane available from Millipore Corporation of Bedford,
Mass., Mustang membranes available from Pall Corporation of East
Hills, N.Y. and Sartobind membranes available from Sartorius GmbH
of Germany.
[0060] In designing a device that is suitable for the present
invention, several factors appear to contribute to the success of
the device and to the achievement of a high Pe value. First, the
membrane must have a Pe of at least 100 by the Pe test defined
herein. Second, the membrane selected should have a relatively
inherently narrow pore size distribution. This allows one to
achieve the maximum achievable Pe number for a given type of
membrane. Also, this determines the minimum number of layers
required to effect the separation. Further, the membrane should
have as even a density of capture sites as possible throughout the
membrane. It is believed that the Pe can be adversely affected by
poor or non-uniform capture mechanism (e.g. poor ligand
distribution) distribution in a membrane.
[0061] Additionally, one should endeavor to select a device design
that will minimize the upstream dead volumes and mixing zones to
maintain as even and uniform a flow as possible. Lastly, one should
design the upstream and downstream fluid paths to minimize
residence time distribution of various fluid paths thereby
maximizing flow and keeping them as uniform as possible from point
in a device to another.
[0062] An additional advantage of a properly designed device of the
present invention is that the device is linearly scalable. By
"linearly scalable" it is meant that one is able to design devices
having a given Pe is sizes that are useful for research, pilot and
production scale processes and that performance of all of the
devices will be essentially the same regardless of the size of the
process used. For example, this means that work done with a small
scale device will allow one to select and use the same device
configuration with additional area and volume and have it work with
the Pe at pilot or production scale. This is a great advantage in
that it eliminates the need to redesign the device at each scale
and provides one wit the knowledge and safety that the selected
device will work for all uses when one attempts to scale up one's
process to production levels. It reduces cost and time required in
the scale up and it also simplifies validation of the process and
device.
[0063] A device of the present invention can contain a flat sheet
membrane such as a pleated filter, a tangential flow filter or a
spiral wound filter. Preferably, the device is formed in a disk
arrangement, more preferably a stacked disk arrangement where one
or more layers of membrane are sealed to each of the two large
surfaces of the disk. One such device is formed of a series of
disks, each disk having two or more, preferably eight layers, of
membranes sealed to each of the two large surfaces of the disk.
These disks are placed within a sealed capsule having an inlet on
one end and an outlet on the other. The disks are sealed so that
all fluid that exits the outlet does so by having first passed
through the membranes on one side of a disk. The disk arrangement
provides one with a parallel arrangement of membranes which provide
for uniform and parallel fluid distribution and flow at relatively
low pressure drops with little mixing or dead volume.
[0064] FIG. 2 shows a first embodiment of the device of the present
invention. This device is a small scale device based on a
MILLEX.RTM. device available from Millipore Corporation of Bedford,
Mass. The device has a top portion 10 having an inlet 12, a bottom
portion 14 having an outlet 16 and a porous membrane support
platform 18. A packet of membrane 20 is sealed to the bottom
portion 14 before the top portion 12 and bottom portion 14 are
sealed together such that all fluid must pass through the membrane
packet 20 before reaching the outlet 16. It has been found that
depending upon the capacity of the membrane selected and the
desired capacity of the device one can use two or more layers of
membrane in the packet 20. In a preferred embodiment, a 25 mm
diameter MILLEX.RTM. device was loaded with 8 layers of 0.65
hydrophilic charged DURAPORE.RTM. membrane to yield a device having
3.5 cm.sup.2 frontal area and 0.35 ml bed volume.
[0065] The packet was sealed around its inner and outer edges. This
was accomplished using a heat seal although other methods such as
epoxy or urethane adhesives, vibrational welding or polyolefin
overmolds could be used. If desired on can seal the packet in the
device separately rather than as part of the device sealing
process.
[0066] FIG. 3 shows a portion of the stacked disk device that is
useful in the present invention. This is used in large scale (pilot
or production) processes. The device as shown is based on a
MILLIDISK.RTM. device available from Millipore Corporation of
Bedford, Mass. In this FIG. 3, a series of disks 30, each having
one or more layers of adsorptive membranes bonded to each of their
two major faces. The disks 30 have been spaced apart from each
other and attached to each other by their outer rims to prevent
distortion by spacing lugs 32. The disks are also sealed to each
other by an inner sealing rim 33 shown in FIG. 5 and attached to an
outlet plate 34 of the device. This outlet plate 34 is comprised of
a relatively flat surface (not shown) having an outlet (not shown)
in the middle of the surface and a outlet neck 36 which contains a
seal 38 in this case an O-ring on its outer surface. The outlet
neck 36 fits into and seals against the inner surface of the device
outlet 38 of the lower housing piece 40 of device.
[0067] FIG. 4 shows the entire device in an exploded view. Those
parts already describe din Figure retain the same numbering as in
FIG. 3. In addition to the disks 30, outlet plate 34 (shown as
mounted in the device outlet 38) there is also a body 41 and an
upper housing piece 42. Also shown are two vents/drains 44A and
44B. The device inlet is shown as 46. The upper housing piece 42,
body 41 and lower housing piece 40 are all sealed together to form
a leak proof housing. If metal, one can weld the sections together.
Preferably they are all made of plastic and solvent bonded, heat
bonded or plastic welded together. While the housing is shown as
three pieces, it could easily be made of more or less pieces
depending upon one's mold design.
[0068] FIG. 5 shows a disk 50 used in the embodiment of FIG. 4. The
inner rim 52 is formed adjacent a central opening 54 which serves
as the outlet of the disk and a inner sealing rim 33 discussed in
relation to FIG. 3. The outer rim 56 and the inner rim 52 have a
flat area 57 and 58 that serve as a sealing point for the
membrane(s). Also shown are a series of radial ribs 60 and
concentric ribs 62 which serve to support the membrane(s) and to
act as channels between the membrane(s) and the opening 54 for the
fluid that has passed through the membranes. Also shown are the
lugs 32 as discussed in relation to FIG. 3.
[0069] FIG. 6 shows a partial cross-section of a device of FIG. 4
with the fluid flow paths. Fluid is fed through the inlet (not
shown) to the outside of the disks 50. Fluid enters the membrane(s)
64 sealed on each side of the disks 50. It passes along the
channels formed by the ribs 60 and 62 to a flow window 66 adjacent
the central opening 54 and from there it exits the device through
the outlet 38.
[0070] FIG. 7 shows an even closer cross-section of the flow path
through one disk of the device of FIG. 4. The flow windows 66 can
be more clearly seen and they allow for the fluid to flow
unhindered to the central opening 54.
[0071] In a preferred embodiment, a 3.25 inch (82.55 mm) diameter
MILLIDISK.RTM. device having 6 disks, each loaded was loaded with 8
layers of 0.65 hydrophilic charged DURAPORE.RTM. membrane on each
side of each disk yielded a device having 0.045 m.sup.2 frontal
area and 0.045L bed volume.
[0072] In another embodiment, a 3.25 inch (82.55 mm) diameter
MILLIDISK.RTM. device having 60 disks, each loaded was loaded with
8 layers of 0.65 hydrophilic charged DURAPORE.RTM. membrane on each
side of each disk yielded a device having 0.45 m.sup.2 frontal area
and 0.45L bed volume.
[0073] Other formats of devices useful in the present invention
include pleated flat sheet cartridges, especially those having two
or more layers of membrane, tangential flow cassettes such as are
shown in U.S. Pat. Nos. 5,147,542, 5,176,828, 5,824,217 and
5,922,200 and which are commercially available as PROSTAK.RTM.,
Pellicon.RTM., Pellicon II .RTM. and Pellicon XL.RTM. cassettes
from Millipore Corporation of Bedford, Mass. and spiral wound
cartridges such as those taught by U.S. Pat. No. 5,128,037 and
available as HELICON.RTM. cartridges from Millipore Corporation of
Bedford, Mass.
[0074] The components of the device such as end caps, inlets,
outlets, housings, disks, etc., can be made of a variety of
materials, such as metal, ceramic, glass or plastic. Preferably,
the components are formed of metal such as stainless steel,
especially 316 stainless steel or aluminum due to their relatively
low cost and good chemical stability or more preferably, plastics,
such as polyolefins, especially polyethylene and polypropylene,
homopolymers or copolymers, and ethylene vinyl acetate (EVA)
copolymers; polycarbonates; styrenes; PTFE resin; thermoplastic
perfluorinated polymers such PFA; PVDF; nylons and other
polyamides; PET and blends of any of the above.
[0075] A method for determining the integrity of a membrane and/or
a device containing one or more membranes is also part of this
invention and is as follows:
[0076] The membrane or device is challenged with an adsorbing
solute. The challenge buffer and solute concentration are selected
such that the binding of the solute to the membrane follows
Langmuir adsorption and the solute concentration is sufficient to
be in the non-linear portion of the adsorption isotherm. The reason
for operating in the non-linear portion of the isotherm is that
favorable adsorption thermodynamics will enhance the sharpness of
the solute breakthrough (to increase the test sensitivity it is
important that the breakthrough be as sharp as possible). The
output of the test would be the breakthrough time at a
predetermined low level of breakthrough (typically less than 10%,
preferably from about 1 to about 5%) and the broadness of the
breakthrough front, (typically measured by the time of 5%
breakthrough to 95% breakthrough relative to 50% breakthrough).
This test is capable of detecting a defect in one layer of an eight
layer stacked disk device as described herein. For integral
membranes and device the low level break through is extremely
sharp. When defects are present, premature breakthrough occurs,
thereby decreasing the low level breakthrough time and in some
instances adversely affecting the broadness of the breakthrough
front. By plotting the data one can see if a defect is present.
[0077] FIG. 8 shows examples of the plots obtained by the present
integrity test using an eight layered stacked disk arrangement
described above with the membranes of the present invention. A
challenge solution consisting of 50 micrograms/ml tosyl glutamic
acid in 2.5 mMTris buffer at a pH of 8.0. Curve 100 shows the
breakthrough curve on an integral membrane. Note the sharp and
continuous solute breakthrough occurring at an onset time of
approximately 160 seconds. Curve 102 shows an eight-layered device
with a defect intentionally induced into the top membrane layer. As
seen in 102, premature breakthrough of the tosyl glutamic acid is
observed, occurring at a breakthrough time of approximately 120
seconds. This premature breakthrough occurs because the
ion-exchange capacity of certain flow paths within the membrane
adsorber become exhausted earlier than other flow paths (namely,
the flow paths that traverse only 7 membrane layers versus the flow
paths that traverse all 8 membrane layers). The remainder of the
adsorption bed becomes exhausted at the 160 second time frame. It
should be noted that the sensitivity for detecting the presence of
small defects can be enhanced by
[0078] (a) using membranes with more uniform flow properties
(membranes with high Pe numbers), which can be obtained, for
example, by using membranes with very narrow pore size
distributions like the Durapore membrane. This is the preferred
method since the membrane essentially determines the maximum
sharpness that is attainable for a breakthrough curve study. The
methods described below can only minimize the decrease in
observable breakthrough curve sharpness that occurs when the
membrane adsorber is placed into a device.
[0079] (b) Change the solute challenge conditions such that the
solute adsorption is further in the non-linear portion of the
adsorption isotherm (for an adsorption isotherm that behave
Langmuir kinetics) (i.e., increase the solute concentration).
[0080] (c) Change the solute and detection system such that lower
solute concentrations can be more easily detected.
[0081] (d) Design devices that have better flow distribution
properties
EXAMPLES
Example 1
[0082] Tosyl glutamic acid breakthrough curves were measured on
three different membrane adsorbers--a 3.5 cm.sup.2 device made of
8-layers of a positively charged 0.65 .mu.m Durapore membrane
(labeled Invention) and two other membrane adsorbers that are
commercially available (labeled Membrane X containing 60 layers of
membrane and Membrane Y containing 3 layers of membrane). All
membrane adsorbers were tested in housings designed to have good
flow distribution properties. The devices were first flushed with
DI water to completely wet the devices and to eliminate any
potential entrapped air that may negatively influence the results.
The membrane adsorbers were then flushed with approximately 20 mL
of 2.5 mM Tris buffer, pH 8.0. After buffer equilibration, the
membrane adsorbers were then challenged with tosyl glutamic acid at
a concentration of 50 .mu.g/ml in 2.5 mM Tris, pH 8.0. The
resulting breakthrough curves are shown in FIG. 9. As seen in the
Figure, the membrane adsorber of the present invention exhibited an
extremely sharp breakthrough, with a Pe number of approximately
4000 (as measured by equation (1) above). Membrane X and membrane Y
exhibited much more diffuse breakthrough curves, with measured Pe
numbers on the order of 100 and 20, respectively. These data
indicate that the flow properties inherent with the Durapore
membrane are much better than the other tested membrane adsorbers.
This has two important implications. First, it is expected that the
removal of trace impurities would be more robust with the Durapore
membrane adsorber. In fact, virus removal data (presented in
Example 2) show that this membrane adsorber is capable of removing
over 5 LRV of virus at residence times of less than 1 second.
Second, the presence of defects that may negatively influence the
retention of trace impurities are much more easy to detect in the
Durapore membrane adsorber. Thus, the Durapore membrane adsorber is
more amenable to being integrity tested, significantly aiding in
virus validation exercises.
Example 2
[0083] Four different 3.5 cm.sup.2 membrane adsorbers made of
8-layers of a positively charged 0.65 .mu.m Durapore membrane were
challenged with .phi.X-174, a 28 nm diameter bacteriophage.
Experimental results (not shown) indicate that this bacteriophage
is an excellent marker for understanding the capabilities of
membrane adsorbers for removing trace levels of mammalian virus.
The devices were first flushed with Dl water to completely wet the
devices and to eliminate any potential entrapped air that may
negatively influence the results. The membrane adsorbers were then
flushed with approximately 20 mL of 25 mM Tris buffer, pH 8.0.
After buffer equilibration, the membrane adsorbers were then
challenged with 300 mL of 1.5.times.10.sup.7 pfu/mL .phi.X-174 in
25 mM Tris, pH 8.0. Each of the four devices was challenged at a
different flow rate, (either 10, 20, 40, or 60 mL/min). After
challenging with the bacteriophage, samples of the feed and
effluent pool were assayed for bacteriophage concentration.
Finally, bacteriophage LRV values were calculated as log.sub.10
(feed concentration/effluent concentration). The bacteriophage LRV
is plotted as a function of challenge linear velocity in FIG. 10.
As seen in the Figure, virus removal is consistently greater than 5
LRV, a result that is essentially independent of flow rate. It
should be noted that a flow rate of 60 mL/min (linear velocity of
approximately 1050 cm/hr), the residence time of the solution
within the membrane adsorber is on the order of 0.35 seconds. These
data indicate that minimal mass transfer and kinetic limitations
exist which may negatively impact virus removal. These data also
indicate that the dispersion properties inherent with this
particular membrane are extremely good.
Example 3
[0084] Tosyl glutamic acid breakthrough curves were measured on
five different membrane adsorbers comprised of various numbers of
layers of a positively charged 0.65 .mu.m Durapore membrane (1, 3,
5, 7, and 8 layers). The membrane adsorbers were first flushed with
DI water to completely wet the devices and to eliminate any
potential entrapped air that may negatively influence the results.
The membrane adsorbers were then flushed with approximately 20 mL
of 2.5 mM Tris buffer, pH 8.0. After buffer equilibration, the
membrane adsorbers were then challenged with tosyl glutamic acid at
a concentration of 50 .mu.g/ml in 2.5 mM Tris, pH 8.0. The membrane
adsorber Pe numbers were calculated based upon 10%, 50%, and 90%
breakthrough times (as described previously). The data are
tabulated below. As seen in the accompanying table, all of the
measured Pe numbers were extremely high, indicating very good flow
distribution properties of this membrane adsorber. This is further
highlighted in the fact that the Pe number for a 1 layer device was
greater than 500. Based on these data, it is expected that this
membrane would be an ideal candidate for use as a membrane adsorber
for trace impurity removal.
1 Number of Membrane Layers Pe Number 1 580 3 >10,000 5
>10,000 7 >10,000 8 >10,000
Example 4
[0085] Seven different 3.5 cm.sup.2 membrane adsorbers made with
differing number of membrane layers of a positively charged 0.65
.mu.m Durapore membrane were challenged with .phi.X-174, a 28 nm
diameter bacteriophage. The devices were first flushed with DI
water to completely wet the devices and to eliminate any potential
entrapped air that may negatively influence the results. The
membrane adsorbers were then flushed with approximately 20 mL of 25
mM Tris buffer, pH 8.0. After buffer equilibration, the membrane
adsorbers were then challenged with 300 mL of 1.5.times.10.sup.7
pfu/mL .phi.X-174 in 25 mM Tris, pH 8.0 at a flow rate of 20
mL/min. After challenging with the bacteriophage, samples of the
feed and effluent pool were assayed for bacteriophage
concentration. Finally, bacteriophage LRV values were calculated as
log.sub.10 (feed concentration/effluent concentration). The
bacteriophage LRV is plotted as a function of number of membrane
adsorber layers in FIG. 11. As seen in the Figure, for devices that
contain greater than 3 membrane layers, virus removal is
consistently greater than 5 LRV, a result which can be attributable
to the high membrane Pe number.
* * * * *