BACKGROUND OF THE INVENTION
[0001] Field flow fractionation is a versatile technique for the high resolution separation
of a wide variety of particulates suspended in a fluid medium. The particulates include
macromolecules in the
105 to the 10
13 molecular weight (O.DO1 to 1 µm) range, colloids, particles, micelles, organelles
and the like. The technique is more explicitly described in U.S. Patent No. 3,449,938,
issued June 17, 1969 to John C. Giddings and U.S. Patent No. 3,523,610, issued August
11, 1970 to Edward M. Purcell and Howard C. Berg.
[0002] In the case of sedimentation field flow fractionation (SFFF), use is made of a centrifuge
to establish a force field for separating the particulates. For this purpose a long,
thin annular belt-like channel is made to rotate about its axis in a centrifuge. The
resultant centrifugal force causes components of higher density than the mobile phase
flowing in the channel to sediment toward the outer wall of the channel. For equal
particulate.density, because of higher diffusion rates, smaller particulates will
accumulate in a thicker layer against the outer wall than will larger particulates.
On the average, therefore, larger particulates are forced closer to the outer wall.
[0003] If now the'mobile phase or solvent is fed continuously from one end of the channel,
it carries the sample components through the channel for later detection'.at the outlet
of the channel. Because of the shape of the laminar velocity profile of the solvent
within the channel and the placement of particulates in that profile, solvent flow
causes smaller particulates to elute first, followed by a continuous elution of components
in the order of ascending particulate mass.
[0004] In order to reduce the separation times required using this technique, it is necessary
to make the channels relatively thin. This creates many problems in that the walls
of the channel must have a microscopically smooth finish to prevent the particulates
from sticking to the walls or being trapped in crevices of the same height as the
particulate distributed. Unfortunately, in the construction of such a thin belt-like
channel for use in a centrifuge, the microfinish cannot be easily obtained or maintained.
Another problem is that, even though the walls of the channel have a microfinish,
the particulates in many instances tend to adhere to the walls of the channel. This
problem is noted by-Purcell et al. at column 4, line 43 who suggest it is necessary
to select for the surface of the channel wall a material to which the particle will
not be adsorbed.
[0005] It would be desirable to obtain a channel whose outer wall has a removeable or replaceable
surface that can be changed for cleaning or according to the particulates being separated
to reduce adhesion of the particulates to the walls. Furthermore, it would be desirable
to provide a channel in which the walls need not be finished to such microscopic surface
in the first instance thereby resulting in lower cost channels.
[0006] To facilitate cleaning, the channels are usually formed of two mating rings. This
creates a problem in that leaks can occur at the interface between the rings.
SUMMARY OF THE INVENTION
[0007] According to one aspect of this invention, an apparatus is constructed for separating
particulates suspended in a fluid medium according to their effective masses. This
apparatus includes an annular, cylindrical channel having a cylinder axis, means for
rotating the channel about the axis, means for passing the fluid medium circumferentially
through the channel, and means for introducing the particulates into the medium for
passage through the channel. The channel is defined by an outer support ring having
a constant inner radius, and a unitary inner ring mating with the outer ring to define
the channel therebetween.
[0008] This channel is improved according to this invention by constructing the channel
to have a replaceable, thin film disposed between the rings and in contact with the
outer ring forming the outer channel wall. This greatly facilitates cleaning of the
channel. With this construction, the outer support ring need not have a microfinish;
rather it can rely simply upon the film to provide such finish. The film is particularly
useful in that it aids in sealing the channel to prevent leakage at the interface
between the rings.
[0009] The particular film may be selected according to the particulates to be separated,
the objective being to prevent the particles from adhering to the film surface of
the outer wall. The outer ring or wall may be formed either by a separate ring or
by the inner wall of a conventional zonal rotor centrifuge. The inner ring may be
continuous or split. The film may be coated with a material or its surface may be
modified using known techniques to reduce adhesion of the particulates to the outer
support ring. Typically, the film may be a polymeric material that is flat, smooth,
of a desired surface chemically to reduce particulate adhesion, resilient, insoluble
in the solvents normally used, and noncorroding.
BRIEF DESCRIPTION OF THE DRAWINGS
[0010] Further advantages and features of this invention will become apparent upon the following
description wherein:
FIG. 1 is an exploded pictorial representation of mating split rings that may be used
to form a sedimentation field flow fractionation channel utilizing a film interfaced
between the rings;
FIG. 2 is a fragmentary cross sectional view of a channel formed in accordance with
this invention utilizing the film insert wherein the outer ring constitutes the inner
wall of a zonal rotor centrifuge;
FIG. 3 is a cross-sectional elevation view of a continuous ring channel, utilizing
the film insert of this invention, constructed to be submerged in a fluid.
DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT
[0011] Typical sedimentation field flow fractionation apparatus with which this invention
may find use are described in U.S. Patent 4,283,276 issued August 11, 1981 to John
Wallace Grant, U.S. Patent 4,353
;795 schedule to issue October 12, 1982, copending application Serial No. 326, 156,
filed November 30, 1981 and entitled .Apparatus and Method for Sedimentation Field
Flow Fractionation" by Dilks et al. (IP-0304) and copending application Serial No.
326,157, filed November 30, 1981 and entitled "Sedimentation Field Flow Fractionation
Channel and Method" by Dilks et al. (IP-0313).
[0012] As is described in the Grant patent, split ring, SFFF channels 10 (FIG. 2) are constructed
to have an outer ring 12, which is in the form of a band having a constant inner radius
and functions to support an inner ring 13. Actually, the outer ring 12 may be supported
by a spider, bowl, or disc which is driven directly by a rotor 14 acting through a
linkage depicted by the dashed line 16. The Romanauskus patent describes one such
mounting in which the outer ring is supported by compression washers which follow
the expansion of the ring during centrifugation. Alternatively, the outer ring may
be eliminated and a bowl type rotor substituted. In this event, the bowl rotor has
a cylindrical inner surface formed thereon to provide the outer channel wall.
[0013] The inner ring 13 is split, i.e., its longitudinal circumference is divided or separated
to have a gap 18 with the-longitudinal ends 20 of the inner ring 13 slightly tapered
to facilitate the use of wedges 22. The wedges 22 retain the inner ring sufficiently
expanded to maintain contact with the outer ring 12 at all times even when stopped.
The radially outer wall 24 of the inner ring 12 and the radially inner wall 26 of
the outer ring 56 are formed to define the channel 10. Normally these walls are formed
as by polishing, for example, or by coating the surfaces with a suitable material
to have a microfinish. This smooth finish tends to reduce the possibility that particles
will stick to the walls or become entrapped in small crevices or depressions and also
insures that the expected sample retention takes place.
[0014] Depending upon the needs of the operation, a groove 28 may be formed in the outer
wall 24 of the inner ring 13 form the flow channel 10. Along the edges of the channel
groove 28, subsidiary grooves 30 may be formed to accommodate a resilient seal 32,
such as an 0-ring, which completely surrounds and tracks along the edges of the channel,
including the end sections. Additionally, the upper edge of the inner ring is formed
with a radial, outwardly extending flange 34, as is seen most clearly in FIG. 2, such
that the inner ring may rest upon and be supported by the outer ring against axially
downward displacement. Inlet and outlet conduits 36 communicate with the ends of the
channel 10 through the inner ring. As is known, fluids containing particulate samples
are passed through these conduits. The outlet conduit is coupled to a detector (not
shown). As noted, the thickness of this channel 10 is relatively small, typically
being in the order of 0.1 cm or less. The dimensions of the channel, both width and
thickness must be very precisely maintained. The actual thickness is selected according
to the separations to be performed as is known.
[0015] In accordance with this invention, a thin, flat film 75 of a resilient, smooth surfaced
material is positioned in-between the inner and outer rings 13, 12. This film typically
is placed over the inner ring, the ring compressed (the wedges 22 being removed) and
placed within the outer ring (which of course may be the inside wall of a bowl rotor)
and the inner ring allowed to release to expand and force the film in contact with
the outer wall. The wedges are then reinserted to stabilize the inner ring as described
by Grant. This film, which may be of any polymeric material or even aluminum or stainless
steel, provides the smooth surface for the outer wall without the need for the normal
polishing that would normally be required. The film extends beyond the 0-ring seals
to aid in sealing to prevent leakage of fluids from the channel. In fact, in many
cases the seals may be omitted when a resilient film'insert is used.
[0016] The films that may be used should be flat, have a smooth surface and the desired
surface chemical characteristics, be resilient, insoluble in the mobile phase used,
noncorroding, and desirably colored for ease of handling. Among the films that may
be used are cellulose, polyethylene terephthalate, polytetrafluoroethylene, aromatic
polyimide, polypropylene, polyvinyl acetates, and polyvinyl propionate polyesters.
These tend to provide hydrophobic surfaces. If hydrophilic surfaces are desired, the
above noted materials in many cases can be treated or coated. Film thickness typically
should exceed 0.005 cm. Thinner films than this have a tendency to wrinkle, which
is highly undesirable for this purpose. Films up to .02 cm thickness have been used
successfully. Films significantly in excess of this thickness may be used but tend
to be unnecessarily wasteful of the film material. Films having a thickness in the
range of .01 cm are particularly desirable. They do not change the dimension of the
channel's thickness, unless the channel is formed by a groove in the outer ring, since
they simply serve to coat the inner wall of the outer ring. Metal films that may be
used include aluminum and stainless steel which may be gold plated if desired.
[0017] In SFFF separations, samples of particulates in suspension are introduced into a
mobile phase which carries the particulates through the channel 10. The particulate
under test will determine the mobile phase and the type of film 75 that is used. Adhesion
between the channel outer wall and the particulates is to be avoided--it reduces the
resolution and sensitivity of the separation.
[0018] In accordance with this invention, such -adhesion can be avoided by properly selecting
or coating or modifying the surface of a film. Adhesion usually occurs where the surface
charge of the particulate and the film's surface polarity are opposite. It also occurs
where there is covalent bonding, chemical adsorption, or van der Waals forces. Ideally
the channel should "look like" the mobile phase, i.e., be of the same polarity.
[0019] It also is important that the mobile phase density be less than the particulate density
and that the mobile phase wets the particulates. If the particulates are hydrophobic,
the mobile phase should be nonaqueous. If the particulates are hydrophilic, the mobile
phase should be aqueous. Although of secondary importance, it is desirable that the
mobile phase wets the channel walls-In the event a hydrophilic surface is desired,
many of the above noted polymeric films which are hydrophobic, may be surface modified.
This typically is accomplished by forming OH or carboxyl groups on-the surface as
by chemically bonding desired molecules to reactive sites on the surface. Alternatively
the hydrocarbons may be oxidized by a corona discharge, the fluorocarbons may be reduced
by sodium in ammonia, esters, amides and imides may be hydrolyzed. Cellulose and polyvinyl
alcohol have hydrophilic surfaces.
[0020] The replaceable film of this invention finds use with any double ring or other type
SFFF channel which can be opened for cleaning and the like. For example, it may be
used in the "floating" type channels described in the copending Dilks and Yau and
Dilks, Yau and Kirkland applications mentioned above. These channels are formed with
a continuous inner ring as is the Romanauskus channel. Dilks and Yau describe a disklike
channel 10 as seen in FIG. 3. This channel 10 is formed by an inner ring or hub 76
and a mating outer ring 80 positioned in the bowl of a rotor 60. The hub 76 and outer
ring 80 are formed to have a diametrical interference fit of about 0.03 centimeters
(cm) so that the outer ring 80 is in constant compressive contact with the hub 76
under static conditions. The inner or mating surface 82 of the outer ring 80 has a
channel or groove 84 formed therein leaving lands 81 on either side of the groove
84. The outside portions 85 of the inner surface 82 are removed to limit the axial
width of the lands 81 and thereby enhance their ability to seal the channel when they
contact the peripheral surface of the hub 76. This groove 84 may be formed to have
different thicknesses, different widths, different lengths, different aspect ratios
(width to thickness ratio).
[0021] In accordance with this invention, a strip of film 75, as described, is positioned
against the inner wall of the outer ring 80. This is accomplished by coooling th einner
ring, wrapping the film about the inner ring, inserting the inner ring 76 into the
outer ring 80 while the film is maintained under tension, and allowing the inner ring
to expand. Air may be removed from the inner wall of the outer ring by forming it
with a matte finish. When liquid is later introduced into the channel 10, it will
expand the film into the groove 84 to define the channel.
[0022] The beginning and end of each channel and the manner in which fluids are fed to and
withdrawn therefrom are preferably those described in U.S. Patent 4,284,498 issued
to Grant et al. on August 18, 1981, the disclosures of which is incorporated herein
by reference. Fluids are fed through tubing 74 to circumferentially spaced radial
bores 83 in the hub 76 to the beginning and end of the channel 10. The beginning and
end of the channel groove 84 is defined by a plastic shim (not shown) having a close
fit with the channel axial width. The shim has inverted V-shaped ends with the apex
of the V slotted to encompass the respective bores 83. The shim may be formed of a
polyphenylene oxide plastic and be cemented into position. It may be slightly thicker
than the depth of the channel groove 84. Thus, when it is compressed by the smooth
outer peripheral surface of the hub 76, it seals and defines the beginning and end
of the channel 10.
[0023] The interior of the bowl-type rotor 60 preferably is filled with a liquid of approximately
the same density as the fluid medium that is forced to flow through the channel. Further,
the outer ring 80 is formed to have a diameter slightly less than the interior diameter
of the bowl 60 so that it does not contact the inside of the bowl even during centrifugation.
On the other hand, the hub 76 is configured so that it fits concentrically over the
interior hub 94 of the rotor 60, so as to be mounted securely thereon, and to have
a nib 96 that engages a receptacle 98 in the cover 70 to center the channel housing
76, 80. The mid-portion 100 of the hub 76 may be in the form of.an annulus having
a reduced thickness to facilitate the radially outward expansion of the hub 76 during
centrifugation to facilitate its following the outer ring expansion.
[0024] Liquid, typically water or other aqueous based liquid, thus surrounds essentially
all of the channel housing 76, 80. Under these conditions, when the rotor 60 is rotated,
centrifugal force causes the liquid pressure exerted by the liquid in the rotor bowl
60, external to the channel, and that exerted internally by the fluid medium within
the channel to be substantially equal. Since, leakage is essentially eliminated at
the interface 81 between the hub and outer ring the width of the film may be selected
to match the axial width of the groove 84. In this manner it finds use in reducing
particulate adherence to the outer channel wall and the ability to remove and replace
films according to the particulates to be separated.
[0025] As described by Dilks et al., the hub 76 and outer ring 80 preferably are each constructed
of a suitable engineering plastic selected such that the effective density φ to tensile
modulus E ratio of the outer ring is somewhat less than the effective density ϕ to
tensile modulus E ratio of the hub. The effective density φ is the density of the
channel material minus the density of the bowl filling liquid. The effective density
φ of course can be negative. This is done so that the hub can expand outwardly to
a greater extent than the outer ring to maintain a good contact, during centrifugation,
with -the-outer ring and thereby maintain the integrity of the channel. In addition,
if the density of the outer ring is less than that of the hub, the density of the
compensating liquid can be selected to be different from the density of the fluid
medium and to lie between the densities of the hub and outer ring. When the compensating
liquid density exceeds that of the outer ring, the outer ring will literally float
under a force field and be forced to have closer contract with the inner ring.
[0026] If the density of the outer ring is greater than that of the inner ring, then the
use is limited to compensation liquid densities less than that of the hub or else
the hub can separate from the outer ring under some operating conditions. With the
effective density ϕ to tensile modulus E ratio of the outer ring less than the effective
density φ to tensile modulus E ratio of the hub, the hub will expand under centrifugal
force at a faster rate than the outer ring and maintain good contact therebetween
during centrifugal operation. Preferably, the density of the compensating liquid within
the rotor is selected to be approximately equal to the density of the outer ring such
that there is little expansion or contraction of the outer ring due to the effects
of the liquid.
[0027] There has thus been described a relatively simple mechanism.for reducing the expense
of preparing SFFF channels. The films may be used typically to coat the inner wall
of the outer ring of any particular channel whether the outer wall be formed by a
zonal rotor, an outer ring or whatever.
[0028] The advantage accruing from the use of this film-are many. The finish of the outer
channel wall need not require costly machining operations to obtain as microfinish-it
is provided by the smooth surface of the film. The film is easily replaceable for
cleaning the channel, to provide surface characteristics matching those of the particulates,
end to reduce channel leakage.
1. In an apparatus for separating particulates suspended in a fluid medium according
to their effective masses, said apparatus having an annular cylindrical channel with
a cylinder axis, means for rotating said channel about said axis, and means for passing
said fluid medium circumferentially through said channel, the channel having an outer
support ring with a constant inner radius, and unitary inner ring mating with the
outer ring to define the channel therebetween, the improvement wherein:
a replaceable thin film is disposed between said rings and in contact with the outer
ring, thereby to provide a replaceable outer wall for said channel.
2. The apparatus of claim 1 wherein said ; inner ring has an outer wall the middle
portion of which defines a circumferential groove of constant depth to form the channel.
3. The apparatus of claim 1 wherein said inner ring is separated at one point along
its circumference and the channel begins and ends on either side of the separation.
4. The apparatus of claim 1, 2 or 3 wherein the film is a resilient,-smooth material.
5. The apparatus of claim 1, 2 or 3 wherein the film is coated with a material that
reduces adhesion of the particulates to the outer support ring.
6. The apparatus of claim 1, 2 or 3 wherein said film has a surface modified chemically
to reduce the adhesion of the particulates to the outer support ring.
7. The apparatus of claims 1, 2 or 3 wherein the film is a polymeric material selected
from the group consisting of polyester, polypropylene, polyethylene, polyvinylacetate,
polyvinyl propionate, polyoxymethylene, polyethylene terephthalate, tetrafluoroethylene,
and aromatic polyimide.
8. The apparatus of claim 1 wherein the film is a resilient, smooth, polymeric material,
thereby to reduce leakage from the channel through the interstitial space between
the rings.
9. The apparatus of claim 8 wherein the surface of the film is modified to be hydrophilic.
10. In a channel for separating particulates by sedimentation field flow fractionation
comprising an outer support ring with a constant inner radius, a unitary inner ring
mating with the outer ring to define the channel therebetween, the improvement wherein
a replaceable thin film is disposed between said rings and in contact with the outer
ring, thereby to provide a replaceable outer wall for said channel.
11. The channel of claim 10 wherein the film -is a resilient, smooth, polymeric material,
thereby to reduce leakage from the channel through the interstitial space between
the rings.
12. The channel of claims 10 or 11 wherein said film has a surface modified chemically
to reduce the adhesion of the particulates to the outer support ring.