Centrifugal Particle Elutriator And Method Of Use

Abstract

A method and apparatus for carrying out centrifugal elutriation using a rotatable cylinder having an annular cavity within. Samples are introduced into the cavity at a central part thereof. A suspending liquid is introduced into the cavity at the centrifugal side. A first portion of particles within the sample moves in the centripetal direction with the flowing liquid and a second portion of larger particles moves in the centrifugal direction. Exit ports at the centripetal and centrifugal sides of the cavity provide a means for continuously removing the separated first and second portions of particles.

Description

BACKGROUND OF THE INVENTION

This invention was made in the course of, or under, a contract with the U.S. Atomic Energy Commission. It relates generally to the art of centrifugal elutriation.

The prior art in many areas of technology has used elutriation as a means of separating particles of similar densities but of different effective diameters. The process is based generally upon an application of Stokes Law of sedimentation. The process is applied to particles smaller than about 100 microns. When particle sizes are below approximately 40 microns centrifugation has been used to speed up the settling process.

In the field of blood separation centrifugation has been used in a batch type operation for separating the various constituents. One such prior art technique is that of Lindahl et al., IVA. Tidskrift for Teknisk Vetenskoplig Forskning 26, 309 (1955). This apparatus comprises a cone-shaped inclined cavity within a rotating disc, with a side loop attached to the cavity. The inclination and side loop serve the purpose of minimizing recirculation currents and thus mixing. However, even with this design recirculation currents and mixing still occur.

Another such apparatus is described by McEwen et al., Analytical Biochemistry 23, 369-377 (1968). This apparatus comprises a rotatable disc having a kite-shaped cavity in a radial portion thereof. The sample to be separated is pumped into the centrifugal side of the cavity while the disc is rotating. The slower settling particles are thus pumped out of the centripetal side of the cavity while the faster settling particles are retained within the cavity. By varying the speed of rotation, various size particles can be pumped out of the cavity one at a time.

Another prior art technique involves the use of a suspending liquid whose viscosity is varied over time. The settling rate of the particles being separated is effected by the viscosity of the suspending medium in accordance with Stokes Law.

The above prior art techniques generally utilize the concept of flow through a finite void within a rotating disc. Several inherent disadvantages result from such operation. In the above techniques the flowing contents of the cavity come into contact with radial walls during the separation process. This gives rise to Coriolis force effects which cause turbulence along the wall fronting rotation and mixing of the particles which are being separated. Another problem with the above prior art is that the fluid is introduced into the cavity at the section of smallest cross section and moves in the centripetal direction into sections of increased cross section. This causes the overall velocity to decrease as the fluid moves in the centripetal direction. It is known from the technology of fluidization that a high fluid velocity leads to a low particle concentration and, therefore, to a low suspension density. As a result, the suspension density within the cavity tends to be low at the outer radius, and to increase in the centripetal direction. Such a density configuration is unstable (like trying to suspend a layer of mercury above a layer of water in a beaker) and will lead to turnover and turbulent mixing. In addition, since laminar flow exists at the radial boundaries of the void, the central portion of the fluid has a greater velocity than that of the fluid in the laminar flow of the boundaries. This tends to cause convective mixing.

A particular problem with the above prior art processes as they are applied to blood separation, as well as to other separations, is that there is a density gradient in the particlees due to the varied velocities in the different cross section area regions of the void. This results in turbulent mixing of the particles. Another result of the radial velocity gradient is that different shear conditions exist in regions of different velocity. Blood particles are composed of aggregate particles. Under conditions of high shear the aggregates are broken up into primary particles. Under conditions of low shear the particles reaggregate and move in the centrifugal direction only to again be broken up.

In addition to the above problems which militate against achieving any separation at all, the processes are only applicable to batch operation. Thus such systems would not be amenable to a continuous process wherein a particular blood constituent is removed from the blood of a donor, and the remaining plasma and blood constituents are returned to the donor in a single continuous operation.

SUMMARY OF THE INVENTION

It is thus an object of this invention to provide a new method and apparatus for centrifugal elutriation in which the fluid does not come into contact with axially extending radial walls.

It is a further object of this invention to provide a method of centrifugal elutriation wherein shear conditions are such that there is no inversion of aggregate particles.

These and other objects are accomplished by introducing a sample to be separated into a toroidal cavity concentrically located within a rotating disc at a point intermediate the centrifugal and centripetal boundaries of the cavity, evenly flowing a suspending medium from the centrifugal boundary of the cavity toward the centripetal boundary removing faster settling solids and suspending medium from the centrifugal portion of the cavity and removing slower settling particles and suspending medium from the centripetal portion of the boundary.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a sectional oblique drawing of a rotor housing according to this invention.

FIG. 2 is a sectional view of an alternative construction of a rotor housing according to this invention.

FIGS. 3 and 4 are graphs used in determining the overall geometry of a rotor housing according to this invention.

DETAILED DESCRIPTION

According to this invention it has been found that axially extending radial walls may be completely dispensed with in a centrifugal elutriator. A cross section view of the elutriator of this invention is shown in FIG. 1. The elutriator is comprised of a housing (1) having a right toroidal cavity (6) enclosed within it. Conduit means (7), (8), (9), and (10) communicate with the cavity at various locations. Conduit means (7) serves as an introductory port for suspending fluid used in the process of this invention. Conduit means (10) serves as a sample introduction port. Conduit means (8) and (9) are respectively the centripetal and centrifugal exit ports. The conduit means are pipe-like orifices which pass from the central entrance to their respective openings in the cavity. For small centrifuges (less than about 15 centimeters in radius) a conduit means every 30.degree. or 12 conduit means for each of those illustrated in FIG. 4 is sufficient for satisfactory flow. However, any other arrangement, such as disc-shaped cavities, which allows for uniform flow may be used.

Pervious baffles (2) and (4) through which suspending medium and particles can flow are provided at the centrifugal and centripetal sides of the cavity. Outer baffle (2) is necessary for assuring that the suspending medium, which is introduced through conduit means (7), flows into the central portion of the cavity at an even and uniform velocity around the cavity. Such even flow minimizes mixing effects which would otherwise be present. Inner baffle (4) is not absolutely necessary for the successful operation of the apparatus. However, it is preferred to incorporate baffle (4) into the apparatus so as to minimize and evenly distribute any suction effects which may arise from conduit means (8).

The pervious baffles may be in the form of wire mesh or perforated sheet. It is preferred, however, that the baffles be constructed of porous material having open porosity with a size on the order of the material being separated. Porous polytetrafluoroethylene having a pore size of about 25 microns, which is commercially available, is desirable for use when blood is being separated. The baffles may also be in the form of a packed bed of beads.

Rotor (1) is of the type of construction as is conventional with centrifuges. Appropriately grooved and drilled stacked plates of stainless steel bolted at the periphery provide a suitable construction. However, when blood is being separated it is necessary for the stainless steel to be coated with an inert material such as polytetrafluoroethylene.

Conventional temperature control and rotating means may be employed. For example, the control system used in the K series centrifuge may be employed with the rotor of this invention. Such means are described by Brantley et al. in "K-Series Centrifuges," Analytical Biochemistry 36, 434-442 (1970).

An alternate form of construction is shown in FIG. 2. In this alternative embodiment, pervious baffle (2') extends only partially across cavity (6) and partition (13) intersects baffle (2') so as to define a flow path for liquid flowing through conduit means (7) through pervious baffle (2'). In this case, conduit means (9') is displaced centrifugally from pervious baffle (2') in areas where no counterflow occurs. Such an arrangement provides for a higher degree of packing of faster settling particles prior to removal.

The process of this invention is generally applicable to particles within the size range of from about 100 microns to 100 A. Although a broad spectrum of particle sizes may exist within the above range, the process of this invention is designed to divide the particles into two groups, one of which is larger than the dividing size and the other of which is smaller than the dividing size. The process of this invention is carried out by the continuous introduction of a sample comprising particles and a suspending liquid into the cavity (6) through conduit means (10) while simultaneously flowing suspending liquid through conduit means (7) while rotating housing (1) at an appropriate velocity. The particles introduced through port (10) tend to move toward centrifugal boundary (12) at varying velocities due to the centrifugal field existing within the rotating housing. Different size particles tend to settle toward centrifugal boundary (12) at different velocities generally as is predicted from Stokes Law. Suspending liquid flowing through conduit means (7) is forced through cavity (6) at a velocity which is intermediate the settling velocities of the particles introduced through conduit means (10). Thus, particles which are settling at a velocity which is greater than the velocity of suspending fluid entering through conduit means (1) move in the centrifugal direction and out through conduit means (9). Particles which are settling at a velocity which is less than the velocity of the flowing suspending liquid move in the centripetal direction and out through conduit means (8). In carrying out the process of this invention there are four flow rates which must be monitored and regulated to achieve satisfactory results. The four flow rates are as follows.

First, the flow of incoming sample through conduit means (10);

Second, the flow of suspending medium through conduit means (7);

Third, the flow of slower settling particles and suspending medium through conduit means (8); and

Fourth, the flow of rapidly settling particles and suspending medium through conduit means (9).

The first and second flow rates are determined by geometry considerations which are discussed below. The third and fourth flow rates are best determined by regulating the fourth flow rate so as to maintain the radial separation interface of the fast and slow settling particles between conduit means (10) and inner pervious baffle (4). This can be done by observing the separated products or by providing transparent windows in the rotor housing so that the interface may be observed either visually or photometrically. The interface is preferably located centrally between conduit means (10) and inner pervious baffle (4).

In carrying out the present invention the overall geometry of the apparatus must be based upon the particular fluid particle system upon which the apparatus will operate. As an example, an apparatus designed to separate while cells from 1.0 cm.sup.3 /sec. of whole blood with a typical red cell volume fraction of 0.45 is described as follows. A maximum volume of cavity (6) is stipulated as 500 cc since this is a safe volume to remove from a donor.

The sedimentation coefficient of white cells in plasma at 37.degree.C is about

S.sub.w = 12 .times. 10.sup.-.sup.8 sec.

The sedimentation coefficient of red cells depends on the degree to which individual cells combine to form large aggregates. This tendency varies from individual to individual. Shearing the blood just before sedimentation tends to break up the larger aggregates and thus to reduce the sedimentation velocity. We consider here the value

S.sub.r = 12 .times. 10.sup.-.sup.7 sec.

which should be readily attainable with blood from most individuals if severe shearing is avoided. This value allows for a moderate amount of shearing in the tubes and seals which deliver the blood to the separation apparatus.

The volume fraction of white cells (C.sub.W) in blood is normally about 0.002. This small value makes possible a convenient approximation that C.sub.W is too small to appreciably affect the sedimentation velocities of either red or white cells. ith the aid of this approximation, the plots of FIGS. 3 and 4 have been prepared to aid in determining the size of the apparatus required and the amount of flow which must be pumped through the porous or perforated baffles.

In FIG. 3, the ordinate is defined as:

red cell throughput = FC.sub.R.sup.F /.omega..sup.2 r.sub.F.sup.2 2.pi.lS.sub.R

where

F = volumetric flow rate of blood in the stream feeding the separator (in cm.sup.3 /sec.)

C.sub.r.sup.f = concentration of red cells in the stream feeding the separator (volume fraction)

r.sub.F is the radial location of conduit means (10) (cm)

l is the axial length of the separator cavity (cm)

In FIG. 4, the ordinate is defined as:

"Elutriation flow" = E/.omega..sup.2 r.sub.F.sup.2 2.pi.lS.sub.R

where

E = volumetric flow rate of plasma through pervious baffle (2) (cm.sup.3 /sec.)

In both FIGS. 3 and 4, the abscissa is the concentration of red cells in the stream feeding the separator, C.sub.R.sup.F.

Since the capacity as shown in FIG. 3 is increased by using lower feed concentrations, we dilute the whole blood to a concentration of C.sub.R.sup.F = 0.30 before introducing it to the separator. We choose a rotor speed of 700 rpm and a feed port radius of r.sub.F = 10 cm. These values are convenient and lead to a radial acceleration of about 55 times gravity which does not damage blood cells.

From FIG. 3 at C.sub.R.sup.F = 0.30

Fc.sub.r.sup.f /.omega..sup.2 r.sub.F.sup.2 2.pi.lS.sub.R = 0.076

The axial length of the cavity required is then

l = 1.9 cm.

From FIG. 4, at C.sub.R.sup.F = 0.30

E/.omega..sup.2 r.sub.F.sup.2 2.pi.lS.sub.R = 0.195

The volumetric throughput of plasma through the porous baffles is then

E = 1.16 cm.sup.3 /sec.

Typical dimensions for an elutriator under the above conditions would then be:

radius of centripetal wall (11) 6 1/2 cm.

radius of centripetal baffle (4) 7 1/2 cm.

interface of red and white cells 8 1/2 cm.

conduit means (10) 10 cm.

centrifugal baffle (2) 10 1/2 cm.

centrifugal wall (12) 11 1/2 cm., and

axial height 1.9 cm.

The above equations and approximations can, of course, be used to determine the elutriator geometry for any system in which separation is desired. The process and apparatus of this invention may also be used in more than one stage. For example, red and white cells may be separated in a first stage and plasma separated from the red and white cells in second and third stages. On the other hand, the products of the first stage may be again separated to achieve a higher separation quality. It is readily apparent that additional stages may be built into the same rotor housing by merely stacking the stages one against the other or that other separate stages may be used.

Claims

What is claimed is:

1. A method for separating particles which are suspended within a suspending liquid, comprising the steps of:

continuously introducing a sample comprising said particles and said suspending liquid into a central portion of a bound rotating cavity, a first portion of said particles having different settling velocities from a second portion of said particles within said suspending liquid;

flowing a liquid, which is the same liquid as said suspending liquid of said sample, from the centrifugal side of said cavity toward the centripetal side of said cavity at a velocity which is intermediate the settling velocities of said first and second portion of said particles, whereby the faster settling portion of said particles move in the centrifugal direction and the slower settling particles move in the centripetal direction along with said flowing liquid;

removing said faster settling particles and suspending liquid from the centrifugal side of said cavity; and

removing said slower settling particles and suspending liquid from the centripetal side of said cavity.

2. The method according to claim 1 wherein said sample is whole blood, said slower settling particles are white blood cells, said faster settling particles are red blood cells and said suspending liquid is plasma.

3. A centrifugal elutriator, comprising:

a rotor housing (1) enclosing a right toroidal cavity (6) concentrically located about an axis of said housing, said cavity having a centrifugal boundary and a centripetal boundary, said centrifugal boundary being located at a radial distance from said axis which is greater than the radial distance of said centripetal boundary from said axis.

first conduit means (7) communicating with said cavity at a point adjacent said centrifugal boundary,

second conduit means (9) communicating with said cavity,

third conduit means (10) communicating with said cavity at a point centripetally located from said second conduit means and generally centrally located on a radius of said cavity,

fourth conduit means (8) communicating with said cavity at a point centripetally located from said third conduit means and adjacent said centripetal boundary, said first, second, third, and fourth conduit means communicating with the exterior of said housing; and

a pervious baffle (2) concentric with said centrifugal and centripetal boundaries axially traversing at least a portion of said cavity and attached to said housing at a radius intermediate said first and third conduit means whereby fluid entering through said first conduit means flows through said baffle to reach any of said conduit means.

4. The apparatus according to claim 3 further comprising a second pervious baffle (4) located between said third and fourth conduit means.

5. The apparatus according to claim 3 wherein said pervious baffle extends across the entire axial height of said cavity.

6. The apparatus according to claim 3 wherein said baffle extends across only a portion of said cavity and a concentric partition extends from said pervious baffle to said centrifugal boundary, whereby said first conduit means communicates with said cavity within the space defined by said pervious baffle, said partition and said centrifugal wall and said second conduit means is located centrifugally of said pervious baffle and on the opposite side of said cavity from said first conduit means.