DE2125027A1 - Device for separating aqueous solutions from suspensions - Google Patents

Description

concerning

Device for separating aqueous solutions from suspensions.

The invention relates to the treatment of relatively small volumes of aqueous solvents, which contain soluble and reactive substances in relatively low concentrations. She relates also to a device which is available at low cost for separating a part of the solution from an aqueous one Suspension of insoluble reaction products formed in an aqueous medium for analysis of the same. The device according to the invention is suitable for insulation .of aqueous media in relatively small volumes for the purpose of investigation, e.g. colorimetric or radioactive investigation of hormones, carbohydrates, enzymes, etc.

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The device according to the invention is a closed arrangement with several liquid-containing zones in which aqueous media are kept separate from one another as long as atmospheric pressure is maintained. the The arrangement consists of a first container made of a suitable material, such as glass, a polymer made from unsaturated hydrocarbons etc. and one with this first container

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type related second container that liquid can pass from one to the other. The first container is an open top and bottom end Röhr- f Chen, whose upper end can be closed by a removable cover. In the communication path for liquid between the two containers, a waterproof film with pores averaging 5 to 10 microns is arranged, which delays the flow of aqueous media from one to the other container until the tension between the pores and the liquid through one on the pressure acting on the aqueous medium inside the device is overcome.

Substances present in very low concentrations can be used on the device according to the invention, for example with the aid of radioimmunological or staining methods, e.g. hormones such as insulin, antibodies, enzymes such as glucose, vitamins and the like, which are present in small volumes of liquid are to be detected and measured. The fluids can come from the human body, such as urine, Sputum, blood plasma, cerebrospinal fluid and the like, or they can be liquid carriers for vitamins, such as vegetable fats or liquids from other sources.

Typical experimental equipment for carrying out the- ' like investigations, e.g. for the detection of insulin previously made from insulin iodized with iodine 125, a binding

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means for insulin and filter discs. The implementation the investigation requires pipettes, syringes, filter holders for the Pyrex microanalysis, test tubes and the like.

The entire equipment must be cleaned extremely carefully for a long time to avoid the effects of contamination to prevent. The transfer of the substances to be examined from one part of the equipment to one Others with large surface areas can cause parts of the test material to adhere to the surfaces Lead to losses due to the small amount of starting material and the low. Concentration the accuracy the examinations can seriously affect.

In addition, in examinations in which the radioactivity is to be measured, a particularly intensive one must be carried out Cleaning treatment take place, and the equipment must be handled with extra care after use Avoid dangers to health.

Another problem with such tests is the loss of fluid and, as a result, of that to be tested solutes associated with the concentrated solids; such losses occur especially when the clear liquid required for the subsequent examination by decanting or pipetting the supernatant liquid takes place over a settled suspension.

It has now been found that the separation of an aqueous solution from a suspension is carried out in one device which can also be used if the examination involves health risks; the device consists of two parts, which are connected in this way

den are that after cleaning the entire investigation in

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a closed system that works as the only closed component. Since this is the If this is the case, an effective pressure can be applied to the liquid, e.g. by centrifugation Conversion of virtually all solids-free liquid can be achieved, and that liquid can be collected in the second container without the losses associated with decanting and pipetting will.

t The device according to the invention consists of a

first container of tubular type, which is open at the top and bottom, but optionally one at the top may have removable closure. A second container is connected to this first container in a liquid-tight manner, whose liquid-receiving zone is in communication with the liquid zone of the first container, however, a perforated hydrophobic film is provided between the two zones, the pore size of which is selected in this way is that water can only penetrate if there is a clear pressure difference on both sides of the film exists; normally the liquid-tight separating film blocks the communication of liquid between the two zones.

The device preferably consists of a plastic container, in which the upper part of the cylindrical side walls in the lower part of the side walls of the upper tubular container engages such that the connection is liquid-tight. A hydrophobic film is arranged in such a way that it allows the flow of liquid between the two containers under normal or atmospheric pressure blocked with the peripheral area of the film wedged in the connection between the two containers.

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Hydrophobic films vary widely in terms of their brittleness and pore size. As a precaution against tearing of more brittle films, whereby the liquid could run from one container into the other, is preferably the film with porous plastic disks sealed. Such a disk can be supported by a support ring or by a flange that has a Part of the cylindrical side walls of the container according to the invention can represent. In this case, flanges formed on the upper and on the lower container can be used can be used to hold the support members in a fixed position. The supporting disks are generally chosen so thick that that they are rigid and consist of plastics such as polypropylene, polystyrene and the like.

According to another embodiment of the device according to the invention contains the first or lower container, i.e. the receiving container for the filtered solution, a path for the Fluid communication in the form of the tip of a syringe. Such a syringe is a container, its contents against leakage through the tip into the first container is blocked by introducing a layer with a hydrophobic film in the barrel of the syringe. Such a unit is formed by making a circular disk of a hydrophobic film, the diameter of which is slightly smaller than the inner diameter of the syringe, between two rigid porous disks of the same diameter and connects the three elements at the periphery with an elastic binder. The tie layer gets so thick made that the binder makes a watertight seal on the inner wall when the unit is inserted into the syringe barrel forms so that water can only reach the outlet of the syringe after passing through the hydrophobic film. To Introducing such a unit containing a hydrophobic film into the syringe barrel is described above

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closed by introducing the usual piston mechanism. This mechanism provides a means for mechanically applying pressure to a suspension contained in the syringe, and then the aqueous solution passes through the hydrophobic film and is transferred to the lower container as described above.

A typical device used to examine substances such as insulin, amylase and the like is usually a unit 12.7 cm in length and about 1.3 cm in outer diameter, with container walls 2, A to 3, 1 are thick. For the separation of filtered liquids of larger volume, larger units can easily be produced, for example plastic pipes with an internal diameter of 7 »6 cm or more.

When using the device according to the invention for Separation of aqueous media, water and reactive substances are introduced into the upper container and the conditions selected so that the reaction products are formed and then subjects the container contents to pressure conditions which the interfacial pressure between the liquid and the membrane pores is exceeded, so that the aqueous Medium or, if insoluble reaction products were formed, an essentially solids-free aqueous solution passes through the hydrophobic film and collects in the lower container.

To carry out investigations, one brings between the first and second containers a hydrophobic film such as a tetrafluoroethylene film (trade name "Mitex") at. A corresponding amount of reagents, e.g. during an insulin test Sodium barbital buffer, hydrochloric acid, human serum

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Albumin buffer, porcine insulin, encoded insulin and insulin antibodies, introduced into the first container. After incubation, charcoal coated with dextran is added and the insulin that has not been converted into a complex is adsorbed or absorbed. The aqueous solution in the first container is then subjected to a force which forces it into the lower container. A pressure caused by centrifugal force, for example at 2,000 to 4,000 rpm, can cause the aqueous solution to run through the "Mitex" (Teflon) film into the second container, which then serves as a counting ampoule, while the solid reaction products remain in the first container. The counting ampoule is then inserted into the shaft of a crystal block. A photocell counts the scintillation caused by the emitted γ-rays that penetrate through the counting ampoule and impact the crystal material _3.

If a chromogen or indicator test is to be carried out, an indicator such as o-toluidine can be placed in the container, the walls of which are made of transparent plastic before using Eruck to force the liquid through the hydrophobic film into the lower container. The liquid then gives the indicator reaction and a comparison with standard stains proves rapid Completion of the investigation.

Typical hydrophobic films which can be used for the purposes of the invention are the continuous mat-like braids of tetrafluoroethylene polymer fibers, known under the protected trade name "Teflon", which are fused together so that they have, for example, a pore size of 5 to 10 / U; Continuous mats of acrylonitrile-polyvinyl chloride copolymers, reinforced with nylon, with a pore size of about 0.45 Ai or more are also useful. Such film material leaves an aqueous medium

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only through when the interfacial tension between filter and liquid is overcome by creating a Pressure difference, e.g. by applying overpressure to the liquid suspension in the upper container or to the lower container Allow negative pressure to act. Sintered braids made of polymer material are called hydrophobic because of their flexibility . Films preferred. Films with a pore size of 0.4-5 to 10 / U allow a liquid medium to pass if a pressure

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difference of 1.41 to 0.18 kg / cm is exerted. For clinical Work best is a film that requires 0.18 to 0.35 kg / cm of pressure to force water through, i.e. having a pore size of 5 to 10 / u. and one

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Bubble point from 0.07 to 0.04 kg / cm when blowing through Has air. The latter condition allows the omission of a valve for releasing air pressure in a lower Compartment during centrifugation or other pressure application may have built up while the liquid was being transferred to this lower compartment.

Other hydrophobic films can also be used, e.g. cellulose fiber filters such as a Whatman filter (1 PS paper) or cellulose derivatives that are mixed with plastics based on alkyl ketene dimers, have been treated to make them water-resistant on the surface (trade name "Aquapel") ·

A hydrophobic film such as a tetrafluoroethylene fiber mat (Trade name "Mitex") is an essential part the invention. The film shows the necessary tightness against liquids at atmospheric pressure, even if the. Pore size is such that solid particles of e.g. 5 / U size can penetrate. Such a hydrophobic film can be used in combination with a hydrophilic filter, such as the cellulose acetate filter available under the protected trade name "Celotate"; the latter is for Microfiltration is useful because it has pore sizes of 0.2, 0.5

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and 1 / U is available. Solids so large that they pass through the pores of the hydrophobic film, are then retained on the hydrophilic film and it collects A solution that is essentially free of solids forms in the lower container.

The invention will now be explained in more detail with reference to the drawing:

Fig. 1 shows a perspective view of a device according to the invention from a tubular upper and a tubular lower liquid container;

Fig. 2 is a longitudinal section through a unit according to Fig. 1 and shows the position of the hydrophobic Films;

Figure 5 is a partial section through a unit in which the hydrophobic film is disks is supported;

Fig. 4 is a partial section through a particular embodiment of the upper liquid container as a syringe; and

Fig. 5 is a partial section showing the hydrophobic Film layer is seen holding back liquid in the syringe before pressure is applied.

As can be seen from FIGS. 1, 2 and 3, there is the inventive Device for separating aqueous solution containing dissolved substances from a suspension of solid formed in .situ. Reaction products from tubular Parts 10 and 11 made of any suitable material,

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such as metal or malleable synthetic plastics, vie methyl methacrylate, polyethylene, polypropylene, etc. The tubular part 10 can optionally be at 12 Wear internal threads so that a cap 13 is more light-tight Completion can be screwed on.

The tubular part 10 merges into an extension Ϊ4 whose outside diameter is slightly smaller than that of the main part. The constriction forms on the pipe ^ shaped part 10 a shoulder 15.

The tube part 11 has a bottom 17 and in its upper part merges into an extension 18 with a somewhat larger one Inner diameter as the main part, so that a shoulder 19 is formed inside the tube 11.

Because of its inner diameter, the extension 18 is in sliding connection with the extension 14 of the tubular part 10 so long that the pipe part 11 rests against the shoulder 15 of the upper pipe part, so that a tight connection consists.

On the extension 14 of the pipe part 10 is a flange-like Arranged extension 20, the inner diameter of which is slightly larger than that of the main part of the tubular part 10; This creates an inner shoulder 21 in which an upper porous protective disk 22 for a flexible, hydrophobic tetrafluoroethylene film 23 intervenes.

A porous polypropylene support disk is seated on the shoulder 19 24 for the film 23, the diameter of which is practically the inner diameter of the extension 18 of the tubular part 11 is equivalent to.

When assembling the device, the flexible becomes hydrophobic

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Film 23 applied to disk 24. Then the porous disk 22 is placed over the film 23 so that when the parts 10 and 11 are plugged together, the peripheral region 25 of the film 23 between the extensions -14 and 18 of the two Pipe pieces is clamped, the shoulder 21 holding the porous disc 22 and the extension 20 on the flexible Film rests where it covers the peripheral area of disk 24 so that the film and the two disks are secure are held in place and a watertight connection is formed.

In Figs. 4 and 5, reference numeral 30 represents a Syringe, which in this case forms the upper part of a unit used for analysis. The syringe 30 has a Tip 31, through whose outflow channel 32 liquid can escape. The plunger 34 of the syringe 30 has corresponding Devices for manual operation, by means of which pressure can be exerted on the contents of the syringe. Between the piston 34 and the mouth of the liquid channel 32 is the one hydrophobic film containing device 35 is arranged.

The device 35 is shown in detail in FIG consists of a filter 36 made of "Aquapel" (a polymer Alkyl ketene dimer), which is arranged between the porous support disks 37 and 38. The filter 36 and the discs 37 and 38 are united at the periphery to form a liquid-tight unit with the aid of a suitable elastic Binders 391 like rubber putty. The binder 39 forms a in the radial direction extending thick layer, which acts as a seal and a liquid-tight connection between the periphery of the filter 36 with the disks 37 and 38 and the Vand of the syringe 30 forms. At the top is the syringe 30 is fastened with a flange 40.

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The tip 31 of the syringe 30 extends inside a lower container 41. The tubular container 4-1 is provided with a flange 42 at the upper end and its length and diameter are slightly larger than that of the syringe 30 so that it can be received therein, with the flange 42 engaging the flange 40. If necessary 'can the two flanges are attached to one another, as indicated by the brackets 43 in the drawing.

A typical analysis can be carried out in the device according to FIGS. 1 and 2 as follows:

A polypropylene tube 10 is covered by a tetrafluoroethylene film ("Mitex") with an average pore size of 5 / u of of liquid communication with "the polypropylene container 11. The periphery of the film 23 is seated on a Ledge of about 0.8 mm on and is supported by the extensions 14 and the shoulder 15, which make a liquid-tight Ensure fit by friction, held firmly.

In the raw part 10 is now an antibody, namely with 125-iodine labeled insulin and a potassium citrate buffer solution inserted and the top of the tube closed with a cap.

For the solution ± 2 pipe part 10 of one provided for comparison A standard solution is added to the unit and the unknown serum is added to the tube part 10 of another unit. The two units are then incubated in a chamber maintained at incubation temperature, e.g. 37 ° C, for 2 to 4 hours. After the incubation, charcoal coated with dextran is added and the contents of the two parts to be compared Shake 10 to make sure the mixture is uniform. The pipe parts, which are a suspension of solid

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containing substances are then placed in a centrifuge and centrifuged for 5 min at 2,000 rpm.

The clear filtrate then contains the entire antibody-antigen complex. It is placed in the container 11 in the shaft of a crystal block of a 101 gamma counter (Abbott Logic) and exposed to γ-ray irradiation. The γ-rays emitted from the sample are counted. A photocell count of 2,636 in 30 seconds on one A 1.0 cnr sample of serum containing insulin indicates 24 microunits of insulin per cnr.

A colorimetric test for serum glucose can be carried out in the device made of clear polystyrene can be carried out as follows:

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In the tubular part 11 are placed 3 cm of an o-toluidene solution and so fitted together, the two parts that a hydrophobic "Mitex" -I therebetween is ¹ ilter.

Now a sample of 0.5 cm glucose is in the pipe part 10

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Solution, 0.1 cnr barium hydroxide and 0.2 cnr zinc sulfate solution brought in and mixed well; after the mixture has been left to stand for 10 minutes, a suspension is formed of precipitated solid proteins.

The container is now centrifuged. A clinical centrifuge, which works, for example, at 2,000 to 2,500 rpm, which means that a pressure of up to about 1.76 kg / cm absolute occurs separates the aqueous solution from the precipitated Proteins. The solution is mixed with the chromophore solution, i.e. o-toluene in the pipe part 11 and 30 min left to stand at -3 C. ' The mixture is tested in a Bausch & Lomb Spectronic 20 spectrometer by

through-

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light with a wavelength of 420 to 450 m / u

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directs. A reading of 0.25 optical density indicates the presence of 30 µg glucose in the sample.

The device according to the invention described above can be modified in detail without departing from the scope of the invention To exceed. A typical modification would be to attach to the outer portion of a removable polyethylene cap, which closes the upper end of the lower container, a polystyrene tube, a tubular polystyrene part attached. An opening can be punched into the cap which is large enough to make it hydrophobic Can record film. The film can be attached in the opening with the help of an epoxy resin adhesive and through from above be protected by a porous polyethylene disc.

When such a unit was used for the colorimetric examination with the aid of a Bausch & Lomb spectrometer, which was calibrated for a passage of light of 440 and 5 ^ 5 m / u, it showed when an iodine-iodine-potassium solution was used detected through a variation in the experimental results of - 2% T between tube and _ 2% within a single unit after a rotation of 90 °. Such a fluctuation remains within the error limits in laboratory tests.

PATENT CLAIMS:

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Claims (6)

DK. IHQ- F. WITESTHO FF _ "I R - DIPL. IMG. G. PULS ^ DB. E. τ. PECHMANN DB. ING. D. BEHRENS FATKNTANWlLT« • MtNCHIN 90 PATENT CLAIMS 1) Device for separating an aqueous solution from solids suspended therein, characterized by several zones (10, 11) for taking up liquid, wherein aqueous media remain separated from one another as long as atmospheric pressure is maintained, namely by a ersoen container (10) in the form of a tube open at the top and bottom / a second container (11), eggs from the first container is separated by a perforated hydrophobic film (23) with pores, the size of which is adjusted is that water only penetrates if there is a noticeable difference in pressure on both sides of the film, wherein the film is fitted in a liquid-tight manner and the communication of liquids between the first (10) and the second (11) container blocked under normal conditions, but free when the pressure is applied accordingly gives. 2) Device according to claim 1, characterized in that the two container parts (10) and (11) are slidably nested and that the film (23) supported by porous discs (22) and (24) is, at the lower end of the first container part (10) by clamping a peripheral region (25) between the walls (14) and (18) of the nested container parts is attached. 3) Device according to claim 1 or 2, characterized in that the film (23) is a braid or fabric made of fused polymer fibers, which has an average pore size between 0.45 and 10.0 Ai . 109849/1308 "'~ ¹⁶ 4) Method according to claim 1, 2 or 3, characterized in that the hydrophobic film (23) is supported by porous discs (22, 24) and that the first container (10) at its upper end by a removable cover (13) is closed. 5) Device according to one of the preceding claims, characterized in that the hydrophobic film (23) has a pore size of about 1Q / u and that a hydrophilic filter with a maximum pore size of 1 Ai is arranged parallel to it. 6) Use of the device according to one of claims 1 to 5, characterized in that one in the first container (10) carries out a reaction in an aqueous medium in which solids precipitate and that the liquid portion of the reaction mixture using pressure through the filter (23) in. The second container (11) transferred. 10-9849 / 1308