Mixing fluid components

Abstract

Portions of fluid components are mixed during transport through a tube by continuously disturbing their flow pattern in a controlled manner during their passage through the tube. The disturbance is created by providing the tube with a radially inward profile which in a preferred form is an internal screw-thread of a thickness 0.25 to 0.75 times the internal diameter of the tube and having a pitch 0.75 to 1.5 times the internal diameter of the tube.

Description

This invention relates to a method and apparatus for mixing batches of fluid components during transport in particular for use in a continuous flow analysis system, in which the batches may be separated by air bubbles.

In 1957, Technicon Corporation developed a continuous flow analysis system, in which a stream of material is divided by air bubbles into segments in which chemical reactions occur. In this system, continuous streams of material, containing samples, reagents, etc., are combined and during transport mixed in mixing coils. The continuously flowing stream is passed through tubing from one apparatus to another, each apparatus performing a different analytical function, such as purifying, heating, incubation, qualitative determination, and recording.

An essential principle of the analysis system is the introduction of air bubbles in the stream of liquid. The air bubbles provide a barrier between liquid segments as they flow through the system, to prevent cross-contamination, they clean the system in that they wipe along the walls of conduits, and also assist in mixing combined streams of material in the mixing coils.

In this technique of analysis, it is a requirement for the streams of material, which contain samples, reagents and the like, and are commonly liquids, to be very intimately mixed. Mixing has hitherto been accomplished by running the combined streams through a horizontally disposed glass mixing coil, it being assumed that mixing is effected due to the repeated inversions of the liquid segments separated by air bubbles (see the Technicon Corporation brochure entitled "Automation in the Clinical Laboratory" (1957), page 7, lines 28- 29).

The glass mixing-coil proposed by Technicon Corporation should have a sufficient number of windings to ensure proper mixing. The necessarily resulting great path length of the mixing tube has been found to be a drawback in this technique, because interaction between the liquid and the tube wall causes transfer of liquid from one liquid segment to the next. This results in material disadvantages, for example, tailing, cross-contamination, and poor separating capacity.

Another disadvantage is that a coiled mixing tube made of an inert plastics, for example, polytetrafluoroethylene, results in poorer mixing.

It is an object of the present invention to provide a method of mixing batches of fluid components during transport, whereby the above drawbacks and disadvantages are overcome.

It is another important object of the invention to provide a method of intimately mixing batches of fluid components during transport in a continuous flow analysis system, using a relatively short length of tubing, thereby to minimize the time required for analysis, and to minimize the amounts of material and reagents required.

It is still another object of the invention to provide apparatus for carrying out the said method.

According to one aspect of the present invention, there is provided a method of mixing fluid components as they are transported through a tube, which comprises providing continuous controlled disturbances in the flow pattern of said components during their passage through said tube.

According to another aspect of the present invention, there is provided a mixing tube for use in mixing fluid components together, said tube having an inner wall surface provided with a radial profile.

In a preferred form, the mixing tube according to the invention is provided with an internal screwthread. To attain proper mixing, the and pitch of the screwthread should have a particular ratio to the diameter of the tube, namely, they should be about 0.5 and about 0.9 to about 1.1 times the tube diameter respectively. It will be clear, for that matter, that the ranges will be determined in part by the material properties of the liquid and the tube wall.

For use in an analysis system, it is of great importance for the corners between the profile and the inner surface of the tube to be uniformly rounded to prevent material being processed from remaining behind in possible "dead corners," which is apt to cause tailing and cross-contamination. Also, turbulence should be avoided by adapting the diameter of the tube to the amount of material to be processed.

The mixing tube according to the invention can be made in full or in part from a synthetic plastics material, and the profile in the tube can be provided in various ways.

In one embodiment the mixing tube according to the invention is made of glass and has a glass enamel coated Kovar wire cemented to its inner surface (Kovar is a registered tradename (Westinghouse) used to denote certain alloys of cobalt, nickel and iron).

The use of a mixing tube according to the invention results in excellent, substantially transverse mixing. The fluid components are mixed in a small unit of volume, and it is no longer strictly necessary to include air bubbles within a sample, since tailing and cross-contamination are considerably reduced, and a better separation is obtained as compared with the prior art.

Another very important advantage over the prior mixing coil is that the mixing tube path length through which the liquid has to travel is shortened by a factor of about 20, as a result of which the following further advantages are obtained.

The greatly shortened analysis path length decreases the time of analysis required, which is important in sito determinations and for the development of enzyme analysis apparatus. Also, less material and less reagent is required, which is of importance for micro-analysis.

The mixing tube of the present invention provides a better signal-to-noise ratio, which results in greater sensitivity. The consequence is that a stable baseline is more rapidly reached. Furthermore, the mixing tube is easier to handle, less vulnerable, and smaller, which opens up possibilities of miniaturizing the existing analysis system.

In the accompanying drawings,

FIGS. 1, 15, 21, 28, 35 and 36 are schematic illustrations of test arrangements used for comparing the performance of a mixing tube according to the invention with that of a mixing coil as hitherto used.

FIGS. 2-14, 16-20, 22-27, 29-34, and 37-48 are reproductions of the graphic results of tests run with the arrangement of FIGS. 1, 15, 21, 28, 35 and 36.

FIG. 49 is a magnification of a schematic cross-sectional view of one exemplary embodiment of a mixing tube according to the invention.

FIG. 49a is a schematic view of a plastic mixing tube.

EXAMPLE I

Mixing liquids of equal and unequal viscosities and specific gravities

In order to properly compare the mixing capacity of a mixing tube (MT) according to the invention and the mixing coil (MS) of the prior art, use was made of a measuring arrangement as shown schematically in the accompanying FIG. 1. As shown, the arrangement comprises:

Proportioning pump 1 with two flexible tubes a and b, whose internal diameters (ID) are indicated in mm in the drawing. With this diameter the rate of transport of the liquid was 0.80 cm.sup.3 /min.

Mixing tube 2 (MT, length 4.5 cm) or mixing coil 2 (MS, length 70 cm).

Colorimeter 3, using a cuvette of 15 mm and a wavelength of 660nm.

Recorder 4, recording the extinction on logarithmic paper moving at a rate of 45.7 cm/hour.

Test I, 1

Mixing liquids of different specific gravities

A solution of 1 cm.sup.3 thymol blue (0.1 g/50 cm.sup.3 ; 0.1 N KOH) in 500 cm.sup.3 0.001 N KOH, pumped through tube a, and different NaCl solutions in distilled water, of 0 M, 0.1 M, 0.5 M, 2.0 M and 5.0 M, respectively, pumped through tube b, were mixed by means of the mixing tube (MT) and then for purposes of comparison by means of the mixing coil (MS).

The extinctions measured are shown in FIGS. 2-11, the noise caused by disturbances due to differences in optical density being a measure for the mixing capacity of the mixing tube and the mixing coil, respectively.

______________________________________ Figures Concentration of the mixed Mixing means NaCl solution (mol.) used ______________________________________ 2 0 MT 3 0 MS 4 0.1 MT 5 0.1 MS 6 0.5 MT 7 0.5 MS 8 2.0 MT 9 2.0 MS 10 5.0 MT ______________________________________

It appears from the noise in FIGS. 2-7 that in mixed NaCl solutions of 0-0.5 M the mixing capacity of the mixing tube corresponded with that of the mixing coil.

It appears from the noise in FIGS. 8-11 that when the NaCl solution of 2 M was mixed the mixing capacity of the mixing tube was somewhat greater and that when the NaCl solution of 5 M was mixed the mixing capacity of the mixing tube was clearly greater than that of the mixing coil.

In this connection it should again be observed that the ratio of MT path length to MS path length was 4.5/70.

Test I, 2

Mixing of liquids of different specific gravities and strongly to very strongly differing viscosities at 20.degree.C.

Distilled water (specific gravity 1; viscosity 1), pumped through flexible tube a, was mixed with 1,2-ethane diol (specific gravity 1.12; viscosity 19.9), pumped through flexible tube b, by means of the mixing tube and afterwards by means of the mixing coil for the sake of comparison.

It clearly appears from FIGS. 12 and 13 and particularly from the noise therein that fluids of different viscosities were mixed considerably better in the mixing tube than in the mixing coil.

This better mixing was obtained although the path length was reduced from 70 cm to 4.5 cm.

Test I, 3

Distilled water (specific gravity 1, viscosity 1), pumped through flexible tube a was mixed with 1,2,3-propane triol (specific gravity 1.26, viscosity 1499) pumped through flexible tube b by means of a mixing tube having a length of 9.5 cm. After this the mixing test was repeated. using a mixing coil instead of the mixing tube.

It is apparent from FIG. 14 and particularly from the noise therein that mixing fluids with extreme differences in viscosity was very well possible by means of the mixing tube.

When the mixing coil was used it was found that the fluids remained separated.

EXAMPLE II

Mixing two fluids for continuous flow analysis, comparing the mixing properties of a mixing tube according to the invention with those of a mixing coil

The test arrangement is schematically shown in FIG. 15 and was composed of

a proportioning pump 1 with two flexible tubes whose internal diameters (ID) are shown in the drawing in mm. The diameter corresponds with a fluid transport of 0.80 cm.sup.3 /min.

a mixing tube 2 (MT, length 4.5 cm) or a mixing coil 2 (MS, length 70 cm),

a colorimeter 3, using a wavelength of 660 nm and a cuvette of 15 mm

a recorder 4 which recorded the extinction on logarithmic paper at a paper velocity of 45.7 cm/hour.

In this arrangement a coloured solution of 1 cm.sup.3 thymol blue (0.1 g/50 cm.sup.3 0.1 N KOH) was mixed by means of the mixing tube with a colourless 0.001 N HCl solution. Afterwards the test was repeated, using for comparison the mixing coil as the mixing means.

Test II, 1

The colourless solution, pumped through one flexible tube, was mixed with distilled water pumped through the other flexible tube, the resulting signal forming the baseline. Subsequently, instead of the distilled water the coloured solution was pumped. The signal rose to a specific steady state, after which instead of the coloured solution again distilled water was pumped. The signal dropped again to the baseline.

The extinction curve obtained by means of the mixing tube is shown in FIG. 16 and the curve obtained by means of the mixing coil in FIG. 17.

Under conditions in which the stable state was obtained it appears from FIGS. 16 and 17 that by means of the mixing tube a more stable signal was obtained. It is also apparent from these figures that the time to attain the steady state from the baseline and subsequently the baseline from said steady state is much shorter when the mixing tube is used than when the mixing coil is used. This time is determined by the degree of tailing.

Therefore, when the mixing tube was used considerably less tailing took place than when the mixing coil was used.

Test II, 2

The colourless HCl solution was mixed with distilled water, the resulting signal forming the baseline. Subsequently, instead of distilled water the coloured solution was pumped for 30 seconds. The signal rose to a specific value which was lower than the steady state signal level. After this, instead of the coloured solution again distilled water was pumped for 30 seconds. The signal dropped to a specific value which was higher than the baseline.

By repetition a pattern of peaks and troughs was formed.

FIG. 18 shows the extinction obtained by mixing means of the mixing coil, FIG. 19 shows the extinction obtained by mixing by means of the mixing tube and FIG. 20 shows the corresponding steady state.

It can be seen from FIGS. 18 and 19 that the tailing which took place in Test 1 influenced the distinquishability of the samples. When the mixing coil is used, the peak levels 1, 2, 3 and 4 (FIG. 18) differ strongly. However, when the mixing tube is used it appears that only peak level 1 was somewhat lower than the level of peaks 2, 3 and 4 (FIG. 19). Besides the difference in level between the peaks and the troughs is much greater when the mixing tube is used than when the mixing coil is used.

Therefore, the mixing tube had a separating capacity far superior to the mixing coil.

EXAMPLE III

Mixing for phosphate determination

The analysis was based on the formation of a phosphomolybdate complex when orthophosphate is added to molybdic acid. The complex was then reduced with a mixture of ascorbic acid (vitamin C), acetone and water to molybdenum blue (absorption at 660 nm).

Literature:

- Strickland, J. D. H., Parsons T. R., A practical Handbook of Seawater Analysis, Bull 167, Fish Res. Bd. Canada, 1968.

- Bradshaw J. S., Spanis C. W., Advances in automated Analysis, Thurman Ass. 1971, pp, 17.

The following reagents were used:

Molybdic acid solution: 8.5 g ammonium molybdate (Merck) was dissolved in 250 cm.sup.3 distilled water. Then 100 cm.sup.3 concentrated H.sub.2 SO.sub.4 was added, cooled and made up with distilled water to 1000 cm.sup.3.

Ascorbic acid solution (vitamin C): 5 g ascorbic acid (Baker Chemicals) was dissolved in 125 cm.sup.3 acetone. Then the volume was made up with distilled water to 250 cm.sup.3.

Before use 20 cm.sup.3 of this starting solution was made up with distilled water to 100 cm.sup.3.

Phosphate solutions: 0.068 g KH.sub.2 PO.sub.4 (Baker Chemicals) was dissolved in 1000 cm.sup.3 distilled water.

The arrangements are schematically shown in FIGS. 21 and 28 and were composed of proportioning pump 1 with flexible tubes whose internal diameters (ID) are shown in mm in these drawings. In the following table the pumped volumes corresponding with the internal diameters are shown:

pumped material ID in mm rates in cm.sup.3 /min. ______________________________________ air 0.045 0.80 phosphate 0.056 1.20 molybdate 0.030 0.32 Vitamin C 0.030 0.32 to waste 0.056 1.20 ______________________________________

A mixing tube 2 (MT) according to the invention with a length of 4.5 cm and for comparison a mixing coil 2 (MS) with a length of 70 cm;

a colorimeter 3 using a wavelength of 660 nm and a cuvette of 15 mm;

a recorder 4 with which the measured extinction was recorded on logarithmic paper at a paper velocity of 45.7 cm/hour.

Tests in which the fluid flow was divided into fluid segments by air (FIG. 21)

Test III. 1

First the reagents and distilled water were pumped as a sample. The resulting signal formed the baseline. Subsequently the phosphate solution was pumped as a sample, during which the signal rose to the steady state. After this again distilled water was pumped as a sample, during which the signal dropped again to the baseline.

FIG. 22 shows the result obtained by means of the mixing coil and FIG. 23 shows the result obtained by means of the mixing tube. It is apparent from these drawings that with the mixing tube a somewhat more stable signal was obtained than with the mixing coil. The mixing capacity of the mixing tube was therefore somewhat superior with 1/14th of the length of the mixing coil. The tailings were comparable.

Test III, 2

First the reagents and distilled water were pumped as a sample. The resulting signal formed the baseline. Subsequently phosphate solution was pumped as a sample for 60 seconds, during which the signal rose to a specific level (lower than the steady state). After this again distilled water was pumped as a sample, during which the signal dropped to a specific value (higher than the baseline). By repetition a pattern of peaks and troughs was obtained, shown for the mixing coil in FIG. 24 and for the mixing tube in FIG. 25.

The mixing properties of the mixing tube and of the mixing coil were comparable, but then their lengths are greatly different.

Test III, 3

This test was analogous to test 2, but sampling took place for 30 seconds instead of 60 seconds.

The result obtained with the mixing coil is shown in FIG. 26 and with the mixing tube in FIG. 27. The mixing properties of the mixing coil and the mixing tube appeared to be substantially the same.

Test in which the fluid flow was not divided into fluid segments by air (FIG. 28).

All tests were analogous to tests III, 1-3, but no air was pumped.

Test III, 4

Analogous to test III, 1. The result obtained with the mixing coil is shown in FIG. 29 and with the mixing tube in FIG. 30. It is apparent from these drawings that the mixing tube produced somewhat more noise than the mixing coil under conditions in which the steady state was obtained. The time necessary to reach the steady state from the baseline and from this state again the baseline was much shorter in the mixing tube than in the mixing coil. This time is determined by the degree of tailing, so that it appears that mixing with the mixing tube involves considerably less tailing than with the mixing coil.

Test III, 5

Analogous to test III, 2. The result obtained with the mixing coil is shown in FIG. 31 and with the mixing tube in FIG. 32. It appears from these drawings that the tailing found in test III, 4 is reflected in this case in the distinguishability of the samples. The difference in level of peaks and troughs was much greater with the mixing tube than with the mixing coil. The mixing tube therefore had a separating capacity far superior to the mixing coil.

Test III, 6

Analogous to test III, 3. The result obtained with the mixing coil is shown in FIG. 33 and with the mixing tube in FIG. 34. FIG. 33 was composed from four samples which were separated by a wash. It appears that the separate samples could no longer be distinguished.

In FIG. 33, which is analogous to FIG. 34, the samples can be distinguished.

Summarizing it can be stated that in actual practice the mixing tube produced a better result.

EXAMPLE IV

Mixing in the determination of hemoglobin in blood

In this series of tests the erythrocytes in the blood were lysed, after which the released hemoglobin was oxidized with potassium ferric cyanide at a pH 7.0 - 7.4 to hemiglobin. The hemiglobin was converted with potassium cyanide to the stable cyano hemiglobin. The extinction of the resulting solution was determined colorimetrically at 540 nm.

Literature:

Henry, R. J.; Clinical Chemistry,

Harper & Row, Publishres 742 (1964).

The following reagents were used:

200 mg K.sub.3 Fe(CN).sub.6 (Merck), 50 mg KCN (B.D.H.), 140 mg KH.sub.2 PO.sub.4 (Baker Chemicals) and 0.5 cm.sup.3 sterox-SE (Lamers & Indemans) were dissolved in 100 cm.sup.3 distilled water. 420 cm.sup.3 venous blood (A-positive) was collected in sterile and pyrogen-free conditions in 80 cm.sup.3 of a solution which contained disodium citrate (2.7%) and glucose (2.3%).

The blood suspension thus obtained was diluted 25 times with a physiological saline solution (0.9% NaCl), and the dilute suspension enabled proper measurement of the hemoglobin concentration.

The test arrangements are schematically shown in FIGS. 35 and 36 and were composed of a proportioning pump 1 with flexible tubes whose internal diameters (ID) are shown in mm. The pumped volumes corresponding with the diameters are listed in the following table.

______________________________________ rates in pumped material ID in mm cm.sup.3 /min. ______________________________________ air 0.030 0.32 cyanide 0.056 1.20 blood 0.020 0.16 ______________________________________

a mixing tube 2 (MT, length 4.5 cm) or mixing coil 2 (MS, length 70 cm)

a colorimeter 3, using a wavelength of 540 nm and a cuvette with

a length of 15 mm;

a recorder 4, with which the measured extinction was recorded on logaritmic paper at a paper velocity of 45.7 cm/hour.

Tests in which the fluid flow was divided into fluid segments by air (FIG. 35)

Test IV, 1

First reagents and distilled water were pumped as a sample. The resulting signal formed the baseline. Subsequently blood was pumped as a sample, during which the signal rose to a specific level namely the steady state, after which again distilled water was pumped as a sample, which caused the signal to drop again to the baseline.

FIG. 36 shows the result obtained with the mixing coil and FIG. 37 the result with the mixing tube. It appears from these figures that with the mixing tube a somewhat more stable signal was obtained than with the mixing coil. Therefore, the mixing tube had a somewhat superior mixing capacity at only 1/14th of the length.

Test IV, 2

First reagents and distilled water were pumped as a sample. The resulting signal formed the baseline. Subsequently blood was pumped as samples for 60 seconds, during which the signals rose to a specific level (lower than the steady state), after which water was again pumped. The signal then dropped to a specific value (higher than the baseline). By repetition a pattern of peaks and troughs was formed.

FIG. 38 shows the result obtained with the mixing coil and FIG. 39 the result with the mixing tube. It is apparent from the drawings that the separating power of the mixing tube was better than that of the mixing coil.

Test IV, 3

The test was analogous to test 2, but sampling took place for 30 seconds instead of 60 seconds.

FIG. 40 shows the result obtained with the mixing coil and FIG. 41 the result with the mixing tube. It is apparent from the drawings that the mixing properties of the mixing tube were much better than those of the mixing coil. The difference in level between the peaks and the troughs was much greater with the mixing tube than with the mixing coil.

Furthermore cross-contamination was much greater with the mixing coil than with the mixing tube.

Tests in which the fluid flow was not divided into segments by air (FIG. 42). 42)

All tests are analogous to tests IV, 1-3 but no air was pumped.

Test IV, 4

Analogous to test IV, 1. FIG. 43 shows the result obtained with the mixing coil and FIG. 44 the result with the mixing tube. It is apparent from these drawings that the mixing tube produced somewhat more noise than the mixing coil under conditions in which the stable state was obtained. The time necessary to reach the stable state from the baseline and from this state again the baseline was much shorter with the mixing tube than with the mixing coil. Therefore, in the mixing tube considerably less tailing occurred than in the mixing coil.

Test IV, 5

Analogous to test IV, 2. FIG. 45 shows the result obtained with the mixing coil and FIG. 46 the result with the mixing tube. The difference in level between the peaks and the troughs was much greater with the mixing tube than with the mixing coil. The mixing tube therefore had a separating power far superior to that of the mixing coil.

Test IV, 6

Analogous to test IV, 3. FIG. 47 shows the result obtained with the mixing coil and FIG. 48 with the mixing tube.

FIG. 47 was composed of 4 samples which were separated by a wash. No separate samples could be observed any longer.

FIG. 48 was composed analogously to FIG. 47. In this case it was possible to distinguish the samples separately.

Summarizing it can be stated that the mixing tube produces a better result in actual practice, although the path length is reduced from 70 cm to 4.5 cm.

Referring now to FIG. 49, there is shown a mixing tube according to the invention, which comprises a glass tube 5 provided with a profile on its inner surface in the form of a helical glass-enamel coated Kovar wire 6 cemented to the tube with glass enamel 7, applied to provide rounded surfaces 8.

In this example the height h of the profile, i.e. the thickness of the coated Kovar wire 6 is equal to half the diameter d of tube 5. The pitch p of the helix is equal to diameter d.

It will be understood that we may use other means to provide the profile on the inner surface of the mixing tube and that, although only one embodiment has been described and shown, our invention is not so limited.

Claims

We claim:

1. A mixing tube for use in transversely mixing fluid components together, said tube having an inner wall surface provided with a rounded male screw-thread configuration that provides a helical bore therethrough and having a height of about 0.5 times the diameter of the tube, and a pitch of from about 0.9 to about 1.1 times the diameter of the tube.

2. A mixing tube as recited in claim 1 wherein angles formed by a profile of said male screw-thread configuration are uniformly rounded.

3. A mixing tube as claimed in claim 1, wherein said tube is made at least in part of synthetic plastics material inert to materials to be processed.

4. A mixing tube as claimed in claim 1, wherein said tube is made at least in part of polytetrafluoroethylene.