CONTINUOUS FLOW REACTOR

Abstract

A continuous flow reactor, a method of performing a continuous flow reaction, and a method of controlling a moveable wall of a reaction chamber of a continuous flow reactor. The reactor comprising: an inlet; an outlet; and a reaction chamber, between the inlet and the outlet and providing a flow path therebetween, the reaction chamber having a moveable wall; the reactor further comprising: a pressure sensor configured to monitor a fluid pressure in the continuous flow reactor; and a controller, operable to adjust the position of the moveable wall, and thereby change a volume of the reaction chamber, based on the monitored fluid pressure.

Claims

1. A continuous flow reactor comprising: an inlet; an outlet; and a reaction chamber, between the inlet and the outlet and providing a flow path therebetween, the reaction chamber having a moveable wall; the reactor further comprising: a pressure sensor, configured to monitor a differential fluid pressure in the continuous flow reactor between the inlet and the outlet; and a controller, operable to adjust the position of the moveable wall, and thereby change a volume of the reaction chamber, based on the monitored fluid pressure.

2. The continuous flow reactor of claim 1, wherein the moveable wall is connected to a drive system including a motor and a lead screw.

3. The continuous flow reactor of claim 2, wherein the drive mechanism includes a stop, configured to provide a minimum volume of the reaction chamber and/or a maximum volume of the reaction chamber.

4. (canceled)

5. The continuous flow reactor of claim 1, wherein the moveable wall includes a frit which at least partially defines a wall of the reaction chamber.

6. The continuous flow reactor of claim 5, wherein the moveable wall includes a fluid pathway disposed therein fluidly connecting the reaction chamber to the outlet.

7. The continuous flow reactor of claim 4, wherein the controller is programmable with a range of pressure changes relative to the set-point over which it will not adjust the position of the sidewall.

8. (canceled)

9. The continuous flow reactor of claim 1, wherein the reaction chamber includes a solid support for use in a flow reaction.

10. (canceled)

11. The continuous flow reactor of claim 1, wherein the pressure sensor includes a first pressure sensor arranged to measure a pressure at the inlet and a second pressure sensor arranged to measure a pressure at the outlet.

12. The continuous flow reactor of claim 1, wherein the pressure sensor monitors a differential pressure between the inlet and the outlet.

13. The continuous flow reactor of claim 1, wherein the controller is programmed with an asymmetric sensitivity to the monitored fluid pressure, such that it adjusts the position of the sidewall faster in response to a change in the monitored fluid pressure in one direction to another.

14. (canceled)

15. The continuous flow reactor of claim 1, wherein the controller is programmable with a value for a maximum volume of the reaction chamber; and is operable to modify the value for the maximum volume of the reaction chamber based on a stage of a continuous flow reaction.

16. (canceled)

17. A method of performing a continuous flow reaction, comprising: supplying a fluid, via an inlet, into a reaction chamber; extracting the fluid from the reaction chamber, via an outlet; monitoring a differential pressure of the fluid within the continuous flow reactor between the inlet and the outlet; and adjusting a volume of the reaction chamber, based on the monitored fluid pressure between the inlet and the outlet.

18. (canceled)

19. The method of claim 17, including the step of: defining a set-point for the monitored pressure and a range of pressure changes relative to the set-point over which the volume of the reaction chamber will not be adjusted.

20. (canceled)

21. The method of claim 17, wherein the volume of the reaction chamber is adjusted with an asymmetric speed, such that when the monitored fluid pressure changes in one direction the volume is changed at a different rate to when the monitored fluid pressure changes in another direction.

22. (canceled)

23. The method of claim 17, comprising the steps of: (a) supplying an amino acid containing fluid into the reaction chamber, the reaction chamber containing a solid support such that the amino acid attaches to the solid support; (b) supplying a reagent into the reaction chamber, the reagent acting to deprotect the amino acid; and (c) supplying a further amino acid containing fluid into the reaction chamber, the further amino acid coupling with the amino acid attached to the solid support.

24-26. (canceled)

27. The method of claim 23, including a step of: continuously recording the volume of the reaction chamber, and calculating therefrom an indication of an extent of coupling.

28. The method of 23, including the steps of: setting a value for a maximum volume of the reaction chamber; and modifying the value for the maximum volume of the reaction chamber based on the stage of the continuous flow reaction.

29-30. (canceled)

31. A method of controlling a moveable wall of a reaction chamber of a continuous flow reactor, the method comprising: monitoring a differential pressure of a fluid within the continuous flow reactor between the inlet and the outlet; adjusting the position of the moveable wall, to thereby change a reaction volume of the reaction chamber, based on the monitored pressure.

32. (canceled)

33. The method of claim 31, including the steps of: defining a set-point for the monitored pressure and a range of pressure changes relative to the set-point over which the volume of the reaction chamber will not be adjusted.

34. (canceled)

35. The method of claim 31, wherein the volume of the reaction chamber is adjusted with an asymmetric speed, when the monitored fluid pressure changes in one direction, relative to another.

36-39. (canceled)

Description

BRIEF DESCRIPTION OF THE DRAWINGS

[0057] Embodiments of the invention will now be described by way of example with reference to the accompanying drawings in which:

[0058] FIG. 1 shows a cross-sectional view of a continuous flow reactor according to the an embodiment of the present invention;

[0059] FIG. 2 shows a cross-sectional view of the continuous flow reactor of FIG. 1 rotated by 90;

[0060] FIG. 3 shows an exploded view of the continuous flow reactor of FIGS. 1 and 2;

[0061] FIG. 4 shows a perspective view of a controller according to an embodiment of the invention;

[0062] FIG. 5 shows a control interface of the controller of FIG. 4;

[0063] FIG. 6 is a graph showing the reactor volume against time during an example synthesis;

[0064] FIG. 7 is an HPLC plot of the peptide resulting from the reaction shown in FIG. 6;

[0065] FIG. 8 is a graph showing the reactor volume against time during a further example synthesis;

[0066] FIG. 9 is an HPLC plot of the peptide resulting from the reaction shown in FIG. 8; and

[0067] FIG. 10 is a detailed view of the reactor volume for the coupling and deprotection of two amino acids during an example synthesis.

DETAILED DESCRIPTION AND FURTHER OPTIONAL FEATURES

[0068] Embodiments of the present invention are described below. Where optional features are set out, these are applicable singly or in any combination with any aspect of the invention.

[0069] FIG. 1 shows a continuous flow reactor 100 according to a first embodiment of the present invention. The reactor includes a feed stream 101 for a first fluid, for example an activated amino acid, and a feed stream 102 for a second fluid which may be a solution used for deprotection of amino acids. The join between the two feed streams 101 and 102 includes a pressure connector 103 for connecting the join to a pressure sensor (not shown) for measuring a fluid pressure upstream of a reaction chamber 104. Generally the join between the two feed streams is considered the inlet of the reactor.

[0070] The reaction chamber is partially defined by a glass column 104 which is contained within a heat exchanger 105. The glass column includes a solid support 115 for use in solid phase peptide synthesis, for example 150 mg of Chlorotrityl chloride resin.

[0071] At one end of the reaction chamber is a moveable wall 106 which, in FIG. 1, is a frit connected to the end of a plunger 107. The frit, sometimes called a fritted glass filter, is a ceramic composition used to filter solid particles from a fluid flow. The frit could equally be made of stainless steel or polytetrafluoroethylene. The plunger contains a channel 108 along its length, so that fluid can flow through the frit 106 and to an outlet. The plunger is threaded, so as to engage a correspondingly threaded drive nut or lead screw 109. The threaded drive nut 109 is connected to and driven by a drive motor 110 which thereby alters the volume of the reaction chamber. The drive motor 110 is connected to a pulley (in this example, an AT3 pulley: 18 tooth, 10 mm wide) which transfers rotation of a drive shaft of the motor to the threaded drive nut 109.

[0072] The drive motor may be a hybrid stepper motor, or a brushless DC motor with encoder feedback. Attached to the drive motor is an anti-rotation device 111, and a slide rod 112 for the anti-rotation device. The anti-rotation device can function as a mechanical stop for the drive motor 110, to provide a maximum volume and/or minimum volume of the reaction chamber. As an alternative to the anti-rotation device (or in addition thereto) a controller connected to the drive motor may be programmed with maximum and minimum volumes of the reaction chamber, and can act to stop the drive motor exceeding these volumes.

[0073] The outlet, provided at the end of the channel 108 in the plunger is connected to a fluid outlet 113 and a pressure connector 114 for connecting to outlet to a pressure sensor (not shown) for measuring a fluid pressure downstream of the reaction chamber 104.

[0074] The drive motor 110 is controllable by the controller which may be external or internal to the continuous flow reactor 100, and is connected to the pressure connectors 103 and 114. In response to the pressure sensed from the pressure connectors, the controller can operate the drive motor so as to increase or decrease a reaction volume of the reaction chamber 104 by rotating the lead screw 109 and thereby moving the plunger 107 and frit 106 up or down.

[0075] FIG. 2 shows the continuous flow reactor 100 shown in FIG. 1 as rotated by 90 about its length (vertical axis). Visible in this view is an inlet 202 and outlet 203 of the heat exchanger 105. The heater exchanger also includes a connector 204 for a temperature sensor 205, for sensing the temperature of fluid provided to the heater exchanger as it passes over the reaction chamber. This sensed temperature can be fed back to the controller.

[0076] FIG. 3 shows an exploded view of the continuous flow reactor 100 discussed above. Like components are indicated by like reference numerals.

[0077] FIG. 4 illustrates a controller 300 according to a further embodiment of the invention. The controller includes a display 17, and buttons for inputting commands to the controller. A gauge pressure manifold 15 allows connection of pressure connectors 104 and 114, thus the controller can monitor the pressure downstream and upstream of the reaction chamber and is able to calculate a differential pressure between the two. The pressure connectors preferably include ceramic fluid contact surfaces. As an alternative, a single differential pressure sensor can be provided which directly measures a differential pressure between the inlet and outlet of the continuous flow reactor. Also connected to the controller is cable bundle 16 which connects to the drive motor 110.

[0078] FIG. 5 shows an example display 17 of the controller. The display includes a status indicator 401, for informing the user whether the controller is operating or not. A target differential pressure 402 is also displayed, together with a current setting of the dead-band 403 discussed above (i.e. the region of pressure changes over which the controller will not alter the volume of the reaction chamber). The target differential pressure is the differential pressure that the controller is adjusting the volume of the reaction chamber in order to achieve. As discussed above, this is usually set within the range 0.1 bar to 15 bar. The actual current position 404 of the moveable wall is shown (in mm), which is measured relative to a starting position (in this example 0.1 mm from the initial starting position). The maximum allowable plunger position is shown at 405, and the minimum at 406. Again these are measured relative to an initial starting position. The positions may be displayed a resolution of 1/10.sup.th of a millimetre, but can be controlled to within 1/100.sup.th of a millimetre. The minimum value may be set so as to prevent the reactor contents from being compressed continually should the current target differential pressure not be achievable (for example in the case where no fluid is flowing or the flowrate is excessively low). The current measured differential pressure is shown at 407.

[0079] As discussed above, the maximum allowable plunger position may be modified so as to track with the actual position experience during coupling during peptide synthesis. In this way the reaction chamber can be allowed to expand as the peptide grows but never excessively during a wash cycle at a high flowrate.

[0080] In an example of a reaction according to an embodiment of the present invention and using the continuous flow reactor of FIGS. 1-3, residues 65-74 of the acyl carrier protein (ACP, H2N-VQAAIDYING-CONH2, ref: R. B. Merrifield, J. Am. Chem. Soc., 1963, 85 (14), 2149-2154) were synthesised. This peptide is well-known as a challenging sequence due to aggregation phenomena, and is commonly used to evaluate the performance of a synthesis platform. Below the ACP synthesis is used to show how the reactor volume changes during synthesis and then using this example the peptide is synthesised using two different methods to show how the data from the compression controller can be used to predict problematic sequences in any peptide synthesis.

[0081] The 10 mer peptide acyl carrier protein (ACP, H2N-VQAAIDYING-CONH2) is synthesised using a continuous flow reactor of the above embodiments using 400 mg Fmoc-Gly-Rink amide resin. It is assumed that the Fmoc-Gly-Rink amide resin has a loading of 0.8 mmol/g. The fully protected peptide ACP has molecular weight of 1879.96 g/mol. Therefore, assuming no losses, after synthesis and before cleavage the 0.4 g of resin will have a mass of 0.4+(1879.960.40.8/1000)=1.002 g. It is clear that the reactor must change in volume to accommodate the additional 0.602 g of mass unless the density of the packed bed is to increase markedly. In reality, when this peptide is synthesised the volume of the reactor is observed to increase by a factor of 2.5 (see FIG. 6). For the example shown in FIG. 6 a solvent modification was implemented for the final two couplings to prevent aggregation of the peptide. FIG. 8 shows the same synthesis but without the solvent modification.

[0082] FIG. 6 shows the data derived from the reactor controller when synthesising the 10-mer peptide ACP using 400 mg of Fmoc-Gly-Rink amide resin. The first 7 couplings were completed using 100% dimethylformamide (DMF) as the solvent. The final two couplings were completed using a 1:1 mixture of DMF and dimethyl sulfoxide (DMSO). The change in the solvent was necessary to prevent aggregation of the peptide. The data from the reactor controller shows a continuous growth for the reactor during all 9 couplings and the corresponding HPLC of the crude peptide (FIG. 7) indicates purity of 90%.

[0083] FIG. 7 shows the data derived from the reactor controller when synthesising the 10-mer peptide ACP using 200 mg of Fmoc-Gly-Rink amide resin. All 9 couplings were completed using 100% dimethylformamide (DMF) as the solvent. The data from the reactor controller shows a continuous growth for the reactor during the first 8 couplings but during the final coupling (Valine) at 137 minutes the volume of the reactor is observed to reduce rapidly. This reduction is volume can be attributed to aggregation of the peptide. This type of aggregation is known to hinder the efficiency of coupling. The HPLC trace (FIG. 9) shows a significant peak at 7.055 minutes which is consistent with a proportion of the peptide without the Valine coupled. The crude peptide indicates purity of 80%. The contrast between the data in FIG. 6 and FIG. 8 clearly shows capability of the reactor controller to monitor the coupling and indicate the exact time when a peptide synthesis has become problematic.

[0084] FIG. 10 is a graph showing the reactor volume logged by the controller in the ACP synthesis for the coupling and deprotection of two of the amino acids, Isoleucine and Tyrosine. Also shown is the UV absorption recorded (dotted line) at 312 nm by a UV cell located immediately downstream of the reactor. The flow rates of solvent/reagent through each coupling and deprotection step was identical. The flowrates for Isoleucine coupling and deprotection are detailed as follows:

Time 35.8 min to 45.1 min, coupling at 1.4 ml/min
Time 45.1 min to 50.4 min, post coupling wash at 3 ml/min
Time 50.4 min to 54.4 min, deprotection at 2 ml/min
Time 54.4 min to 59.9 min, post deprotection wash at 2 ml/min

[0085] The coupling, wash steps and deprotection are at different flowrates which is why the reactor volume changes rapidly immediately after coupling is complete. It is important to note that the two amino acids have very different height/time profiles. At a given flowrate, the profile during coupling is a combination of a number of parameters; molecular weight of the protected amino acid, amount of swelling of the resin by a particular amino acid, rate of coupling, rate of loss of the peptide fragment from the resin (this can occur with some resins and is not desirable). The data obtained during the coupling stage is most valuable in determining the extent and speed of coupling but also in identifying changes in the structure of the peptide.

[0086] While the invention has been described in conjunction with the exemplary embodiments described above, many equivalent modifications and variations will be apparent to those skilled in the art when given this disclosure. Accordingly, the exemplary embodiments of the invention set forth above are considered to be illustrative and not limiting. Various changes to the described embodiments may be made without departing from the spirit and scope of the invention.

[0087] All references referred to above are hereby incorporated by reference.

LIST OF FEATURES

[0088] 100 Continuous flow reactor [0089] 101, 102 Feed stream [0090] 103, 114 Pressure connector [0091] 105 Heat exchanger [0092] 106 Moveable wall [0093] 107 Plunger [0094] 108 Channel [0095] 109 Lead screw [0096] 110 Drive motor [0097] 111 Anti-rotation device [0098] 112 Slide rod [0099] 113 Fluid outlet [0100] 115 Solid support [0101] 202 Heat exchanger fluid inlet [0102] 203 Heat exchanger fluid outlet [0103] 204 Temperature connector [0104] 205 Temperature sensor [0105] 300 Controller [0106] 17 Display [0107] 15 Pressure manifold [0108] 16 Cable bundle [0109] 401 Status indicator [0110] 402 Target pressure [0111] 403 Dead-band setting [0112] 404 Current plunger position [0113] 405 Maximum plunger position [0114] 406 Minimum plunger position [0115] 407 Current measured pressure