NOVEL POROUS POLYMER MONOLITHS ADAPTED FOR SAMPLE PREPARATION

Abstract

A porous polymer monolith comprises a polymer body having macroporous through-pores that facilitate fluid flow through the body and an array of mesopores adapted to bind from the fluid flow molecules of a predetermined range of sizes, wherein the surface area of the monolith is predominantly provided by the mesopores. Also disclosed is a method of making a porous polymer monolith. The method includes forming a polymer body by phase separation out of a solution containing at least a monomer, a crosslinker and a primary porogen, whereby the body contains multiple macroporous through-pores, wherein the solution further contains a secondary porogen comprising oligomers inert with respect to the monomer and cross-linker but chemically compatible with the monomer so as to form mesostructures within the polymer body during said phase separation, and washing the mesostructures from the body to provide an array of mesopores such that the surface area of the monolith is predominantly provided by the mesopores.

Claims

1-28. (canceled)

29. A method of making a porous polymer monolith comprising: forming a polymer body by phase separation out of a solution comprising a monomer and a primary porogen, whereby the body contains multiple macroporous through-pores, wherein the solution further comprises a secondary porogen comprising oligomers inert with respect to the monomer but chemically compatible with the monomer so as to form mesostructures within the polymer body during said phase separation, wherein the secondary porogen has a molecular weight of not more than 5000, and washing the mesostructures from the body to provide an array of mesopores such that the surface area of the monolith is predominantly provided by the mesopores.

30. The method according to claim 29 wherein the secondary porogen is an oligomeric form of the monolith polymer.

31. The method according to claim 29 wherein the secondary porogen comprises styrene oligomers.

32. The method according to claim 31 wherein the styrene oligomers are structured to minimise or prevent their participation in the primary phase separation polymerisation reaction that forms the monolith body.

33. The method according to claim 30 wherein the monomer is a mix of methacrylates and the secondary porogen is a HEMA oligomer.

34. The method according to claim 29 wherein there are no or substantially no residual mesostructures formed from the secondary porogen.

35. The method according to claim 29 wherein the macoporous through-pores are substantially unmodified and unaffected by the addition of the secondary porogen during the formation of the polymer body.

36. The method according to claim 29 wherein the pore size profile of the mesopores is predetermined by the molecular size and therefore molecular weight of the secondary porogen oligomers.

37. The method according to claim 29 wherein the secondary porogen is soluble in the solution comprising the monomer and primary porogen.

38. The method according to claim 29 wherein the mesopores have a pore size in the range 20-120 Å or 40-120 Å.

39. The method according to claim 29 wherein the macroporous through-pores have a pore size in the range of 2-3 microns or 1-5 micron.

40. The method according to claim 29 wherein the mesopores contain sites adapted to bind molecules of predetermined character, structure, chemistry or size.

41. The method according to claim 29 wherein greater than 65% of the surface area of the monolith is provided by the mesopores.

42. The method according to claim 29 carried out in situ in a sample preparation or analytic device.

43. The method according to claim 29 further comprising surface grafting onto a surface of the monolith body to provide a hydrophilic external surface.

44. The method according to claim 43, wherein the surface grafting comprising grafting poly(ethyleneglycol)methylether methacrylate (PEGMA) onto the surface of the polymer body.

45. The method according to claim 44 wherein the monomer is divinyl benzene.

46. The method according to claim 29 wherein the monomer is divinyl benzene.

47. The method according to claim 29 wherein the secondary porogen has a molecular weight of not more than 2000.

48. A method for separating an analyte from a solution, the method comprising passing the solution comprising the analytye through the polymer body prepared according to the method of claim 29.

Description

[0039] The results for the synthesised monoliths were compared to corresponding results obtained with commercially available DVB solid phase extraction devices—Oasis from the Waters Corporation and Sola from Thermo Fisher Scientific.

[0040] FIG. 1 demonstrates the increased binding capacity for the increasing molecular weight analytes on the various synthesised monoliths. The monolith designated “original” was made by the standard process with no secondary porogen. As the molecular weight of the secondary porogen was increased a corresponding increase in binding capacity was observed for the increasing molecular weights of the analytes.

EXAMPLE 2

[0041] Caffeine molecular weight 194, was measured in whole human capillary blood. A calibration curve was constructed using dilutions of 0.5 mg/mL caffeine stock solution. The following concentrations were chosen: 3.125 μg/mL, 5 μg/mL, 31.25 μg/mL and 62.5 μg/mL. Calibration analysis was carried out by High Pressure Liquid Chromatography (HPLC) under the conditions listed below and gave a correlation of R.sup.2=0.9993 All caffeine concentrations were calculated comparison to the calibration curve obtained.

TABLE-US-00002 LC Conditions Column: 250 × 4.6 mm enable C18G HPLC system: Shimadzu Prominence 20A Flow rate: 1.0 ml/min Mobile phase: 16% acetonitrile with 0.1% TFA Detection: 270 nm Temperature: 25° C. Sample temp: 15° C. Injection volume: 1 μL

[0042] Whole blood samples were obtained from two volunteers. The whole blood of the first volunteer was obtained 3 hours after the volunteer had consumed a cup of coffee. The second volunteer had not consumed coffee for the past 24 hours. In each case, 150 μL of blood was lysed with 1350 μL of water. A solid phase extraction (SPE) procedure was conducted according to the following sequence of steps: [0043] Precondition cartridge using 2 mL methanol and 2 mL water [0044] Load 2 mL of samples [0045] Wash with 9 mL water [0046] Elute with 1 mL methanol [0047] HPLC analysis

[0048] A 10 μL sample was injected for HPLC analysis.

[0049] The results are shown in FIG. 2.

[0050] The caffeine concentration in the blood of volunteer 1 was found to be 2.43 μg/mL. Volunteer 2 was correctly shown to have no measurable caffeine. The healthy level is 1-10 μg/mL. The lethal level is 80 μg/mL.

Example 3

[0051] To demonstrate the utility of the hydrophilic surface functionality of a porous polymer monolith with PEGMA surface graft, a 0.5 mg/mL of 3-nitroaniline sample was loaded onto porous polymer monolith 1681 (example 1) and PEGMA grafted monolith 1681. The result showed that sample solution stayed on top of unmodified monolith 1681 but was easily absorbed onto the grafted monolith 1681 without conditioning and equilibration due to the hydrophilic surface allowing the sample to transfer through the polymer.

Example 4

[0052] To demonstrate the utility of grafting of the porous polymer monolith to prepare a mixed-mode ion exchange capability, 2-Acrylamido-2-methylpropane sulfonic acid (AMPS) was used as the functional monomer for surface grafting of porous polymer monolith 1681 (example 1). In order to demonstrate the degree of grafting on the mesoporous structure of monolith 1681, amitriptyline (pka=9.4, MW=277.4) was chosen as the target analyte to investigate the binding of different AMPS-grafted 1681 monoliths.

[0053] FIG. 3 shows the increased ion exchange functionality of the increased degree of AMPS grafting. The amount of amitriptyline loaded was 100 μg and was completely bound on all samples. The original 1681 is purely hydrophobic and all adsorbed amitriptyline could be eluted with methanol. As the amount of grafted AMPS was increased the amount of amitriptyline that was retained through ionic interactions was increased too. This fraction will not desorb with methanol but requires basic elution conditions. At 3% grafting the binding capacity for amitriptyline is equally shared between the reversed phase and the ionic mode. At 10% grafting and above, the material has turned from a mixed mode sorbent to a true ion exchanger. The high recovery results demonstrate a higher degree of grafting on the mesopores compared to micropores.