WO1993023356A1

WO1993023356A1 - Process for recovering 2,3-dihydroxy-1-chlorocyclohexa-4,6-diene from aqueous solutions using a solid sorbent

Info

Publication number: WO1993023356A1
Application number: PCT/US1993/004354
Authority: WO
Inventors: Antonio A. Garcia; Gregory M. Whited; Lawrence D. Kwart
Original assignee: Genencor International, Inc.
Priority date: 1992-05-12
Filing date: 1993-05-06
Publication date: 1993-11-25
Also published as: AU4239793A

Abstract

There are described processes for the removal and recovery of polar oxygenated organic compounds (e.g. 2,3-dihydroxy-1-chlorocyclohexa-4,6-diene) from aqueous solutions. Such processes comprise sorption/leaching processes whereby polar oxygenated uncharged organic compounds can be removed from aqueous solution and recovered from the sorbent.

Description

PROCESS FOR RECOVERING 2,3-DIHYDR0XY-l-CHL0R0CYCL0HEXA-4,6-DIENE FROM AQUEOUS SOLUTIONS USING A SOLID SORBENT

FIELD OF THE INVENTION:

This invention relates to recovery of polar uncharged oxygenated organic compounds from aqueous solutions using solid sorbents. Traditional methods for recovering such compounds include liquid-liquid extraction, distillation and membrane filtration, which are often inefficient and not practical on an industrial scale.

BACKGROUND OF THE INVENTION:

Several sorbents or resins have been used to remove certain types of compounds from aqueous medium, however such known resins have not successfully been employed in removal and recovery of polar uncharged compounds such as those described in the present invention. Furthermore, although some resins have been used to remove certain organic compounds from aqueous solutions, often the organic compounds are quite difficult or impossible to recover from the resin on which they are adsorbed.

US Patent 4,450,294 (Feldman) discloses a process for recovering an oxygenated organic compound from a dilute aqueous solution using a crosslinked polyvinylpyridine resin. US Patent 4,064,043 (Kollman) discloses a process for separating an organic component from a non-body liquid medium using particles of a partially pyrolyzed macroporous synthetic polymer produced by the controlled thermal degradation of a macroporous synthetic polymer containing a carbon-fixing moiety and derived from one or more ethylenically unsaturated monomer(ε) .

US Patent 4,267,055 (Neely) describes a process for removing planar molecules from a mixture containing planar and non- planar molecules using a particulate adsorbent which is the product of controlled thermal degradation of a macroporous synthetic polymer. The planar molecules removed using the described resin, may be halogenated planar molecules.

US Patent 4,BS3,596 (Agui, et al.) discloses a method for the removal of pyrogenic matter dissolved in water using a carbonaceous adsorbent prepared by carbonizing porous beads of a cross-linked polymer. The carbonaceous resin described by Agui, et al., is comparable to AmbersortP resins, commercially available from Rohm and Haas Co.

The cited art is illustrative of the use of particular resins or adsorbents to remove certain organic compounds from aqueous solutions, however, none of the references describe a useful resin or sorbent for removal and recovery of the compounds of interest in the present invention. It is an object of this invention therefore to provide acceptable processes for the recovery of certain compounds from aqueous solution such as from a fermentation broth.

SUMMARY OF THE INVENTION:

In accordance therewith, it has been discovered that the recovery of polar uncharged oxygenated organic compounds from aqueous solutions can be done using the processes described herein. The invention relates to a process for recovering polar uncharged organic oxygenated compounds from aqueous solution, the process comprising:

a) contacting the organic compound with a solid sorbent; and

b) leaching with one or more polar solvents or mixtures of polar solvents.

BRIEF DESCRIPTION OF THE DRAWINGS:

Figure 1 is a graph showing leaching of DHCD from XEN-575 using various solvents at various temperatures.

Figure 2 is a graph showing leaching of DHCD from XEN-575 using acetonitrile at various temperatures. DETAILED DESCRIPTION OF THE INVENTION:

Biocatalytic transformations such as lipase resolutions, hydroxylations, reductions, and oxygenations often result in the formation of polar oxygenated organic compounds as products. A common problem with biocatalytic reactions is that they are conducted in aqueous media and, therefore, when polar products are formed as reaction products, separation of the products from the reaction media is problematic. When polar reaction products are not charged (i.e., nonionic) , separations are generally accomplished by liquid-liquid extraction. These extractions may be inefficient and have process limitations due to the formation of emulsions. When whole cells are contained in the aqueous medium to be extracted, cell lysis occurring during the biotransformation or from solvent contact may form an emulsion which cannot be processed.

Another problem with the recovery of many of these compounds is that they are unstable to extremes of pH and temperature which could facilitate extraction or distillation respectively.

The polar oxygenated organic compounds recovered by the processes described herein may be useful in synthesis of pesticides, herbicides, pharmaceuticals and other therapeutic agents. Preferred polar compounds are those having at least one hydroxyl group and at least one double bond, although it is understood that any polar oxygenated compound having no double bond or hydroxyl or more than one double-bond or hydroxyl are also useful in this invention.

More preferred are polar uncharged oxygenated organic compounds including, but are not limited to, halogenated oxygenated cyclohexadienes such as described in PCT applications WO 91/12257 and WO 91/16290, which are incorporated herein by reference. Because of the polarity of these compounds and the fact that they are uncharged, it is unexpected that they will be attracted to any resin, preferring to remain in solution.

Useful sorbents in the present invention are high surface area (>100m2/g) , porous polymer adsorbents or high surface area, porous pyrolyzed polymers. For example, sorbents, as described by Kolman, et al., in US Patent 4,064,063, and carbonaceous adsorbents, such as described by Agui, et al., in US Patent 4,883,596, and those commercially available, such as Ameberlite® XAD and AmbersorkP XE-347 or XE-348 (now called Ambersor s>575) , which are commercially available from Rohm and Haas. Preferred sorbents are the porous pyrolyzed polymer adsorbents commercially available from Rohm and Haas, particularly Ambersorb XE-347 and XE-348 (AmberεorkP575) .

Useful solvents or solvent mixtures for the present invention are those fully miscible in water. The solvent to be used for a particular adsorbate will depend on the chemical nature of the absorbed species. The polar solvents may include but are not limited to: methanol, ethanol, acetonitrile, acetone, tetrahydrofuran (THF), propanol, isopropanol, or water immiscible compounds, such as butanols and ethyl acetate when used in combination with an appropriate solvent(ε) , such that the mixture is miscible in water. For example, a mixture of ethyl acetate and ethanol.

Preferred solventε for recovery of a halogenated oxygenated cyclohexadiene are mixtures of ethyl acetate and ethanol or methanol.

The polar uncharged oxygenated compounds recovered by the procesεeε of the present invention may be synthesized by fermentation as described in D.T. Gibson and V. Subramanian, 1984, Microbial Degradation of Aromatic Hydrocarbons, pp. 181- 252; in Microbial Degradation of Organic Compounds, ed. D.T. Gibson, Marcel Dekker, Inc., or in non-growing cell suspensions as reported by A.J. Brazier, M.D. Lilly and A.B. Herbert, 1990, Enzyme Microb. Technol., 12:90-94. Typically, product concentration in fermentation broths for the production of such organic compoundε are low and these low concentrations, coupled with the fact that media components, buffers, proteins and metabolites can remain in solution after cell separation, usually dictate a multistage product recovery operation. For example, a commercial process for recovery of such compounds would typically include the following steps: 1) synthesiε compriεing fermentation, tissue culture and/or immobilized enzymes; 2) isolation compriεing homogenization, centrifugation, filtration, and/or extraction; 3) gross purification comprising adsorption, precipitation and/or extraction; 4) polishing comprising chromatography, adsorption and/or extraction; 5) solvent removal comprising evaporation and/or crystallization; and 6) drying. Such processes are cumbersome and cost inefficient on an industrial scale.

It has been found that using the processes of the present invention, certain organic compounds, as described herein, can be recovered by a much easier and much more efficient process, whereby one or more of the steps listed above (1-6) are eliminated. Specifically, the isolation and polishing steps, which are often very rate limiting and costly, have been eliminated from the present recovery process. EXPERIMENTAL:

Example 1 Binding

A whole fermentation broth containing 2,3-dihydroxy-l- chlorocyclohexa-4,6-diene (DHCD) was used in this experiment. The fermentation medium was a mineral salts medium containing potassium phosphate, ammonium sulfate, ferrouε sulfate, magnesium sulfate, trace minerals and fructose. The microorganism was a mutant strain of Pseudomonas putida which oxidizes chlorobenzene to DHCD. Gibson, D.T., et al., 1984 Microbial Degradation of Aromatic Hydrocarbons, pp. 181-252. This whole fermentation broth (including cells) contained approximately 22 g/1 DHCD. 100 ml of whole fermentation broth was contacted with 20 grams (dry weight) of once-used XEN-575 (commercially available from Rohm and Haas) in a vesεel with paddle εtirring. Samples were taken at the intervals indicated and assayed for DHCD, the results are summarized in Table I.

Table I

Example 2 Binding

A whole fermentation broth containing 2,3-dihydroxy-l- chlorocyclohexa-4,6-diene (DHCD) was used in this experiment. The fermentation medium was a mineral saltε medium containing potassium phosphate, ammonium sulfate, ferrous sulfate, magnesium sulfate, trace minerals and glucose. The microorganism was a recombinant strain of Escherichia coli expreεεing the necesεary enzymeε to oxidize chlorobenzene to DHCD (Zylεtera, G.J. ; Gibson, D.T. (1989) J. Bio. Chem. 264:14940-14946). This whole fermentation broth (including cells) contained approximately 103 g/1 DHCD. 100 ml of whole fermentation broth was contacted with 100 grams (dry weight) of once-used XEN-575 (commercially available from Rohm and Haas) in a vesεel with paddle stirring. Samples were taken at the intervals indicated and assayed for DHCD, the results are summarized in Table II.

Example 3

56.7g of a whole cell fermentation broth containing 2,3- dihydroxy-l-chlorocyclohexa-4,6-diene (DHCD) was contacted with 6.8 grams (dry weight) of once-used XEN-575 (commercially available from Rohm and Haas) in a vesεel with gentle agitation with an "orbit-type" εhaker for about 1 hour at 22"C. After adεorption, the broth was decanted from the resin whereby 53.9g (47.6ml) of broth was collected. A first batch water wash of 25.3ml water was done on the resin, no measurable amount of DHCD was found. A second batch water wash with 24 ml water was done on the resin and again no measurable amount of DHCD was found. The DHCD was leached from the resiπ using three batches of ethanol (95%) of about 24ml each. The three leachings were performed at 61°C. Results of the leaching are shown in Table III below. TABLE III

Leaching Cumulative Recovery ( % )

1st 42.4% 2nd 67.8% 3rd 80.0%

Example 4

The same experiment as set forth in Example 3 was carried out except that a total of five leachings were performed using five batches of ethanol (95%) of about 24ml each. The three initial leachings were carried out at 22"C. The fourth leaching was performed at 44^βC and a fifth leaching was performed at 61°C. Resultε of the leaching are provided in Table IV.

TABLE IV

Leaching Cumulative Recover % lεt

2nd

3rd

4th*

5th**

* Leaching performed at 44^βC ** Leaching performed at 61^βC Example 5

Similar experiments to Example 3 were carried out using various solvents or mixtures of solvents in the leaching phase as well as under various temperature conditions. The solvents, temperatures and number of leachings used in this example are set forth in tabular form below in Table V. Results showing percent recovery are provided in Figure 1.

TABLE V

Solvent em erature Number of Leachings

1-4 1-4 1-4 1-4 1-4 1-4 1-4 1-4 1-4 1-4

Following the process set forth in Example 3, 50ml of fermentation broth containing DHCD was contacted with 6.8g XEN-575 fresh (unused resin) for 1 hour with gentle stirring. After adsorption the broth was decanted off and the resin was washed with water and the leaching was performed with Ethyl Acetate at both 22 ^βC and 40"C. The percentage of recovery of DHCD from the broth is provided in Table VI.

TABLE VI

Leaching Cumulative Recovery f%)

1st 22'C 34.83% 2nd 22*C 52.61% 3rd 22'C 61.52% 4th 22°C 66.86% 1st 40^βC* 14.24% 2nd 40°C 24.18% 3rd 40°C 26.62% 4th 40^βC 27.99%

* Resin XEN-575 was reused for the second experiment run at 40°C, fresh XEN-575 was used for 22"C.

Example 7

Following the process set forth in Example 3 , adsorption and leaching was performed using XEN-575 and acetonitrile with four leachings at a temperature of 22°C, 40°C and 60°C. Percentage recovery of DHCD is provided in Figure 2.

Example 8

Approximately 100 grams of once used XEN-575 resin with approximately 7 grams of DHCD bound was washed with water and placed in a jacketed column operated at 65*C. The resin bed was about 22 x 2.6 cm. The DHCD was eluted from the resin with a solution of EtOH:EtOAc, 1:1; and collected in 25 ml aliquots. DHCD was quantified and the results are summarized in Table VII.

Aε expected, although batch elution iε quite good, elution of the resin when placed in a column is much more efficient than batch elution. Table VII showε that acceptable recovery is achieved with only a portion of the solvent used in a single batch leaching step.

In addition to the above examples we have also tried and obtained successful binding and leaching results from benzene dihydrodiol, toluene dihydrodiol, bromobenzene dihydrodiol and benzonitrile dihydrodiol.

The previous examples and disclosure are intended to serve as a repreεentation of embodiments herein and should not be construed as limiting the scope of this application.

Claims

WHAT IS CLAIMED IS:

1. A process for recovering polar uncharged oxygenated organic compounds from aqueous solutions, the procesε comprising:

a) contacting the polar oxygenated compound with a solid sorbent; and

b) leaching with one or more polar solvent or a mixture of polar εolventε in which the organic compound iε soluble.

2. A procesε of Claim 1 wherein the polar oxygenated organic compound comprises one or more hydroxyl groups and one or more double bond.

3. A process of Claim 2 wherein the organic compound is a halogenated oxygenated cyclohexadiene.

4. A process of Claim 1 wherein the polar solvent is selected from the group consiεting of: ethanol, methanol, ethyl acetate, acetonitrile or mixtures thereof.

5. A process of Claim 1 wherein the aqueous solution comprises fermentation broth, resuspended fermented cells or other biocatalysts.

6. A process of Claim 5 wherein the fermentation broth comprises cells or other impurities.

7. A process of Claim 1 wherein the polar organic compound is a product of a reaction catalyzed by a whole cell or cell free catalyst.

8. A process of Claim 7 wherein the catalyst is a mutant or recombinant organism or is derived from a recombinant Organism.

9. A process of Claim 8 wherein the recombinant organism is Escherichia coli.

10. A process of Claim 1 wherein the leaching is carried out at elevated temperatures, increaεing efficiency.

11. A process for recovering 2,3-dihydroxy-l-chlorocyclohexa- 4,6-diene (DHCD) the procesε comprising:

a) contacting a fermentation broth containing DHCD with a solid adsorbent; and

b) leaching the DHCD from the solid adsorbent using one or more polar solvent or mixtures of solvents.

12. A procesε of Claim 11 further compriεing batchwiεe adεorption.

13. A process of Claim 11 wherein the solid adsorbent is a carbonaceous adsorbent.

14. A process of Claim 11 wherein the polar solvent used in step (b) is selected from the group consiεting of: ethanol, methanol, ethyl acetate, acetonitrile or mixtures thereof.

15. A process of Claim 11 wherein the leaching is carried out at elevated temperatures, increasing efficiency.