Process Biochemistry 40 (2005) 969–974

Patent survey

Patent survey on process biochemistry

Marten Terpstra∗

Technisch-Wetenschappelijke, Publicaties Vakliteratuuronderzoek, Landrestraat 53, Den Haag AB 2551, The Netherlands

A chemo-enzymatic synthesis of optically enriched roseoxides

European patent application 1 373 548 corresponding to In-ternational Patent Application WO 03/087387 to the Coun-cil of Scientific and Industrial Research, New Delhi, India

Shubash C. Taneja and co-biochemists, all of Jammu, In-dia, developed a process for the synthesis of optically en-riched dextro- and levo-rotary isomers of rose oxide fromracemic citronellol.

Although there are numerous ways to synthesize roseoxides, most of the presently known routes involve acid-catalyzed cyclisation of (E)-3,7-dimethyl-5-octen-1,7-diolproduced in various ways from citronellol [G. Ohloff, Lien-hard, Helv. Chem. Acta 48 (1962) 182]. Ohloff prepared(E)-3,7-dimethyl-5-octen-1,7-diol by the photosensitizedair oxidation of citronellol to give alkyl hydroperoxidewhich on reduction and subsequent cyclisation with anacid gave a mixture ofcis- and trans-rose oxide. Eschinasiprepared [E.H. Eschinasi, J. Org. Chem. 35 (1970) 1097]a rose oxide mixture by the acid-catalysed cyclisation of(E)-8-acetoxy-2,6-dimethyl-1,3-octadiene obtained fromthe pyrolysis of 2,6-dimethyl-2,3,8-triacetoxy octane.

The invention provides a process for the preparation ofoptically enriched (−)-(2S, 4R)-rose oxide and its isomer(+)-(2R, 4S)-rose oxide from racemic citronellol, compris-ing cohalogenating racemic citronellol with a halogenationreagent in anhydrous alcohol to obtain an alkoxy halide, de-hydrogenating the alkoxy halide to obtain the corresponding3-octenol derivative, acylating the alcoholic function of the3-octanol derivative with an acylating agent in the presenceof a base to give the corresponding acylate, subjecting theacylate so obtained to kinetic resolution using a biocatalystor an enzyme, separating the mixture of reaction productscomprising of optically enriched hydrolyzed alcohol and un-

∗ Tel.: +31-70-3259101; fax:+31-70-3252824.E-mail address: sa017971@worldonline.nl (M. Terpstra).

hydrolyzed acylate derivatives, hydrolyzing the optically en-riched acylate with a base to furnish optically enriched pri-mary alcohol and cyclized the alcohol so obtained with anacid catalyst to produce dextrorotatory (2R, 4S)-rose oxide,the optically enriched hydrolyzed alcohol being directly cy-clized with an acid catalyst to produce levo-rotatory (2R,4S)-rose oxide.

The alkyl group in the alkoxy halide may be selected fromthe group consisting of methyl, ethyl,n-propyl andn-butyl,and the halide may be selected from the group consisting ofchloro, bromo, iodo.

Furthermore, the 3-octanol formed, preferably, is 2-alkoxy-3-halo-2,6-dimethyl-8-octanol.

Dehydrohalogenation of the alkoxy halide is done us-ing a strong base or an alkali to provide (E)-2-alkoxy-2,6-dimethyl-3-octen-8-ol.

The enzyme the inventors used for their process is selectedfrom a hydrolase and lipase fromPseudomonas sp. lipase(PSL) and Candide cylinderacae lipase (CCL).

15 pages; 31 claims; International search report included.

ReferencesIzumi T, et al. J Chem Technol Biotechnol 1997;68:57–64.Frater G, et al. Tetrahedron 1998;54:7633–703.Chapuis C, et al. Appl Catal 2001;221:93–117.UK patent publications GB 1010057 and GB 1054420.US patent publication 5892059.

A process and device of treating wastewater containingnitrogen-containing dyes

European patent application 1 375 437 to Kuraray Co. Ltd.,Okayama-Pref., Japan

Fig. 1 is a flow chart illustrating a nitrogen dye con-taining wastewater purifying system developed by TadaoShiotani and co-bioengineers, all of Osaka, Japan. Theirsystem comprises an obligatory anaerobic tank for bringingthe wastewater into contact with sulfate-reducting bacteria

0032-9592/$ – see front matterdoi:10.1016/j.procbio.2004.03.001

970 M. Terpstra / Process Biochemistry 40 (2005) 969–974

Fig. 1. A flow chart illustrating the waste apparatus of Tadao Shiotani et al.

under obligatory anaerobic conditions, a nitrification tankfor bringing the wastewater into contact with nitrifying bac-teria under aerobic conditions and a denitrification tank forbringing the wastewater into contact with the denitrifyingbacteria under anaerobic conditions.

A wastewater treatment apparatus was manufactured insuch a manner that an anaerobic tank (1), a denitrificationtank (2), and a nitrification tank (3) were disposed in the or-der mentioned and 75% of nitrified wastewater dischargedfrom the nitrification tank was returned circulated to thedenitrification tank. Each tank had a capacity of 150 L.The anaerobic tank (1) was filled with, as a microorganismimmobilized support, 10% of acetalized polyvinyl alcoholhydrogel (“PVA gel”) having sulfate reducing bacteria im-mobilized thereon. The PVA gel was made to flow by anagitator. Aeration was conducted via an air diffusing tube.Actual wastewater containing Acid Orange 7 and Acid Blue126 was employed.

The actual wastewater had a chromaticity of 360, BODconcentration of 1545 mg/L and T–N concentration of408 mg/L. The chromaticity of the wastewater at the out-let of the nitrification tank was 18, clearly suggestingthe decolorization of the wastewater. The BOD and T–Nconcentrations of the wastewater were 18 and 20 mg/L,respectively.

In a manner similar to that employed above except thatthe obligatory anaerobic tank was omitted, wastewater treat-ment was conducted. The BOD concentration of the treatedwastewater at the outlet of the nitrification tank was 19 mg/L,almost a level similar to that described above, but the T–Nconcentration was 68 mg/L and chromaticity was 203, indi-cating that the wastewater treated in a comparative examplewas not decolorized.

11 pages; 10 claims; 2 figures; European search reportincluded.

ReferencesKrull R, Hempel DC. Chemie Ingenieur Technik 2000;

72(9):1113–4.Stolz A. Appl Microbiol Biotechnol 2001–7;56(1–2):

69–80.O’Neill C et al. Appl Microbiol Biotechnol 2000;53(2):

249–54.

US patent publications 4384956; 6054044.European patent publications 426933; 861808.

A process for the synthesis of synthons for the manufac-ture of bioactive compounds

US preliminar patent application 2003/0232416 toChi-Huey Wong et al., San Diego, CA, USA

Chi-Huey Wong and co-biochemists discovered that2-deoxyribose-5-phosphate aldolase (DERA, EC 4.1.2.4)and variants thereon, can be used to catalyze sequentialasymmetric aldol reactions between a wide variety of donorand acceptor aldehydes. The reaction products typicallycontain at least two new stereogenic centers and can be pro-duced in enantiomerically pure form. As such, the DERAcatalyzed asymmetric adol chemistry can be exploited toproduce synthons for the synthesis of a variety of bioactivemolecules.

More specifically, their process involved the productionof bioactive compounds, one of which being anenantiomeri-cally pure pyranose. The process comprises contacting a firstachiral aldehyde, a second achiral aldehyde and the thirdone with 2-deoxyribose-5-phosphate aldolase (DERA) or aderivative thereof under conditions which are suitable to fa-cilitate sequential asymmetric aldol reactions.

The first aldol reaction between the first and the sec-ond achiral aldehydes forms the first reaction product andwherein the second aldol reaction between the first reactionproduct and the third achiral aldehyde forms a second re-action product. The second reaction product spontaneouslyundergoes an intermolecular cyclization reaction to form theenantiomerically pure pyranose.

Fig. 2 illustrates the mechanism of DERA catalyzedaldol reaction between the natural donor acetaldehydeand acceptord-glyceraldehyde-3-phosphate to generated-2-deoxyribose-5-phosphate.

22 pages; 28 claims; 7 figures.

Not yet available.Note: More information can be obtained from Gray

Cary Ware & Freidenrich LLP, Suite 1100, San Diego, CA92121-2133 (USA).

M. Terpstra / Process Biochemistry 40 (2005) 969–974 971

Fig. 2. DERA catalyzed aldol reaction of Chi-Huey Wong.

A novel antagonistic yeast useful in controlling spoilageof agriculture produce, and methods of use thereof

European patent application 1 372 384 to The State of Israel,Ministry of Agriculture, Beit-Dagan, Israel

Samir Droby, a biochemist and assignor to the aboveIsraelian Ministry, discovered that a yeast of the speciesMetschnikowia fructicola, identified as NRRL Y-27328 (Na-tional Center for Agricultural Utilization Research Peoria,Illinois, USA) is capable of inhibiting growth of a deleteri-ous micro-organism on a portion of a plant to which a bio-logically effective amount of a culture of the yeast is applied.

The inventor describes a composition for use in the pro-tection of citrus fruits and tomatoes, containingM. fructicolaand a carrier, and he carried out several laboratory tests, ofwhich the following will be discussed.

M. fructicola is propagated under aerobic conditions attemperatures ranging from 5 to 37◦C. Optimal growth tem-perature is 20–27◦C. The growth is in liquid medium (nu-trient broth; Droby et al., 1989) with a neutral pH. Thecell density of the yeast reaches its maximum (stationarystage) growth in 24–48 h. For laboratory and small scaletests growth in Erlenmeyer flasks containing the mediumand shaken on a rotary shaker is suitable. For large scale andcommercial tests, fermentation tanks and industrial growthmedia are preferred. The yeast cells are harvested by cen-trifugation using conventional laboratory or industrial cen-trifuges.

Individual grapes or cherry tomatoes were removed fromclusters. Surface disinfection was accomplished by dippingfor 1 min in 1% (v/v) sodium hypochlorite (pH 11.5). Dis-infected fruit was mounted on masking tape strips glued toPVC pads with an incubation box. The fruit was puncturedwith a pin to a depth of 2 mm and 10�l of an antagonist (M.fructicola or as indicated) cell suspension and were pippetedonto the wound site and left to dry for 1–2 h. Fruit was theninoculated with 10�l of conidial suspension of an appropri-ate fungal pathogen (B. cinerea or as indicated). Conidialsuspensions were obtained from 1-week-old pathogen cul-tures incubated at room temperatures. Spore concentrationwas adjusted to 1−5×104 conidia/mL. Each treatment wasapplied to three replicates of 7–10 individual fruit. Followingthe treatment, wet filter paper was placed in the incubationboxes which were covered with polyethylene to maintainhigh relative humidity. The percentage of decayed berries/fruits in each replicate was evaluated after 4–5 days at 20◦C.

Fig. 3 is a graph of cell number as a function of timefor the yeastsM. fructicola (Mf), Metschnikowia reukafii(231) andkluyveromyces thermotolerance (414) incubatedat 40◦C indicating that Mf grows faster than other yeasts inthe cold.

46 pages; 21 claims; 27 figures.

ReferencesDe Curtis et al. Ann Microbiol Enzymol 1996;46:

45–55.

972 M. Terpstra / Process Biochemistry 40 (2005) 969–974

Fig. 3. Cell number as a function of time for the yeastsM. fructicola,M. reukafii and K. thermotolerance according to Samir Droby.

Piano et al. Postharvest Biol Technol 1997;11:131–40.7 US patent publications.International patent publications WO 92/18009; WO

91/01641.

An artificial intelligence (AI) control system and methodfor treating sewage/wastewater by means of a neutralnetwork and a back-propagation algorithm

Fig. 4. Block diagram illustrating the constitution of the Al control system of Ik-Bae Yang.

European patent application 1 376 276 to H2L Co. Ltd.,Aanyang-City, South Korea

Fig. 4 shows a block diagram of an AI control systemthat has been developed by the South Korean bioengineerBae Yang, whileFig. 5 illustrates the computer program theinventor used for his process.

In Fig. 4the overall constitution of the AI control system isillustrated and in such constitution, data obtained from eachmeasuring instrument (10–12), such as BOD, an incomingstream flow and a water temperature as attributes of theinflow water, and each DO of the first and second story of theexhalation tank, MLSS of the aeration tank, a concentrationof sludge, a drawn stream flow and BOD of the outflowwater for controlling each SP, are transmitted to A/D card(101) of PLC (14) in analog signals.

Such data constitute the system steps according to the flowchart as shown inFig. 5. First, the program starts and it isinitialized (201), then the existing data are collected (202);the data are filtered (203); attributes of the inflow water,the internal condition of the reaction tank and the outflowwater as measured are inputted to PLC (205); and the cur-rent condition is identified (204). Then, each identified PVand each SP input value (206) are operation-processed (207,210, 211) for control output and then control-outputted (209,212) in analog and digital signals, respectively, and at thesame time saved (218) in the memory of PLC (14). Then, incase the data are judged to be in conformity with the crite-ria for efficiency (214), the data are operation-processed to

M. Terpstra / Process Biochemistry 40 (2005) 969–974 973

Fig. 5. The program schedule of Ik-Bae Yang.

convert them into the data of a physical quantity through themoving average process (217), and then the data are savedby dates (221) and each initial value is saved (219). Fur-ther, the data are bi-directionally communicated by meansof TCP/IP (transmission control protocol/internet protocol)through RS232C communication means (106) as shown inFig. 4 so that they are displayed on the monitor screen ofthe computer set (220).

The inventor discusses several steps in his control systemand calculates the entire error by a rate of change againstthe weight of sludge and/or wastewater.

28 pages; 20 claims; 16 figures; European search reportincluded.

ReferencesCote M et al. Water Research, Elsevier Science Publishers,

Amsterdam, NL. vol. 29, Nr. 4, p. 995–1004.Bhat N et al. In: Proceedings of the American Control

Conference, Pittsburgh, June 21–23, 1989, IEEE, US, vol.2 Conf. 8, p. 1342–7.

Pirsing A. Engineering and Automation, Siemens AG,Berlin, DE, vol. 19, Nr. 3, p. 25–7.

Minderman PA et al. In: Proceedings of the AmericanControl Conference, San Francisco, June 2–4, 1993, NewYork, IEEE, US, vol. 2, p. 1480–4.

Japanese patent publication 06335694.German patent publication 19914277.

An integrated technology for treating and valorizatingorganic waste

Canadian Patent Application 2 435 319 to Bio-Terre Sys-tems, Inc., Canada

An integrated process for treating and valorization of or-ganic waste have been proposed by Gerard Laganière andco-bioengineers. Their proposal broadly comprises

(a) treating the organic waste with a microorganism treat-ment for producing gas;

(b) separating waste resulting from step (a) into a liquidfraction and a solid fraction; and

(c) using the liquid fraction resulting from step (b) as afertilizer.

Populations living around porcine production are com-plaining about odors that are greatly affecting their qualityof life when it is not their health that is affected.

Pigs producers and other animal producers are using dif-ferent types of barns and buildings to grow their animals.Some producers cover all stages of production and othersare finishers only (from 20 to 110 kg). These animals pro-duce manure that is managed as solid or liquid dependingon the producer’s operations and installations.

Liquid management of the manure is the most popular. Itneeds less manpower and is most cost-effective. However,the qualities of the manure to manage are enormous (from0.35 to 1.0 m3 per pig produced) and generate odors, con-tain pathogens, phosphorous, and other substances contam-inating the environment and being of potential danger forpublic health.

Nevertheless, the inventors found that a proper valoriza-tion of pig manure in an integrated process, according towhich biogas is produced by the action of anaerobic bac-teria, will result in a substantial reduction of the aboveproblems.

To explain their idea in more detail reference is made toFig. 6 that illustrates the process used for the valorizationof animal manure. Animal manure is collected from a barnand is transferred periodically to bioreactors to match theoperation of the farm. The manure feeding system can eitherfunction by gravity or by pump and the manure can be fed toone bioreactor or to a plurality of bioreactors in concordancewith the type and volume of manure and the size of thebioreactors. Raw manure is fed to the bioreactors and itscomposition depends on the mix volume of many types ofmanure (sow, piglet or hogs) to obtain a COD (chemicaloxygen demand) content that is chosen for the design of thebioreactors. The COD value depends on the temperature andthe volume of the bioreactors. Preferred COD values in thepresent invention are 7–120 kg/m3. The animal manure inthe bioreactors is treated by microorganisms. In a preferredsystem, the animal manure is treated in accordance with thetechnology of anaerobic digestion under low temperature.The residence time of the manure in the bioreactors is a

974 M. Terpstra / Process Biochemistry 40 (2005) 969–974

Fig. 6. Block diagram of the biogas production according to Gerard Laganiere et al.

period of time sufficient for a full production cycle of biogasby the microorganism. In the preferred system, this periodof time is sufficient for the microorganism to digest 80% ofthe organic matter and the biogas produced has an averageof 70% of CH4. This period of time also allows reduction ofCOD, elimination of manure pathogens and elimination ofthe odors related to manure composition. In a more preferredsystem of the invention, the residence time is between 7 and21 days.

As illustrated inFig. 6, the biogas produced during themanure treatment is captured by any gas collection system(14) that is suitable for collecting gas. The biogas collectedis sent to a flare (16) or is sent to a purification unit (18). Theflare (16) is used to burn the excess volume of gas that cannotbe processed and during emergency or plant shut down formaintenance. For security reasons, a flame arrester (20) ispreferably located in the gas entrance to prevent the flamesfrom the flare (16) from going back to the bioreactors (12).

The biogas produced is sent to the purification unit (18)to eliminate the hydrogen sulfide and some of the watervapor present in the biogas. The hydrogen (H2S) concen-tration varies in time and is related to methane produc-

tion. The methane and H2S production are related since themethanogens and the sulfate reducer bacteria both use aceticand propionic acid as a substrate to produce methane andH2S. Tests done on the semi-industrial scale bioreactors hasshown that the H2S concentration varies between 1000 and6000 ppm during a cycle of anaerobic digestion and H2Sconcentration follows the same profile as biogas productioncurve.

The purification unit (18) consists in a combination ofmore than one filtration units. It is designed to cut off thepeaks of H2S concentration and maintain an acceptable H2Slevel at the exit of the process. The filters preferably used arecommercially available filters such as SulfatreatTM 410HP(The Sulfatreat Company, Chesterfield, MS, USA), whichuses a fixed bed with iron oxide type media.

The lab results has shown that under stable H2S concen-tration of 3000 ppm, the filtration efficiency remain at 83%after 7 days of operation and the efficiency of this treatmentis enhanced when the biogas is saturated with water.

16 pages; 5 figures; 20 claims.US patent publication 5863434.