| 1. |
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| 2. |
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| 3. |
- Lipnizki, Frank, et al.
(författare)
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Einsatz von Pervaporation-Bioreaktor-Hybridprozessen in der Biotechnologie
- 1998
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Ingår i: Chemie-Ingenieur-Technik. - : Wiley. - 0009-286X .- 1522-2640. ; 70:12, s. 1587-1595
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)abstract
- Pervaporation is a membrane separation process with considerable innovative possibilities in the area of biotechnology. Above all, the combination of bioreactor and pervaporation has potential in the longer term as an alternative to conventional batch processes. This article considers the state of the art of pervaporation/bioreactor hybrid processes. The possible applications of such hybrid processes are discussed and compared with conventional processes. It becomes apparent that the use of pervaporation/bioreactor hybrid processes can avoid product inhibition and greatly enhance the productivity of biotechnological processes.
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| 4. |
- Behravan, Gity, et al.
(författare)
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Formation of a free radical of the sulfenylimine type in the mouse ribonucleotide reductase reaction with 2'-azido-2'-deoxycytidine 5'-diphosphate
- 1995
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Ingår i: Biochimica et Biophysica Acta, Gene Structure and Expression. - : Elsevier BV. - 0167-4781 .- 1879-2634. ; 1264:3, s. 323-329
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Tidskriftsartikel (refereegranskat)abstract
- Mouse and Escherichia coli ribonucleotide reductases (RR) both belong to the same class of RR, where the enzyme consists of two non-identical subunits, proteins R1 and R2. A transient free radical was observed by EPR spectroscopy in the mouse RR reaction with the suicidal inhibitor 2′-azido-2′-deoxycytidine 5′-diphosphate. The detailed hyperfine structure of the EPR spectrum of the transient radical is somewhat different for the mouse and previously studied E. coli enzymes. When the positive allosteric effector ATP was replaced by the negative effector dATP, no transient radical was observed, showing that ‘normal' binding of the inhibitor to the substrate binding site is required. Using the mouse protein R2 mutants W 103Y and D266A, where the mutations have been shown to specifically block long range electron transfer between the active site of the R1 protein to the iron/radical site in protein R2, no evidence of transient radical was found. Taken together, the data suggest that the radical is located at the active site in protein R1, and is probably of the sulfenylimine type
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| 5. |
- Dunuwila, D.D., et al.
(författare)
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ATR FTIR spectroscopy for in situ measurement of supersaturation
- 1997
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Ingår i: Journal of Crystal Growth. - 0022-0248 .- 1873-5002. ; 179:1-2, s. 185-193
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Tidskriftsartikel (refereegranskat)abstract
- The current contribution establishes the technical feasibility of Attenuated Total Reflection (ATR) Fourier transform infrared (FTIR) spectroscopy for the in situ measurement of supersaturation in crystallization processes. The approach was inspired by recent advancements in ATR spectroscopy by way of various light transfer systems for remote sensing and by the increasing availability of ATR configurations well suited for remote, in situ measurements. The feasibility of the technique was investigated using a DIPPER-210® immersion probe manufactured by Axiom Analytical, Inc. Initial experiments conducted using aqueous maleic acid proved that ATR FTIR spectroscopy can be successfully employed to measure supersaturation, solubility and the metastable limit, in situ, with sufficient accuracy and precision.
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| 6. |
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| 7. |
- LeCaptain, D.J., et al.
(författare)
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Applicability of second harmonic generation for in situ measurement of induction time of selected crystallization systems
- 1999
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Ingår i: Journal of Crystal Growth. - 0022-0248 .- 1873-5002. ; 203:4, s. 564-569
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Tidskriftsartikel (refereegranskat)abstract
- The nonlinear optical technique of second harmonic generation (SHG) is introduced as a novel technique for monitoring particle formation in batch crystallizations. SHG is more sensitive and is less prone to interference than turbidometric methods. The studies presented show the applicability of SHG as a method for in situ measuring the induction time of a number of noncentrosymmetric crystal systems.
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| 8. |
- Miranda, E.A., et al.
(författare)
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Development of precipitant agents for precipitation of proteins based on hydrophobic interaction
- 1995
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Ingår i: Brazilian journal of chemical engineering. - 0104-6632 .- 1678-4383. ; 12:1, s. 1-12
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Tidskriftsartikel (refereegranskat)abstract
- Precipitation agents were developed to perform protein separation by precipitation based on hydrophobic interaction. They consisted of ligands, saturated linear chains of fatty acids, attached by esterification to a carrier molecule, methylcellulose. Precipitation of bovine serum albumin was achieved at 50 percent saturation of ammonium sulfate. The butyric acid derivative showed a higher efficiency in precipitating this protein than other derivatives tested. There is evidence that the interaction between the protein and the derivatives is hydrophobic.
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| 9. |
- Pan, Borlan, et al.
(författare)
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Time-resolved fluorescence and anisotropy of covalently coupled 1-pyrenebutyric acid for monitoring the crystallization conditions of lysozyme
- 1997
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Ingår i: Journal of Crystal Growth. - 0022-0248 .- 1873-5002. ; 171:1-2, s. 226-235
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Tidskriftsartikel (refereegranskat)abstract
- Time-resolved fluorescence and anisotropy measurements of trace amounts of 1-pyrenebutyric acid labeled hen egg-white lysozyme (PBA-HEL) were used to characterize hen egg-white lysozyme (HEL) crystallization conditions. The effects of sodium chloride and protein concentrations on the fluorescence lifetimes and rotational correlation times of the labeled protein were examined. These results were compared with the effects of the salts ammonium acetate and ammonium sulfate. Addition of protein precipitants caused increases in the rotational correlation times which were attributed to a combination of steric, hydrodynamic, general electrostatic and specific ionic interactions. This decrease in the rotational mobility of HEL appears to be a necessary but not sufficient condition to allow the formation of specific interactions leading to crystallization. The results demonstrated that fluorescence measurements are effective in characterizing and monitoring protein crystallization processes prior to the appearance of macroscopic crystals.
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| 10. |
- Rasimas, J. P., et al.
(författare)
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A molecular lock-and-key approach to detecting solution phase self-assembly : a fluorescence and absorption study of carminic acid in aqueous glucose solutions
- 1996
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Ingår i: Journal of Physical Chemistry. - : American Chemical Society (ACS). - 0022-3654 .- 1541-5740. ; 100:17, s. 7220-7229
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Tidskriftsartikel (refereegranskat)abstract
- We introduce a novel approach to the study of complex ternary systems where a fluorescent chromophore contains a functionality that incorporates into precrystalline aggregates in concentrated solutions. We demonstrate the feasibility of this approach by using carminic acid, a fluorescent molecule possessing a pendant glycosyl moiety, in aqueous glucose solutions. We report the steady state absorption and emission response of carminic acid as well as its picosecond dynamical response. These data, taken collectively, show that saturated glucose solutions exhibit anomalous molecular scale organization and that the persistence time of this organization is significantly less than a nanosecond. Our results indicate that kinetic contributions to crystallization are expected to play an important, sometimes dominant, role in this technologically important process.
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| 11. |
- Rasimas, J. P., et al.
(författare)
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Measuring self-assembly in solution : incorporation and dynamics of a "Tailor-made impurity" in precrystalline glucose aggregates
- 1996
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Ingår i: Journal of Physical Chemistry. - : American Chemical Society (ACS). - 0022-3654 .- 1541-5740. ; 100:42, s. 17034-17040
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Tidskriftsartikel (refereegranskat)abstract
- We have studied the onset of crystallization from solution using a fluorescent probe molecule that incorporates selectively into precrystalline glucose aggregates that form in supersaturated aqueous glucose solutions. We achieve incorporation of the fluorophore into the aggregates by virtue of the fluorophore pendant glycosyl moiety and compare the rotational diffusion data for this molecule to that for the nonglycosylated, native probe molecule. This experimental approach, in conjunction with semiempirical calculations to understand the electronic response of the fluorescent probe, provides insight into the formation and size of precrystalline glucose aggregates. Our data indicate that the aggregates effectively isolate the fluorophore from the solution over a range of glucose concentrations spanning the saturation point and that the lifetime of these aggregates is on the order of a nanosecond for aggregates that include the glycosylated probe molecule. The subtle but important differences between these results and those we reported previously for carminic acid in aqueous glucose solutions point to the significant role of labile protons in mediating the formation and dynamics of precrystalline glucose aggregates.
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| 12. |
- Rova, Ulrika, et al.
(författare)
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Evidence by mutagenesis that Tyr370 of the mouse ribonucleotide reductase R2 protein is the connecting link in the intersubunit radical transfer pathway
- 1999
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Ingår i: Journal of Biological Chemistry. - : Elsevier BV. - 0021-9258 .- 1083-351X. ; 274:34, s. 23746-23751
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Tidskriftsartikel (refereegranskat)abstract
- Ribonucleotide reductase catalyzes all de novo synthesis of deoxyribonucleotides. The mammalian enzyme consists of two non-identical subunits, the R1 and R2 proteins, each inactive alone. The R1 subunit contains the active site, whereas the R2 protein harbors a binuclear iron center and a tyrosyl free radical essential for catalysis. It has been proposed that the radical properties of the R2 subunit are transferred ~35 Å to the active site of the R1 protein, through a coupled electron/proton transfer along a conserved hydrogen-bonded chain, i.e. a radical transfer pathway (RTP). To gain a better insight into the properties and requirements of the proposed RTP, we have used site-directed mutagenesis to replace the conserved tyrosine 370 in the mouse R2 protein with tryptophan or phenylalanine. This residue is located close to the flexible C terminus, known to be essential for binding to the R1 protein. Our results strongly indicate that Tyr370 links the RTP between the R1 and R2 proteins. Interruption of the hydrogen-bonded chain in Y370F inactivates the enzyme complex. Alteration of the same chain in Y370W slows down the RTP, resulting in a 58 times lower specific activity compared with the native R2 protein and a loss of the free radical during catalysis.
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| 13. |
- Rova, Ulrika, et al.
(författare)
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Evidence by site-directed mutagenesis supports long-range electron transfer in mouse ribonucleotide reductase
- 1995
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Ingår i: Biochemistry. - : American Chemical Society (ACS). - 0006-2960 .- 1520-4995. ; 34:13, s. 4267-4275
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Tidskriftsartikel (refereegranskat)abstract
- Mammalian ribonucleotide reductase consists of two nonidentical subunits, proteins R1 and R2, each inactive alone. The R1 protein binds the ribonucleotide substrates while the R2 protein contains a binuclear iron center and a tyrosyl free radical, essential for activity. The crystal structures of the corresponding Escherichia coli proteins suggest that the distance from the active site in R1 to the tyrosyl radical buffed in R2 is about 35 Å. Therefore, an electron pathway was suggested between the active site and the tyrosyl radical. Such a pathway could include a conserved tryptophan on the suggested RI interaction surface of R2 and a conserved aspartic acid hydrogen bonded both to the tryptophan and to a histidine iron ligand. To find experimental support for such an electron pathway, we have replaced the conserved tryptophan in mouse R2 with phenylalanine or tyrosine and the aspartic acid with alanine. All the mutated R2 proteins were shown to bind metal with the same affinity as native R2 and to form the binuclear iron center. In addition, the W103Y and D266A proteins formed a normal tyrosyl free radical while only low amounts of radical were observed in the W103F protein. Neither the kinetic rate constants nor the equilibrium dissociation constant of the R1/R2 complex was affected by the mutations as shown by BIAcore biosensor technique. However, all mutant R2 proteins were completely inactive in the enzymatic assay, supporting the hypothesis that the tryptophan and aspartic acid residues are important links in an amino acid residue specific long-range electron transfer.
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| 14. |
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| 15. |
- Schmidt, Peter Paul, et al.
(författare)
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Kinetic evidence that a radical transfer pathway in protein R2 of mouse ribonucleotide reductase is involved in generation of the tyrosyl free radical
- 1998
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Ingår i: Journal of Biological Chemistry. - : Elsevier BV. - 0021-9258 .- 1083-351X. ; 273:34, s. 21463-21472
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Tidskriftsartikel (refereegranskat)abstract
- Class I ribonucleotide reductases consist of two subunits, R1 and R2. The active site is located in R1; active R2 contains a diferric center and a tyrosyl free radical (Tyr()), both essential for enzymatic activity. The proposed mechanism for the enzymatic reaction includes the transport of a reducing equivalent, i.e. electron or hydrogen radical, across a 35-Å distance between Tyr() in R2 and the active site in R1, which are connected by a hydrogen-bonded chain of conserved, catalytically essential amino acid residues. Asp266 and Trp103 in mouse R2 are part of this radical transfer pathway. The diferric/Tyr() site in R2 is reconstituted spontaneously by mixing iron-free apoR2 with Fe(II) and O2. The reconstitution reaction requires the delivery of an external reducing equivalent to form the diferric/Tyr() site. Reconstitution kinetics were investigated in mouse apo-wild type R2 and the three mutants D266A, W103Y, and W103F by rapid freeze-quench electron paramagnetic resonance with ≤4 Fe(II)/R2 at various reaction temperatures. The kinetics of Tyr() formation in D266A and W103Y is on average 20 times slower than in wild type R2. More strikingly, Tyr() formation is completely suppressed in W103F. No change in the reconstitution kinetics was found starting from Fe(II)-preloaded proteins, which shows that the mutations do not affect the rate of iron binding. Our results are consistent with a reaction mechanism using Asp266 and Trp103 for delivery of the external reducing equivalent. Further, the results with W103F suggest that an intact hydrogen-bonded chain is crucial for the reaction, indicating that the external reducing equivalent is a H(). Finally, the formation of Tyr() is not the slowest step of the reaction as it is in Escherichia coli R2, consistent with a stronger interaction between Tyr() and the iron center in mouse R2. A new electron paramagnetic resonance visible intermediate named mouse X, strikingly similar to species X found in E. coli R2, was detected only in small amounts under certain conditions. We propose that it may be an intermediate in a side reaction leading to a diferric center without forming the neighboring Tyr().
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| 16. |
- Schwartz, A.M., et al.
(författare)
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Use of Raman spectroscopy for in situ monitoring of lysozyme concentration during crystallization in a hanging drop
- 1999
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Ingår i: Journal of Crystal Growth. - 0022-0248 .- 1873-5002. ; 203:4, s. 599-603
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Tidskriftsartikel (refereegranskat)abstract
- Fiber optic Raman spectroscopy combined with a partial least-squares regression model was investigated as a means to monitor lysozyme concentration during crystallization in a hanging drop experiment in real time. Raman spectral features of the buffer and protein were employed to build the regression model. This model was used to calculate the compositional changes within the hanging drop. The use of fibre optic technology coupled with Raman spectroscopy, which is ideal for use with aqueous media, results in a powerful noninvasive probe of the changing environment within the solution. These preliminary findings indicate that solubility as well as supersaturation measurements can be made.
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| 17. |
- Uusi-Penttilä, Marketta S, et al.
(författare)
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Spectroscopic monitoring of environmentally benign anti-solvent crystallization
- 1996
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Ingår i: Journal of Crystal Growth. - : Elsevier BV. - 0022-0248 .- 1873-5002. ; 166:1-4, s. 967-970
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Tidskriftsartikel (refereegranskat)abstract
- There are very few studies dealing with the fundamentals of anti-solvent crystallization, even though this crystallization method is widely used in the pharmaceutical industry. Anti-solvent crystallization is accomplished by adding a miscible anti-solvent into a mixture of solute and solvent, effectively reducing the solubility of the solute in the solvent, and thus, causing the crystallization of the solute. Unfortunately, many of the anti-solvents used today are chlorinated hydrocarbons suspected of environmental damage. The current research demonstrates the use of environmentally benign solvents for anti-solvent crystallization and new approaches for monitoring of the solvent behavior during an anti-solvent crystallization. Results are presented confirming the efficacy of various water and ester systems for anti-solvent crystallization. Furthermore, the application of fluorescence spectroscopy for monitoring these crystallizations is demonstrated.
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| 18. |
- Uusi-Penttilä, Marketta S., et al.
(författare)
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Spectroscopically determined dielectric constants for various esters
- 1997
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Ingår i: Industrial & Engineering Chemistry Research. - : American Chemical Society (ACS). - 0888-5885 .- 1520-5045. ; 36:2, s. 510-512
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Tidskriftsartikel (refereegranskat)abstract
- Polarity of a solvent can be defined by the Onsager function or the Debye function, which both are functions of the static dielectric constant. Polarity has also been shown to be related to the solvatochromic shifts of the absorption and fluorescence spectra. In this study, the emission maxima of the probe molecule Nile Red were taken in different solvents of known dielectric constants, and relationships between the emission maxima and the Onsager and Debye functions were established. These relationships were used to estimate dielectric constants for various environmentally benign esters.
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| 19. |
- Yedur, Sanjay K., et al.
(författare)
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Synthesis and testing of catalysts for the production of maleic anhydride from a fermentation feedstock
- 1996
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Ingår i: Industrial & Engineering Chemistry Research. - : American Chemical Society (ACS). - 0888-5885 .- 1520-5045. ; 35:3, s. 663-671
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Tidskriftsartikel (refereegranskat)abstract
- It is necessary to develop alternate pathways for the production of chemicals that are traditionally produced from fossil fuels to reduce our dependency on nonrenewable energy sources. In this paper, an alternate technology is presented for producing maleic anhydride from a fermentation feedstock. The process involves the catalytic oxydehydrogenation of fermentation-derived succinic anhydride to produce maleic anhydride. Various catalysts have been synthesized and tested for the oxydehydrogenation reaction. Iron phosphate based catalysts are found to be the best on the basis of high conversions and selectivities obtained. The effects of temperature, oxygen concentration, contact time, and the total time on stream on the performance of the catalyst are investigated, and an optimum set of conditions for the operation of the bench-scale reactor is presented. The bulk and surface compositions, the surface areas, and the bulk crystallographic structure of the catalysts are also reported.
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| 20. |
- Yedur, Sanjay K, et al.
(författare)
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Use of fluorescence spectroscopy in concentration and supersaturation measurements in citric acid solutions
- 1996
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Ingår i: Applied Spectroscopy. - : SAGE Publications. - 0003-7028 .- 1943-3530. ; 50:7, s. 866-870
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Tidskriftsartikel (refereegranskat)abstract
- Measurement of supersaturation is of critical importance in the operation and control of crystallizers. In this work, we report a novel spectroscopic technique to achieve the measurement of concentration and supersaturation in crystallizing solutions. In order to develop a sensor for this measurement, citric acid is chosen as the model solute, and the analytical technique involves fluorescence spectroscopy. Citric acid is a common food-grade compound with a wide range of applications that is exclusively produced by crystallization. The fluorescent properties of a probe, 8-hydroxy-1,3,6-pyrenetrisulfonate (pyranine), are used to provide concentration measurements in aqueous citric acid solutions, thereby providing for supersaturation estimation. The change in the relative emission peak intensities of the probe in different solute concentrations gives an excellent calibration curve for concentration measurements. It is also shown that, although pyranine responds to both its solvent microenvironment and the pH of the solution, it is still possible to measure concentration and supersaturation by using this fluorescence technique.
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| 21. |
- Bachinger, Thomas
(författare)
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Multisensor arrays : for bioprocess monitoring
- 1999
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Bioprocess engineering, the technology that is focused on the exploitation of the metabolic potential of biological agents, has attracted growing interest throughout the past 50 years from both scientific and industrial communities. The products that have been brought to market range from pharmaceuticals and enzymes to food products and vitamins. The quality of human life has been improved through these efforts in many ways.Despite a strong research effort and the fact that microbial transformations often reach yields close to the theoretical maximum. many bioprocesses still operate at relatively low yields. One of the obstacles in effective operation is the extraction of useful information from the bioprocess. Sensors that acquire real-time information about the cells' state and their interaction with the environment in the bioreactor are seldom available. Hence, the implementation of sophisticated process control is prevented.In this thesis a new approach of non-invasive on-line bioprocess monitoring is evaluated. Chemical multisensor arrays (i.e. electronic noses) are used to extract information from the composition of volatiles emitted from the cell culture. The focus is on two specific areas: (i) monitoring of key variables in the bioreactor environment and (ii) monitoring of cell states and physiological events. The overall concern is, besides the increase of yield and reproducibility, the safety operation of bioprocesses.To cover a comprehensive area of modern bioprocessing, several organisms are investigated under different modes of operation in laboratory- and production scale processes. In repeated batch cultivations of recombinant Escherichia coli it is shown that an electronic nose can monitor biomass and specific growth rate with high accuracy. Glucose and ethanol concentration are monitored in batch cultivations of Saccharomyces cerevisiae. Bioproduct monitoring is presented in production-scale mammalian cell cultivation. The concentration of a therapeutic protein is monitored on-line in this long-term bioprocess thereby also outlining the stability of the sensor technique.In production-scale mammalian cell culture it is possible to follow cell transition states and monitor the reproducibility of the process. The physiological state of the cell population is revealed in laboratory-scale cultivations. It is shown that microbial contamination can be identified earlier than with conventional methods. Finally, the metabolic burden imposed on bacterial cells through strong overexpression of recombinant protein is monitored in fed-batch cultivation.
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| 22. |
- Bülow, Leif, et al.
(författare)
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The metabolic effects of native and transgenic hemoglobins on plants
- 1999
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Ingår i: Trends in Biotechnology. - 0167-7799. ; 17:1, s. 4-21
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Tidskriftsartikel (refereegranskat)abstract
- The strictly aerobic bacterium Vitreoscilla expresses a hemoglobin-like protein, VHb, when subjected to oxygen stress. When expressed in plants, this has several intriguing physiological effects, such as improving the overall growth rate, speeding germination and flowering, and increasing the productivity of certain oxygen-requiring metabolic pathways. Although the mechanisms behind the effects of VHb in heterologous hosts are not yet fully characterized, it has been suggested that VHb facilitates oxygen transport and/or storage. This hypothesis is supported by the kinetic properties of VHb, which allow very rapid dissociation of oxygen from the protein.
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| 23. |
- Glassner, David A., et al.
(författare)
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Purification process for succinic acid produced by fermentation
- 1995
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Ingår i: Applied Biochemistry and Biotechnology. - 0273-2289 .- 1559-0291. ; 51-52:1, s. 73-82
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Tidskriftsartikel (refereegranskat)abstract
- Succinic acid is a versatile four-carbon dicarboxylic acid. It can be used commerically as an intermediate chemical for the manufacture of 1,4-butanediol, maleic anhydride, and many other chemicals. Succinic acid can be produced by the fermentation of carbohydrates. A complete process for the production and purification of succinic acid from carbohydrates has been developed. The process includes fermentation, desalting electrodialysis, water-splitting electrodialysis, and crystallization to produce a pure crystalline succinic acid. This article will present experimental work performed in the development of this process.
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| 24. |
- Kwon, Yun Joong, et al.
(författare)
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Protein separation using metal ion-bound particles in aqueous two-phase system
- 1999
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Ingår i: Biotechnology Techniques. - 0951-208X. ; 13:2, s. 145-148
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Tidskriftsartikel (refereegranskat)abstract
- Metal ion affinity partitioning of protein in aqueous two-phase systems was studied using Sepharose as ligand carrier as an integrated adsorption partitioning. Cu(II)-bound Sepharose was mixed with protein solution and an aqueous two-phase system. The affinity sorbent was distributed quantitatively to the upper side or the interface. The binding studies of lysozyme to copper-bound gel in PEG/dextran two-phase systems demonstrate the feasibility of this bioseparation process. PEG/dextran system did not affect binding and elution of lysozyme to and from the Cu(II)-Sepharose particles.
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| 25. |
- Mamma, D., et al.
(författare)
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An alternative approach to the bioconversion of sweet sorghum carbohydrates to ethanol
- 1995
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Ingår i: Biomass and Bioenergy. - 0961-9534 .- 1873-2909. ; 8:2, s. 99-103
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Tidskriftsartikel (refereegranskat)abstract
- The ethanol fermentation of juice and press cake, resulting from the squeezing of sweet sorghum stalks at high pressure, was investigated. The juice was fermented by Saccharomyces cerevisiae and yielded 4.8 g ethanol per 100 g of fresh stalks. The press cake was fermented directly to ethanol by a mixed culture of Fusarium oxysporum and Saccharomyces cerevisiae and yielded 5.1 g ethanol per 100 g of fresh stalks. An overall ethanol concentration and yield of 5.6% (w/v) and 9.9 g of ethanol per 100 g of fresh stalks respectively was obtained. Based on soluble carbohydrates, the ethanol yield from press cake was doubled while the overall theoretical yield was enhanced by 20.7% due to the bioconversion of a significant portion of cell wall polysaccharides to ethanol. The process was found promising for further investigation.
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