Effects of temperature on microbial ethanol production. II. A thermodynamic interpretation of the temperature profile curve of ethanol production

An attempt is made to give a thermodynamic interpretation of the complete temperature profile curve of ethanol formation. Taking into consideration an enhancing competition between thermal activation and thermal deactivation of ethanol formation at increasing temperatures and supposing that the ethanol production is affected by a reversible and an irreversible term of thermal deactivation of a modified ARRHENIUS equation being current for the total biokinetic sphere may be derived: . The quantities ΔH and ΔH^D_2T are identical with the temperature functions of the change of entropy caused by reversible and irreversible deactivation of ethanol formation, respectively. Accordingly for the yeast strain Saccharomyces cerevisiae Sc 5 the calculated entropy coefficients of reversible and irreversible thermal deactivation of ethanol formation amount to C = (0.245± 0.013) kJ/mol · deg.² and C = (1.657 ± 0.046) kJ/mol · deg.².     

Polyiodine units in starch–iodine complex: INDO CI study of spectra and comparison with experiments

The starch–iodine blue complex formation does not involve negatively charged iodine species like I, I, or I; rather, neutral iodine units are involved. The heat of reaction is determined to be about ?110 kJ for every mole of I-I unit in the amylose helix, which suggests that the dissociation of I₂ (binding energy 149 kJ/mol) does not take place during the complex formation. Quantum mechanical (INDO CI) calculations indicate that the linear as well as nonlinear polyiodine units, I₆, with interiodine distance of 3.0 Å are responsible for characteristic absorbance bands of the starch–iodine complex. Based on our previous article [(1989) J. Polym. Sci. A 27 , 4161] and the present studies we identify (C₆H₁₀O₅)_16.5I₆ to be the polymeric unit responsible for the characteristic blue color of the complex.     

How Large is the Probability for the Estimate of a Variance Component to be Negative?

For a balanced one-way classification, where the normally distributed observations obey a random model y_ij=μ+b_i+c_ij with two variance components var (b_i) = δ and var (c_ij) = δ, the probability is given that the analysis of variance estimate of δ will be negative. This probability depends on δ/δ and the degrees of freedom in the ANOVA table. Tables for this probability are given. If the normally distributed observations obey an intra-class correlation model, the probability that the Mean Square between groups is smaller than the Mean Square within groups can also be evaluated from the given tables.     

Gas balance method for testing of microbial growth efficiency after carbon substrate pulse

The present paper deals with the analysis of the amount of oxygen utilized for oxidation of a small dose of carbon substrate in carbon limited Brevibacterium flavum culture. The ratio of the measured oxygen consumption (mo₂) to the amount of added carbon substrate (m_s) gives a stoichiometric coefficient of the biological oxidation equation. A linear relationship between mo₂ and m_s was observed. To compare the efficiency of different carbon substrate utilization there has been introduced a normalized value β = m/m. There exists a simple relationship between β and the thermodynamical growth efficiency η The theoretical considerations are proved by experimental results with β, η and Y_x/s in a chemostat culture at various medium flow rates.     

Whole‐cell‐based CYP153A6‐catalyzed (S)‐limonene hydroxylation efficiency depends on host background and profits from monoterpene uptake via AlkL

Living microbial cells are considered to be the catalyst of choice for selective terpene functionalization. However, such processes often suffer from side product formation and poor substrate mass transfer into cells. For the hydroxylation of (S)‐limonene to (S)‐perillyl alcohol by Pseudomonas putida KT2440 (pGEc47ΔB)(pCom8‐PFR1500), containing the cytochrome P450 monooxygenase CYP153A6, the side products perillyl aldehyde and perillic acid constituted up to 26% of the total amount of oxidized terpenes. In this study, it is shown that the reaction rate is substrate‐limited in the two‐liquid phase system used and that host intrinsic dehydrogenases and not CYP153A6 are responsible for the formation of the undesired side products. In contrast to P. putida KT2440, E. coli W3110 was found to catalyze perillyl aldehyde reduction to the alcohol and no oxidation to the acid. Furthermore, E. coli W3110 harboring CYP153A6 showed high limonene hydroxylation activities (7.1 U g). The outer membrane protein AlkL was found to enhance hydroxylation activities of E. coli twofold in aqueous single‐phase and fivefold in two‐liquid phase biotransformations. In the latter system, E. coli harboring CYP153A6 and AlkL produced up to 39.2 mmol (S)‐perillyl alcohol L within 26 h, whereas no perillic acid and minor amounts of perillyl aldehyde (8% of the total products) were formed. In conclusion, undesired perillyl alcohol oxidation was reduced by choosing E. coli's enzymatic background as a reaction environment and co‐expression of the alkL gene in E. coli represents a promising strategy to enhance terpene bioconversion rates. Biotechnol. Bioeng. 2013; 110: 1282–1292. © 2012 Wiley Periodicals, Inc.     

Conformation study of poly(γ-methyl-L-glutamate) in helicogenic solvents by small-angle X-ray scattering

Small-angle x-ray scattering of poly(γ-methyl-L -glutamate), [Glu(OMe)]_n, in m-cresol and in pyridine was measured to determine the mass per unit length, M_q, and the radius of gyration of the cross section, 〈S〉^1/2. It was confirmed from the values of M_q that [Glu(OMe)]_n exists in an α-helical conformation in these solvents. It was elucidated from the calculations on 〈S〉^1/2 that the side chains come in moderately close contact with the main chain in these solvents. It was indicated from the analysis of the outer portion of the scattering curves that the side-chain conformation varied depending on the solvent.     

Zur Kinetik der Mischsubstratutilisation bei der Citronensäure-akkumulation durch Saccharomycopsis lipolytica EH 59

The mechanism and the kinetic of the assimilation of mixed substrates during the organism growth and the product excretion by strain of Saccharomycopsis lipolytica have been studied. The assimilation of citric acid for the organism growth was prevented when glucose and/or n-paraffins are present as substrates. Citric acid concentrations higher than 30 g/l in the fermentation medium decrease the growth rate on the substrate glucose. Kinetic studies of the mixed substrate assimilation by means of a n-tetradecane-1-C-labelled paraffin fraction proved that in discontinuous as well as in continuous 1-stage-processes for microbial production of citric acid glucose as substrate is only used for the organism growth whereas the n-paraffin fraction is only used for the acid excretion.     

Determination of enantiomerization barriers of hypericin and pseudohypericin by dynamic high‐performance liquid chromatography on immobilized polysaccharide‐type chiral stationary phases and off‐column racemization experiments

Direct enantiomer separation of hypericin, pseudohypericin, and protohypericin was accomplished by high‐performance liquid chromatography (HPLC) using immobilized polysaccharide‐type chiral stationary phases (CSPs). Enantioselectivities up to 1.30 were obtained in the polar‐organic elution mode whereby for hypericin and pseudohypericin Chiralpak IC [chiral selector being cellulose tris(3,5‐dichlorophenylcarbamate)] and for protohypericin Chiralpak IA (chiral selector being the 3,5‐dimethylphenylcarbamate of amylose) gave favorable results. Enantiomers were distinguished by on‐line electronic circular dichroism detection. Optimized enantioselective chromatographic conditions were the basis for determining stereodynamic parameters of the enantiomer interconversion process of hypericin and pseudohypericin. Rate constants delivered by computational simulation of dynamic HPLC elution profiles (stochastic model, consideration of peak tailing) were used to calculate averaged enantiomerization barriers (ΔG) of 97.6–99.6 kJ/mol for both compounds (investigated temperature range 25–45°C). Complementary variable temperature off‐column (i.e., in solution) racemization experiments delivered ΔG = 97.1–98.0 kJ/mol (27–45°C) for hypericin and ΔG = 98.9–101.4 kJ/mol (25–55°C) for pseudohypericin. An activation enthalpy of ΔH^# = 86.0 kJ/mol and an activation entropy of ΔS^# = ?37.7 J/(K mol) were calculated from hypericin racemization kinetics in solution, whereas for pseudohypericin these figures amounted to 74.1 kJ/mol and ?82.6 J/(K mol), respectively. Although the natural phenanthroperylene quinone pigments hypericin and pseudohypericin as well as their biological precursor protohypericin are chiral and can be separated by enantioselective HPLC low enantiomerization barriers seem to prevent the occurrence of an excess of one enantiomer under typical physiological conditions—at least as long as stereoselective intermolecular interactions with other chiral entities are absent. Chirality 2010. © 2009 Wiley‐Liss, Inc.     

On the blue color of triiodide ions in starch and starch fractions. I. Characterization and assignment of absorption and circular dichroism spectra of triiodide ions in amylose

Four fundamental Raman lines were observed at 159, 111, 55 and 27 cm^-1 corresponding to the I bound (I) in amyloses with DP from 20 to 100, regardless of the degree of polymerization of I and the excitation wavelength. The spectral resolution was based on the molar extinction coefficient and molar ellipticity spectra of I. Eight bands, named, S₁, S₂, ?, S₈ from long to short wavelength, were isolated. These were found regardless of the DP. By a resonance excitation Raman study, the characteristics of S₃ and S₄, comprising the shoulder around 480 nm, were found to be different from those of S₁ and S₂, comprising the blue band. The assignment of the spectra was based on the electronic states of the monomeric I in the exciton-coupled dimeric unit. It was concluded that the blue band (S₁,S₂) belonged to the long-axis transitions and the shoulder band (S₃,S₄) to the short-axis ones on the monmeric coordinate system.     

Calculation of thermodynamic properties of protein in Escherichia coli K-12 grown on succinic acid, energy changes accompanying protein anabolism, and energetic role of ATP in protein synthesis

Using an average of the results from three methods of calculation, estimations are made of the thermodynamic properties of a unit carbon formula weight (UCFW) of Escherichia coli K-12 protein. These resulted in values fro DeltaG(f) of -38.09 kJ (-9.10 kcal)/ UCFW, for DeltaH(f) of -68.18 kJ (-16.29 kcal)/UCFW, and for DeltaS(f) of -94.2 J (-22.5 cal)/UCFW deg. The absolute entropy of one UCFW of E. coli K-12 protein is calculated to be 73.8 J/UCFW deg. Using these values, the corresponding changes in thermodynamic properties accompanying the anabolism of protein by this microorganism to from one UCFW of protein by this microorganism to from one UCFW of protein are calculated to be 1.97 kJ (0.47 kcal)/UCFW for DeltaG, 0.75 kJ (0.18 kcal)/UCFW for DeltaH, and -4.09 J (-0.98 cal)/UCFW deg for DeltaS. All these values are sufficiently close to zero that they may be considered to be so. The question is raised as to the quantity of ATP energy conserved within the substance of the protein as it is synthesized from succinic acid. It is calculated that only 3.8% of the total free energy available from ATP that is required during protein anabolism can have been conserved within the substance of the protein, there being a net conversion of the remaninder into heat and entropy.     

Application of the nearest-neighbor Ising model to the helix-coil transition of polypeptides in mixed organic solvents.

Using the formalism of nearest-neighbor Ising model and assuming that the allowed states for a monomeric unity of a polypeptide chain in solutions containing strong acids are E (helix), C (coil), and CS (solvent-bonded coil), the partition function of the system was deduced analytically. Equations were obtained which permitted the prediction of the characteristic thermodynamic behavior of the helix–coil transition under these conditions. These equations were used to examine critically the possible correlations between experimental data obtained using different techniques. Particular attention was devoted to quantities called “transition enthalpies,” obtained from the slope of the transition curves at the point where the helix fraction is one-half (ΔH), or for measurements of the heat of solution of the polymer over the total range of solvent composition (ΔH), or from heat capacity measurements taken at various temperatures (ΔH). Literature data of ΔH(j = opt, sol, cal) for the system poly-γ-benzyl-L -glutamate in mixtures of dichloroacetic acid and 1,2-dichloroethane were carefully analyzed.     

Interplay of reactive oxygen species,intracellular Ca2+ and mitochondrial homeostasis in the apoptosis of prostate cancer cells by deoxypodophyllotoxin

The limited treatment option for recurrent prostate cancer and the eventual resistance to conventional chemotherapy drugs has fueled continued interest in finding new anti‐neoplastic agents of natural product origin. We previously reported anti‐proliferative activity of deoxypodophyllotoxin (DPT) on human prostate cancer cells. Using the PC‐3 cell model of human prostate cancer, the present study reveals that DPT induced apoptosis via a caspase‐3‐dependent pathway that is activated due to dysregulated mitochondrial function. DPT‐treated cells showed accumulation of the reactive oxygen species (ROS), intracellular Ca surge, increased mitochondrial membrane potential (MMP, ΔΨ_m), Bax protein translocation to mitochondria and cytochrome c release to the cytoplasm. This resulted in caspase‐3 activation, which in turn induced apoptosis. The antioxidant N‐acetylcysteine (NAC) reduced ROS accumulation, MMP and Ca surge, on the other hand the Ca²⁺ chelator BAPTA inhibited the Ca overload and MMP without affecting the increase of ROS, indicating that the generation of ROS occurred prior to Ca²⁺ flux. This suggested that both ROS and Ca signaling play roles in the increased MMP via Ca‐dependent and/or ‐independent mechanisms, since ΔΨ_m elevation was reversed by NAC and BAPTA. This study provides the first evidence for the involvement of both ROS‐ and Ca‐activated signals in the disruption of mitochondrial homeostasis and the precedence of ROS production over the failure of Ca²⁺ flux homeostasis. J. Cell. Biochem. 114: 1124–1134, 2013. © 2012 Wiley Periodicals, Inc.     

Thermodynamic analysis of growth of Methanobacterium thermoautotrophicum

Growth of Methanobacterium thermoautotrophicum, an anaerobic archaebacterium using methanogenesis as the catabolic pathway, is characterized by large heat production rates, up to 13 W g⁻¹, and low biomass yields, in the order of 0.02 C‐mol mol⁻¹ H₂ consumed. These values, indicating a possibly “inefficient” growth mechanism, warrant a thermodynamic analysis to obtain a better understanding of the growth process. The growth‐associated heat production (Δ_rH) and the growth‐associated Gibbs energy dissipation per mol biomass formed (Δ_rG) were −3730 kJ C‐mol⁻¹ and −802 kJ C‐mol⁻¹, respectively. The Gibbs energy change found in this study is indeed unusually high as compared to aerobic methylotrophes, but not untypical for methanogens grown on CO₂. It explains the low biomass yield. Based on the information available on the energetic metabolism and on an ATP balance, the biomass yield can be predicted to be approximately in the range of the experimentally determined value. The fact that the exothermicity exceeds vastly even the Gibbs energy change can be explained by a dramatic entropy decrease of the catabolic reaction. Microbial growth characterized by entropy reduction and correspondingly by unusually large heat production may be called entropy‐retarded growth. © 1999 John Wiley & Sons, Inc. Biotechnol Bioeng 64: 74–81, 1999.     

Conformational analysis of acetyl-L-phenylalanine p-acetyl and p-valeryl anilides

Empirical force-field calculations and ir and ¹H-nmr spectra indicate that five-membered (C₅) and seven-membered (C) hydrogen-bonded rings are the preferred conformations of acetyl-L -Phe p-acetyl and p-valeryl anilides in nonpolar media. The C₅/C ratio was found to be dependent on the dryness of the solute and the solvent. This fact and the results from conformational-energy calculations suggest that a molecule of water participates in the stabilization of the C conformation.     

The thermodynamics of solvent exchange

A model for solvation in mixed solvents, which was developed for the free energy and preferential interaction [J. A. Schellman (1987), Biopolymers, Vol. 26, pp. 549–559; (1990), Biophysical Chemistry, Vol. 37, pp. 121–140; (1993), Biophysical Chemistry, Vol. 45, pp. 273–279], is extended in this paper to cover the thermal properties: enthalpy, entropy, and heat capacity. An important result is that the enthalpy of solvation H? responds directly to the fraction of site occupation. This differs from the free energy ? and preferential interaction Γ₃₂, which are measures of the excess binding above a random distribution of solvent molecules. In other words, the enthalpy is governed by K while ? and Γ₃₂ are governed by (K ? 1) where K is the equilibrium constant on a mole fraction scale [Schellman (1987)]. The solvation heat capacity C?p consists of two term: (1) the intrinsic heat capacity of species in solution with no change in composition, and (2) a term that accounts for the change in composition that accompanies solvent exchange. Binding to biological macromolecules is heterogeneous but experiementalists must use binding isotherms that assume the homogeneity of sites. Equations are developed for the interpretation of the experimental parameters (number of sites n_exp, equilibrium constant K_exp, and enthalpy, Δh_exp), when homogeneous formulas are applied to the heterogeneous case. It is shown that the experimental parameters for the occupation and enthalpy are simple functions of the moments of the distribution of equilibrium constants over the sites. In general, n_exp is greater than the true number of sites and K_exp is greater than the average of the equilibrium constants. The free energy and preferential interaction can be fit to a homogenious formula, but the parameters of the curve are not easily represented in terms of the moments of distributions over the sites. The strengths and deficiencies of this type of thermodynamic model are discussed. © 1994 John Wiley & Sons, Inc.     

Kinetics of ethidium's intercalation in packaged bacteriophage T7 DNA: effects of DNA packing density

The kinetics of ethidium's intercalative binding to DNA packaged in bacteriophage T7 and two T7 deletion mutants have been determined, using enhancement of fluorescence to quantitate binding. At a constant ethidium concentration, the results can be described as first-order binding with two different rate constants, k (= k₁ + k_?1) and k (= k₂ + k_?2). The larger rate constant (k) was at least four orders of magnitude smaller than the comparable first-order forward rate constant for binding to DNA released from its capsid. At 25°C values of k decreased as the amount of DNA packaged per internal volume increased. This latter observation indicates that the rate of ethidium's binding to packaged T7 DNA is limited by an event that occurs inside of the DNA-containing region of T7, not by the crossing of T7 capsid's outer shell. Arrhenius plots of kM are biphasic, indicating a transition for packaged DNA at a temperature of 20°C. The data indicate that k s are limited by either sieving of ethidium during its passage through the packaged DNA or subsequent hindered intercalation.     

Relationships Between Kendall's Coefficient of Concordance and a Nonparametric Measure of Phenotypic Stability with Implications for the Consistency in Rankings as Affected by Variance Components

For a two-way classification with rows (= genotypes) and columns (= environments) relationships between Kendall's coefficient of concordance W and a nonparametric measure of phenotypic stability S (= variance among the ranks over the environments) are presented. This provides an analogy between Wricke's ecovalence and the sum of squares of genotype-environment interactions on the parametric side and S and Kendall's W on the nonparametric side: S is a genotype's contribution to the discordance (1 — W) in the data set, while Wricke's ecovalence is a genotype's contribution to the genotype-environment interaction sum of squares. Genotype x environment interactions may lead, but must not necessarily lead to non-identical rank orders of the genotypes in different environments (crossover versus noncrossover interactions). When does interaction become rank interaction? The similarity of the rank orders (measured by W) can be approximately expressed by the ratio of the genotypic variance and sum of genotypic variance plus residual variance. For the consistency of rankings this relation leads to an approximate test of significance which is based on variance components. Finally, a numerical example is given using grain yield data of 20 genotypes (varieties) of winter wheat in 10 environments (locations) from German registration trials.     

A population genetic study of the Banks and Torres Islands (Vanuatu) and of the Santa Cruz Islands and polynesian outliers (Solomon Islands)

As part of a multidisciplinary survey of populations in the Banks and Torres Islands of Vanuatu and the Southern and Central Districts of the Solomon Islands, nearly 2,400 persons have been tested for ABO blood groups and a number of serum protein and red cell enzyme genetic marker systems. For the ABO system, the populations are characterized in general by high gene O and low gene B frequencies except in two of the Polynesian Outlier Islands, Rennell and Bellona, which have high frequencies of B. Among the serum proteins, several alleles have distributions indicating significant movement of people between islands. These include Albumin ^{New Guinea} and the transferrin alleles Tf, and Tf, and Tf. Similar specific alleles for red cell enzymes also show distributions reflecting interisland population movement as well as contact with persons from outside the southern Pacific region. Examples are ACP in the acid phosphatase system, PGM and PGM, PGM and PGM, PGK⁴ and also HbJ^Tongariki. The data available for 11 polymorphic systems were used to generate genetic distances. Of the four Polynesian Outlier Islands, Anuta is most remote genetically, with Rennell and Bellona also relatively isolated. The fourth Polynesian Outlier, Tikopia, occupies a position genetically close to the Melanesian populations of the Banks and Torres Islands and the southern Solomons. The history of early European contact and voyaging in the Pacific, as well as archaeological and linguistic evidence and local legends, indicate that significant movements of people occurred between islands and provided opportunities for genes to be introduced from Europeans, Africans, and Asians. The genetic marker studies give evidence for genes from all these sources, though at a low level. Despite this admixture, the Polynesian Outlier and Melanesian populations have preserved their own distinctive genetic patterns.     

Apparent molar volumes of some amino acids and peptides in aqueous urea solutions

Densities of solutions of several α-amino acids and peptides in 3 and 6m aqueous urea solvents have been determined at 298.15 K. These data have been used to evaluate the infinite-dilution apparent molar volumes of the solutes and the volume changes due to transfer (V ) of the α-amino acids and peptides at infinite dilution from water to aqueous urea solutions. The sign and magnitude of the V values have been rationalized in the framework of Friedman's cosphere-overlap model. The V values for the glycyl group (? CH₂CONH? ) and alkyl side chains have been estimated.     

The influence of environmental conditions on the macromolecular composition of Candida utilis

Glucose-limited chemostat cultures of Candida utilis were cultivated at various pH levels (3.0–7.5), temperatures (15–37.5°C), dilution rates (0.06–0.42 hr^?1), and with different nitrogen sources (NH and NO). The ratio of total nucleic acid to protein increased with increase in dilution rate at constant temperature and decreased with increase in temperature at constant dilution rate. The pattern of these variations is consistent with the hypothesis that the nucleic acid to protein ratio is a function of the ratio of the actual dilution rate to the critical dilution rate corresponding to each one of the cultivation temperatures. This ratio is called “reduced dilution rate.” A basis is proposed on which various microorganisms may be compared with respect to the ratios of cell protein to nucleic acid, RNA, ribosomal RNA, and polysomes.