bluuebunny/crossref_metadata_2025 huggingface dataset:
Annual Summary of Vital Statistics—2003
Authors: Martin Joyce A. , Kochanek Kenneth D. , Strobino Donna M. , Guyer Bernard , MacDorman Marian F.
Published: March 2005

The crude birth rate rose slightly in 2003 to 14.1 births per 1000 population, from 13.9 in 2002. The 2002 rate was the lowest ever reported for the United States. The total number of births and the fertility rate (66.1) also increased. The birth rate for teenaged mothers dropped 3% to another record low in 2003, to 41.7 per 1000 females aged 15 to 19 years. The teenage birth rate has fallen by one third since 1991. The birth rate declined for women 20 to 24 years old but rose for women aged 25 to 44 years. The number, rate, and proportion of births to unmarried women all increased in 2003.

Smoking during pregnancy declined to 11%, down from 19.5% in 1989. Prenatal care utilization improved slightly for 2003; 84.1% of women began care in the first trimester of pregnancy. The cesarean delivery rate jumped 6% to 27.6% for another US high. The primary cesarean rate rose 6%, and the rate of vaginal birth after a previous cesarean delivery plummeted 16% from 2002 to 2003. The percent of infants delivered preterm continued to rise (12.3% in 2003). The preterm birth rate is up 16% since 1990. The percentage of children born at low birth weight rose slightly in 2003 to the highest level reported since 1970 (7.9%). The twinning rate increased, but the rate for triplet/+ births declined slightly between 2001 and 2002. Multiple births accounted for 3.3% of all births in 2002.

The infant mortality rate rose to 7.0/1000 live births in 2002 from 6.8 in 2001, marking the first increase in this rate in >4 decades. Increases were distributed fairly widely across age, racial/ethnic groups, and geographic areas. The rise in infant mortality was attributed to increases in <750-g births in both singleton and multiple deliveries. Although the downward trend in infant mortality rates in many developed nations may have stabilized, the United States still ranked 27th among these nations in 2001.

Expectation of life at birth reached a record high of 77.3 years for all gender and race groups combined in 2002. Death rates in the United States continue to decline. Between 2001 and 2002, death rates declined for the 3 leading causes of death: diseases of heart, malignant neoplasms, and cerebrovascular diseases. Death rates for children 1 to 19 years old decreased by 8% for suicide; the death rate for chronic lower respiratory diseases increased by 33% in 2002. Rates for unintentional injuries and homicide did not change significantly for children aged 1 to 19 years. A large proportion of childhood deaths continues to occur as a result of preventable injuries.
Synthesis and Characterization of the Off‐Stoichiometric Compound (W0.8Al0.2)C0.7
Authors: Yan J. , Ma X. , Zhao W. , Tang H. , Zhu C. , Cai S.
Published: March 2005

A new solid solution of Al in WC with carbon vacancieshas been synthesized by a mechanical alloying and high pressure (MAAHP) technique. ESEM‐EDAX is used to investigate the morphology and the quantitative material composition of the sintered sample. The Vickers micro hardness of the sample is tested to be 3000 kg mm–2 that is much harder than WC, while the density of sintered body is about 12.649 g cm–3, which is lighter than WC.
TEM Investigation of Spherical Primary Si Particles in As‐Quenched Al60Si40 Ribbons
Authors: Zhao Z. K. , Yao K. F.
Published: March 2005

Al‐Si alloys with high Si content are attractive in engineering application, especially in automobile industry. Their mechanical properties and machinability are quite sensitive to the morphology of the primary Si phase. In order to improve the mechanical properties of Al‐Si alloys, optimizing the shape and distribution of the primary Si are quite important. In this paper, the microstructure of the primary Si in the Al60Si40 alloys prepared by rapid solidification method has been studied used the transmission electron microscope (TEM).
Acid‐dependent selective formation of trans‐aryl(germyl)ethenes or 1‐aryl‐1‐germylethenes by the protodestannylation of (Z)‐germyl(stannyl)ethenes
Authors: Nakano Taichi , Senda Yoshiya , Fukaya Kayo , Sugiuchi Naoyuki , Ni‐imi Shinobu , Takahashi Yutaka , Kurihara Hiroyuki
Published: March 2005

Protodemetallation of (Z)‐1‐aryl‐2‐germyl‐1‐stannylethenes using hydrochloric acid in the presence of tetraethylammonium chloride (TEACl) proceeds at 0 °C with retention of configuration to afford trans‐1‐aryl‐2‐germylethenes with high yields. In contrast, the reaction using hydroiodic acid in the presence of tetrabutylammonium iodide (TBAI) brings about a novel 1,2‐germyl migration accompanying the destannylation to form 1‐aryl‐1‐germylethenes. These 1,2‐germyl migration products result with especially high selectivity from adducts bearing a substituent at the ortho‐ or para‐position on the aromatic ring in the adduct. However, the germyl(stannyl)ethenes bearing a substituent at the meta position on the ring produce a regioisomeric mixture of a trans‐germylethene and a 1‐aryl‐1‐germylethene. Copyright © 2005 John Wiley & Sons, Ltd.
Unprecedented self‐assembled cyclic hexamer of ferrocenyldimethylsilanol, [FcSiMe2OH]6 (Fc = (η5‐C5H5)Fe(η5‐C5H4))
Authors: Sharma Hemant K. , Cervantes‐Lee Francisco , Haiduc Ionel , Pannell Keith H.
Published: March 2005

Ferrocenyldimethylsilanol, FcSiMe2OH, Fc = (η5‐C5H5)Fe(η5‐C5H4), features a self‐assembled (through intermolecular hydrogen bonding) cyclohexameric supermolecule with a chair conformation. Copyright © 2005 John Wiley & Sons, Ltd.
Mixed aryl–alkyl organotin compounds, ArnMeSnCl3−n (Ar = RC6H4, R = H, ethyl, i‐propyl, t‐butyl; n‐hexyl, n‐octyl) and the effect of R upon antibiotic activity
Authors: Kapoor Ramesh N. , Apodaca Paula , Montes Miguel , Gomez Fabiola D. , Pannell Keith H.
Published: March 2005

The synthesis of a new series of arylmethyltin chlorides is reported, ArnMeSnCl3−n (Ar = RC6H4, R = H, ethyl, i‐propyl, t‐butyl; hexyl, octyl). The synthesis involves initial formation of triarylmethyltin compounds, Ar3MeSn, via Grignard techniques followed by HCl–Et2O aryl group cleavage, preferably in a stepwise manner. Preliminary biological activity against Staphylococcus aureus illustrates the importance of the para‐alkyl substituents and reinforces that an optimal hydrophobic character is needed for maximum efficacy. Copyright © 2005 John Wiley & Sons, Ltd.
Postprostatectomy adjuvant versus salvage radiotherapy
Authors: Jani Ashesh B. , Kao Johnny
Published: March 2005

BACKGROUND

Radiotherapy (RT) has been used with success after radical retropubic prostatectomy (RRP), both in the adjuvant and salvage settings. The purpose of the current investigation was to systematically compare adjuvant versus salvage RT in a manner that incorporates both treatment efficacy and complications.

METHODS

A literature review was performed of reports of post‐RRP salvage and adjuvant RT, and 12 trials comprising 1060 patients met the appropriate inclusion criteria. The biochemical failure‐free survival in each study/arm was tabulated, and these values were entered into a model to compute an unadjusted number‐needed‐to treat (NNT). RT complications were then considered, accounting for differences in toxicity incidences in the salvage versus adjuvant setting, to compute complication‐adjusted NNTs. In all the trials, the signs and magnitudes of the NNTs obtained were used to compare adjuvant with salvage RT.

RESULTS

The absolute NNT analysis showed an advantage of adjuvant compared with salvage RT. After adjustment for RT complications, however, the advantage shifted to salvage RT. This transition point from superiority of adjuvant RT to superiority of salvage RT was sensitive to the estimated incidence and severity of RT side effects.

CONCLUSIONS

Adjuvant post‐RRP RT was advantageous in comparison to salvage RT if the side effects of RT were estimated to be negligible. However, with moderate incidence/severity of RT side effects, salvage RT was advantageous. The findings herein must be tested in a prospective study in which both health‐related quality of life and cancer control are documented in patients receiving adjuvant versus salvage post‐RRP RT. Further work is needed to better estimate parameters entered into the model to determine the precise transition point between adjuvant and salvage RT with modern RT techniques. Cancer 2005. © 2005 American Cancer Society.
Increased plasma MMP9 in integrin α1‐null mice enhances lung metastasis of colon carcinoma cells
Authors: Chen Xiwu , Su Yan , Fingleton Barbara , Acuff Heath , Matrisian Lynn M. , Zent Roy , Pozzi Ambra
Published: March 2005

Inhibitors of matrix metalloproteinases (MMPs) were developed as anticancer agents based on the observation that MMPs facilitate local tumor spread and metastasis by promoting matrix degradation and cell migration. Unfortunately, these inhibitors were unsuccessful in the clinical treatment of several cancers, including lung cancer. A possible reason contributing to their failure is that MMP activity is critical for the generation of inhibitors of tumor angiogenesis, including angiostatin. Thus, MMPs might play opposing roles in tumor vascularization and invasion. To determine which effect of elevated MMP levels dominates in the progression of metastatic cancer, experimental lung metastasis assays were performed in integrin α1‐null mice, a genetic model for increased plasma levels of MMP9 and MMP9‐generated angiostatin (Pozzi et al., Proc. Natl. Acad. Sci. USA 2000;97:2202–7). We show that while the number of lung colonies in integrin α1‐null mice was significantly increased compared to their wild‐type counterparts, tumor volume was markedly reduced. In vivo treatment with the MMP inhibitor doxycycline resulted in a significant decrease in the number of lung colonies in both genotypes, but the tumors that formed were bigger and more vascularized. Increased tumor vascularization paralleled decreased plasma levels of MMP9 and consequent decreased angiostatin synthesis. These results demonstrate that while inhibition of MMPs prevents and/or reduces tumor invasion and lung metastasis, it has the paradoxical effect of increasing the size and vascularization of metastatic tumors due to decreased generation of inhibitors of endothelial cell proliferation. The continued growth of these large well‐vascularized tumors may explain the poor efficacy of MMP inhibitors in lung cancer clinical trials. © 2005 Wiley‐Liss, Inc.
Correlation between fetal gastric size and amniotic fluid volume
Authors: Ben‐Haroush Avi , Yogev Yariv , Peled Yoav , Bar Jacob , Hod Moshe , Pardo Josef
Published: March 2005

Purpose

Since abnormal conditions of the fetal digestive tract may alter both amniotic fluid volume and fetal gastric volume, we sought to determine whether amniotic fluid volume is correlated with fetal gastric volume in normal pregnancy.

Methods

A total of 280 fetal gastric size measurements were made prospectively from routine sonographic examinations of women with normal singleton pregnancies between 16 and 42 weeks of gestation. The fetal stomach was defined as the largest area including the pyloric site on transverse or oblique real‐time sonographic scans. Gastric volume was calculated according to the formula for a prolate ellipsoid. The amniotic fluid index (AFI) was used for the evaluation of amniotic fluid volume.

Results

Both fetal gastric volume and AFI were significantly correlated with gestational age (R2 = 0.422 and R2 = 0.128, respectively). Only a weak correlation was found between gastric volume and AFI (R2 = 0.036, p < 0.001). On multivariate linear regression analysis adjusting for gestational age and fetal biometric measurements, gastric volume was not an independent and significant predictor of AFI.

Conclusions

Although sonographically determined fetal gastric volume measurements appear to be useful in the assessment of fetal digestive tract anomalies, fetal gastric volume has no clinically significant effect on the amniotic fluid volume in normal pregnancy. © 2005 Wiley Periodicals, Inc. J Clin Ultrasound 33:119–122, 2005
Synthesis and characterization of novel nano size electroactive poly 4,4′‐diaminodiphenyl sulphone
Authors: Manisankar P. , Vedhi C. , Selvanathan G.
Published: March 2005

The modification of electrodeposited polyaniline film by subsequent electrodeposition of 4,4′‐diaminodiphenyl sulfone (DDS) leads to a new material having nanostructure. The coated polymer films were treated with various pH solutions. The film adherent characteristics and surface morphology were studied using SEM. The electrochemically synthesized polyDDS revealed good redox behavior. The DDS was also polymerized by the chemical oxidation method using potassium persulphate. The polymer was characterized by UV‐Vis and FTIR spectral studies. The formation of polymer through the NH group was understood from the single NH stretching vibrational frequency at 3459 cm−1. The X‐ray diffraction studies revealed the formation of nano sized (28 nm) crystalline polymer. The conductivity of the polymer was determined to be 1.07 × 10−4 S.cm−1. The solubility of the chemically polymerized powder was ascertained, and polyDDS showed good solubility in DMF and DMSO. © 2005 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 43: 1702–1707, 2005
Synthesis of Davankov‐type hypercrosslinked resins using different isomer compositions of vinylbenzyl chloride monomer, and application in the solid‐phase extraction of polar compounds
Authors: Fontanals Núria , Cortés Jaime , Galià Marina , Maria Marcé Rosa , Cormack Peter A. G. , Borrull Francesc , Sherrington David C.
Published: March 2005

Two different gel‐type resins have been prepared by suspension polymerization using 2 wt % divinylbenzene (DVB) with either p‐vinylbenzyl chloride (pVBC) or a mixture of VBC isomers (∼ 70% m‐; ∼ 30% p‐). Significant difference in the chlorine content was observed, which was attributed to a more favored hydrolysis process when p‐VBC was used. The presence of hydroxyl groups has been confirmed by elemental microanalytical data and solid‐state 13C cross‐polarization/magic angle spinning (CP‐MAS) nuclear magnetic resonance (NMR) spectra. Hypercrosslinked resins were prepared from both gel‐type precursors by treatment with FeCl3 in 1,2‐dichloroethane (DCE) at 80 °C. The resultant resins showed differences in specific surface area and degree of hydrophilicity. The performance of the hypercrosslinked resins was evaluated in solid‐phase extraction (SPE) of polar compounds, and better results were obtained for the hypercrosslinked resin prepared from p‐VBC that combines a relatively high specific surface area (908 m2 g−1) and somewhat higher oxygen content (3.96 wt % O). © 2005 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 43: 1718–1728, 2005
Characterization of a new 60 kDa apical protein of Plasmodium falciparum merozoite expressed in late schizogony
Authors: Grellier Philippe , Précigout Eric , Valentin Alexis , Carcy Bernard , Schrével Joseph
Published: March 2005

Summary— Immunological cross‐reactivity studies between the Apicomplexa Babesia divergens and Plasmodium falciparum allowed usto identify a P falciparum 60 kDa protein (Pf60) using an antiserum directed against a B divergens 37 kDa culture‐derived exoantigen. In immunofluorescence assays (IFA), Pf60 appears as a doublet of fluorescent spots associated to the apical pole of merozoites. The doublet co‐locates with two rhoptry components: the protein RAP‐1 and the 140/130/110 (105) kDa rhoptry protein complex suggesting the rhoptry location of Pf60. The biosynthesis of Pf60, established by labeling experiments with [35S]methionine on synchronized cultures, and by immunofluorescence detection, occurred during late schizogony. The physico‐chemical properties of Pf60, the absence of identified precursor forms and the absence of co‐precipitation with other proteins indicated a new class of rhoptry protein. Pf60 was detected in all the different geographic P falciparum strains so far tested, with a slight variability in molecular mass ranging from 58 to 60 kDa. During the invasion process of erythrocytes by merozoites, the IFA showed the presence of the Pf60 in the apex of free merozoites, but not in invading merozoite, as well as in new ring‐infected erythrocytes. Furthermore, immunoprecipitation assays indicated the presence of Pf60 in the culture medium, and its absence in new ring‐infected erythrocytes. All together these results suggest a possible involvement of the Pf60 protein in the invasion process.
TRADE-OFFS BETWEEN FORAGE YIELDS AND FEED QUALITY OF LABLAB PURPUREUS, AND MILK YIELDS IN RELATION TO PLANTING AND HARVESTING SCHEDULES
Authors: AGYEMANG K. , OKIKE I. , MAKUN H. J. , MAGAJI S. O.
Published: March 2005

This study, designed to investigate trade-offs between forage yields, feed quality and labour use in fitting a newly introduced crop, Lablab purpureus, into an existing farming system, adopted distinct applied and adaptive research approaches. The applied research component compared forage yields, nutrient contents and feeding qualities of L. purpureus harvested at 100, 114, 128 or 142 d after planting in an on-farm trial. The adaptive research investigated how farmers incorporated L. purpureus cultivaton into their farming calendars given the constraints of labour for planting and harvesting.

Harvested dry matter yields from the on-farm experiment were 1.28, 1.98, 1.74 and 1.44 t ha−1. The resultant hays were fed to four groups each of five Bunaji lactating cows in an on-farm feeding trial over a 10-week period. Ranking the various harvests for yield, the hays for feed value, animal responses to the feed consumed and the quality of animal products on a linear scale (1 to 4) to estimate a composite utility index, showed that harvesting at 114 d after planting gave the most desirable overall benefits.

The survey for adaptive research showed bimodal rainfed and irrigated cultivation patterns for L. purpureus. Peaks were observed in September (31% of farmers) and November (24%) for planting, and in December (39.4%) and March (15%) for harvesting. Forage yields were, however, highest for the 3% of farmers who afforded labour to plant in July and harvest in October. The lack of congruence between highest forage yield periods and the peaks of cultivation of L. purpureus by farmers in the study area was mostly explained by very significant differences in the cost of hired and household labour in different months. Thus, socio-economic and cultural factors, including stakeholders' participation, need to be considered along with applied research results when recommending forage species for farmers' adoption.
Sugar Separations on a Pilot Scale by Continuous Annular Chromatography
Authors: Byers Charles H. , Sisson Warren G. , Decarli Joseph P. , Carta Giorgio
Published: March 2005

A pilot‐scale continuous annular chromatograph has been developed for scale‐up and optimization studies. The chromatographic apparatus consists of a slowly rotating annular bed of sorbent material. The feed mixture which is to be separated is continuously introduced at a stationary point at the top of the bed, while eluent flows over the remainder of the annulus. The rotation of the sorbent bed coupled with the elution development induced by the downflow of eluent causes the feed components to appear as individual helical bands, each of which has a characteristic, stationary exit point. The separation of sucrose, glucose, and fructose, using the calcium form of a cation‐exchange resin, was investigated using water as the eluent. Experiments were carried out both with a synthetic mixture of the sugars as well as with an industrial feedstock. The separation was explored first in a bench‐scale conventional fixed‐bed column and then in the pilot‐scale continuous chromatograph. Factors such as column loading, feed‐to‐eluent ratio, and feed concentration were investigated both experimentally and theoretically. The results of bench‐scale experiments were carried out in the limit of infinite dilution, almost perfectly scaled to the pilot unit when loading was low and feed mixtures were of low viscosity. Very significant deviations from ideality were, however, observed at high feed loadings, when the feed viscosity exceeded that of the eluent by more than a factor of 2.
Affinity Cross‐Flow Filtration: Some New Aspects
Authors: Herak David C. , Merrill Edward W.
Published: March 2005

A variety of affinity cross‐flow filtration (ACFF) experiments were conducted to evaluate the technique as a means of effectively separating biomolecules. Agarose particles, which contained a specific affinity ligand for the targeted protein, were used as “affinity escorts” in the ACFF process. Both conventional agarose particles (40–150; μm) and small agarose particles (Superose, 11–15 μm.) were used as the basis for affinity escorts. Batch ACFF washing experiments were conducted in both constant‐volume and volume‐reduction modes and compared with model predictions. The different affinity ligand‐adsorbate systems studied include Cibacron Blue and human serum albumin, Cibacron Blue and lysozyme, protein A and immunoglobulin G, and concanavalin A and horseradish peroxidase.
Selective Extraction Using Functionalized Poly(organosiloxanes)
Authors: England R. , Abed‐Ali S. S. , Brisdon B. J.
Published: March 2005

There are two major separation problems that frequently impede the successful isolation of fine chemicals and biological materials from aqueous media. These concern either (a) the selective recovery of low concentrations of products from complex mixtures or (b) the purification of high‐value products by selective and efficient removal of contaminants. The recovery of biotransformation products with frequently small chemical and physical differences between reactant and product, from bioreactor broths containing low product concentrations as well as nutrients and extraneous salts, typifies the former problem. We have used fluid polymers functionalized with molecular receptors capable of interacting specifically with the required products in the fermenter broth, to provide a basis for separation. Thus, attachment of various crown ethers to a hydrophobic polysiloxane backbone via an alkyl spacer chain yields functionalized, liquid, linear polymers of the type Me3Si(OSiMe2)x(OSiMe(CH2)n — crown)yOSiMeg3, which are able to preferentially complex, and hence extract, specific types of amines from aqueous mixtures. This molecular recognition technique has been successfully applied to the separation of N‐demethylated drug molecules produced by Cunninghamella sp. in bioreactors. The efficiency of extraction from the fermenter broth and recovery of the product from the functionalized polymer has been determined for this system. Attachment of other receptors to polysiloxane fluids is currently being investigated in order to extend the range of application of this class of discriminating extractant. Initial studies have also been made on applying the technique to problem b, using similarly functionalized polymers to selectively decontaminate complex macromolecular systems. Results indicate that amine contaminants can be removed from some proteins in a single‐batch extraction but that an interaction occurs on direct contact between the protein and functionalized polymers. Studies utilizing supported liquid membranes for the separation of various amines and amino acids are currently under investigation, and preliminary results show that the method is superior to direct liquid/liquid extraction.
Shear Sensitivity of Hybridoma Cells in Batch, Fed‐Batch, and Continuous Cultures
Authors: Petersen Jonathan F. , Mclntire Larry V. , Papoutsakis Eleftherios Terry
Published: March 2005

Previously, we observed that CRL‐8018 hybridoma cells were more sensitive to well‐defined viscometric shear during the lag and stationary phases than during the exponential phase of batch cultures. Some potential hypotheses for explaining the increase in shear sensitivity are (1) nutrient limitations that result in a decrease in production of specific cellular components responsible for the mechanical strength of the cell, (2) nutrient limitations that lead to synchronization of the culture in a cell cycle phase that is more sensitive to shear, or (3) a link between cell growth and shear sensitivity, such that slowly growing cells are more sensitive to shear. Here, the duration of the exponential phase was increased with use of fed‐batch, and the effect on shear sensitivity of the cultures was measured with a viscometric technique. Extension of exponential growth resulted in an increased period during which the cells were insensitive to shear. Additionally, the shear sensitivity of the cells was constant over a wide range of growth rates and metabolic yields in chemostat cultures. These observations suggest that as long as the cells are actively (exponentially) growing, their shear sensitivity does not depend on the growth rate or metabolic state of the cell as expressed by metabolic yields. Thus, hypothesis 3 above can be dismissed.
Biosynthesis of p‐Hydroxylated Aromatics
Authors: Salvo Joseph J. , Mobley David P. , Brown Daren W. , Caruso Lisa A. , Yake Andrew P. , Spivack Jay L. , Dietrich David K.
Published: March 2005

A bioconversion process with Aspergillus parasiticus has been developed to enhance p‐hydroxylation activities with respect to aromatic substrates such as m‐terphenyl. Through modified growth conditions, we are improving product concentration, while streamlining fermentation and downstream processing requirements. We routinely carry out biosynthetic conversions with m‐terphenyl at the 300‐L scale and produce over 200 g of 4,4″‐dihydroxy‐m‐terphenyl at a concentration of 0.7 g/L in the fermentation vessel.
Simulation and Control of Glucose Concentration in Hot‐Water Blanching of Potatoes
Authors: Tomasula P. M. , Kozempel M. F. , Craig J. C.
Published: March 2005

The methodology for online monitoring and computer control of glucose concentration in potatoes during hot‐water blanching is demonstrated. Process conditions, control parameters, and setpoints are determined with the aid of an unsteady‐state model. Glucose concentration in the blanch water is monitored and maintained at a setpoint value by adjustment of the fresh water flow rate to the blancher, while the glucose concentration of the potatoes is calculated by mass balance. Control is effected through feedback and anticipatory feedback control. Results indicate that glucose concentration in potatoes exiting the blancher may be maintained within 10% of the setpoint value. The use of computers to control glucose concentration in potatoes should prove useful in the control of the color and therefore the quality of processed potato products.
Sizing Biological Samples by Photosedimentation Techniques
Authors: Middelberg Anton P. J. , Bogle I. David L. , Snoswell Mark A.
Published: March 2005

The performance of the Joyce‐Loebl disk centrifuge in the sizing of Escherichia coli cells, protein inclusion bodies, and cell debris is evaluated. The need for a density gradient that extends throughout the entire spin fluid is highlighted, and a set of standard conditions that fulfill this requirement is defined. E. coli cells experience a reduction in their Stokes diameter when exposed to ethanol, indicating that a spin‐buffer fluid combination such as glycerol‐water is to be preferred for the sizing of bacteria. The instrument baseline is influenced by the presence of particles, and a method of estimating the baseline is described. The sizing of small particles is further complicated by baseline drift due to temperature sensitivity of the optical yoke. An analysis of diffusion in the spin fluid is conducted, and an expression for the sedimentation:diffusive flux ratio is derived. For the current samples, it is shown that diffusion within the spin fluid does not lead to significant errors for 0.15‐μm particles, whereas the phenomenon may be significant at the manufacturer's size limit of 0.01 μm.
N‐Laurylbiotinamide as Affinity Surfactant
Authors: Coughlin Robert W. , Baclaski Jeffrey B.
Published: March 2005

N‐laurylbiotinamide (NLB), which retains strong affinity for the protein avidin, was synthesized from biotin and N‐laurylamine via the biotin ester of N‐hydroxysuccinimide and characterized by NMR. When the synthesized NLB was used as a cosurfactant with AOT to form a reverse micellar system in isooctane, it was found to extend the pH range over which avidin can be transferred from a continuous aqueous solution to the reverse‐micellar phase. This behavior is similar to that already reported for a different affinity surfactant, n‐octyl β‐D‐glucopyranoside.
Improved Adsorption to Starch of a β‐Galactosidase Fusion Protein Containing the Starch‐Binding Domain from Aspergillus Glucoamylase
Authors: Chen Luojing , Ford Clark , Kusnadi Ann , Nikolov Zivko L.
Published: March 2005

We have previously shown (Chen et al., 1991) that a β‐galactosidase (β‐gal) fusion protein (BSB133) containing 133 amino acids (aa) from the C‐terminus of Aspergillus glucoamylase (GA) adsorbs strongly to starch compared to β‐gal, due to the presence of the GA starch‐binding domain. We have now made deletions at the N‐terminus of this 133‐aa region to test the minimal size required for starch binding of β‐gal fusion proteins. Three fusion proteins (BSB119, BSB103, and BSB80) were genetically engineered, containing 119, 103, and 80 C‐terminal aa from GA, respectively. The fusion proteins were expressed in Escherichia coli and purified. Purified BSB119 adsorbed to native starch at least 2‐fold more strongly than did BSB133 or fusion proteins with shorter tails. Adsorption isotherms generated over a wide range of initial concentrations indicated a 10‐fold difference in the loading capacity of starch for BSB119 (36.5 mg of protein/g of starch) compared to β‐gal (3.7 mg of protein/g of starch). Adsorption constants calculated from the initial slopes of the isotherms indicated a nearly 30‐fold difference in affinity to starch for BSB119 (Kad = 63 mL/g of starch) compared to β‐gal (Kad = 2.3 mL/g of starch). BSB119 in the presence of crude enzyme extracts also bound to starch with a high affinity compared to a β‐gal control. Potential applications of the starch‐binding tail include enzyme immobilization to starch or recovery and purification of target proteins from crude extracts.
Mathematical Modeling of Biochemical Changes during Processing of Liquid Foods and Solutions
Authors: Datta Ashim K
Published: March 2005

Thermal processing of liquid food materials and biochemical solutions always results in a distribution of biochemical changes such as bacterial and nutrient concentrations due to variation in temperature history from one liquid element to another. To estimate such distributions, the liquid elements need to be “tagged” and followed, which is computationally difficult for most flow situations of interest. This paper describes a computational procedure for obtaining the full range of biochemical changes during processing without explicitly following liquid elements. By use of the method developed, transient distributions of bacterial death have been obtained for sterilization of liquid in a nonagitated cylindrical enclosure heated from all sides. The conventional procedure of obtaining sterilization based on temperature at the slowest heating zone is shown to be lower than the actual least sterilization of a real fluid element in the system. Nutrient distributions can be similarly calculated by following this method.
Prediction of Moisture Diffusivity in Granular Materials, with Special Applications to Foods
Authors: Vagenas G. K , Karathanos V. T
Published: March 2005

The effective diffusion coefficient of two granular starch materials was determined from drying experiments by using the method of slopes. A method was developed, which permits the combination of the diffusion coefficient of the solid phase and the binary diffusion coefficient of water vapor in air for the prediction of the effective moisture diffusivity of the system. Several structural models of the literature were used to predict the effective moisture diffusivity of the two granular starches in the ranges of moisture content 0.0‐0.6 kg of water/kg of dry solids, porosity 0.0‐0.6, and temperature 20–100 °C. The results showed that the structural models can be successfully used for the prediction of the effective moisture diffusivity, and especially the Behrens model showed considerable merit. The diffusion coefficient of the solid phase (granule) was estimated by fitting the theoretical model to the experimental data.
Evaluation of Immunoglobulins from Plant Cells
Authors: Hein Mich B , Tang Ying , Mcleod Donald A , Janda Kim D , Hiatt Andrew
Published: March 2005

Expression of cDNA constructs encoding full‐length mouse immunoglobulin chains with their native leader sequences or fusion constructs substituting the native leader with a pre‐pro sequence derived from Saccharomyces cerevisiae yielded blocked N‐termini on the γ chain or the correct amino terminal sequence on the mature k chain. Lectin binding assays revealed that assembled immunoglobulin complexes contained a glycosylated heavy chain. The attached glycan was resistant to digestion by endoglycosidase H and its lectin binding pattern was distinguishable from that of the mammalian glycan. The results indicated processing of the immunoglobulin carbohydrate in the tobacco Golgi to yield a complex oligosaccharide. Secretion of antibody by protoplasts isolated from regenerated transgenic plants or from suspension callus cells was demonstrated by pulse‐chase labeling experiments. When purified, the tobacco‐produced antibody was found to possess the antigen binding and catalytic properties of the murine monoclonal antibody. Kinetic parameters (Km, Ki, Vmax, and kcat) of the tobacco‐derived antibody were comparable to those of the mouse‐derived antibody. The results in general show that the endomembrane system of tobacco cells possesses cognate mechanisms for the recognition of diverse leader sequences. These signals can be used to initiate the assembly, processing, and secretion by plant cells of complex foreign proteins.
Plasmid Stabilization of an Escherichia coli Culture through Cycling
Authors: Stephens M. L. , Christensen C. , Lyberatos G.
Published: March 2005

The problem of plasmid instability of fermentations that involve plasmid‐bearing recombinant organisms is dealt with in this work. Previous theoretical work demonstrated that under certain conditions (where plasmid‐bearing species are slower in responding to changes in the fermentation environment than the wild species) the washout of the plasmid‐bearing species can be prevented. In the sequel, Weber and San showed that cycling the dilution rate can delay the washout of plasmid‐bearing species for a plasmid‐bearing Escherichia coli culture. This work shows that it is indeed possible to secure the presence of the plasmid‐bearing species at all times through appropriate cycling.
Release and Recovery of Porcine Pepsin and Bovine Chymosin from Reverse Micelles: A New Technique Based on Isopropyl Alcohol Addition
Authors: Carlson A. , Nagarajan R.
Published: March 2005

After complete solubilization by the direct method, porcine pepsin was not released from AOT in isooctane reverse micelles even under aqueous‐phase conditions which would not ordinarily allow uptake. Similarly, bovine chymosin, once forward‐transferred at a pH below its isoelectric point, was not back‐transferred into an aqueous contact phase buffered at a pH value above its isoelectric point. These results show that there is significant hysteresis in the forward‐and backward‐transfer processes and further imply that kinetics, and not equilibrium, control uptake or release processes for these enzymes. The addition of 10–15 % isopropyl alcohol to the aqueous phase increases the rate of protein release dramatically and allows for nearly complete back‐transfer of porcine pepsin and 70% back‐transfer of bovine chymosin. IPA addition does not destroy the functional integrity of the system since forward transfer of bovine chymosin still occurs at pH values below (but not above) the pI of the protein.
Transient Association of the First Intermediate during the Refolding of Bovine Carbonic Anhydrase B
Authors: Cleland Jeffrey L. , Wang Daniel I. C.
Published: March 2005

Many proteins which aggregate during refolding may form transiently populated aggregated states which do not reduce the final recovery of active species. However, the transient association of a folding intermediate will result in reduced refolding rates if the dissociation process occurs slowly. Previous studies on the refolding and aggregation of bovine carbonic anhydrase B (CAB) have shown that the molten globule first intermediate on the CAB folding pathway will form dimers and trimers prior to the formation of large aggregates (Cleland, J. L.; Wang, D. I. C. Biochemistry 1990, 29, 11072–11078; Cleland, J. L.; Wang, D. I. C. In Protein Refolding; Georgiou, G., De‐Bernardez‐Clark, E., Eds.; ACS Symposium Series 470; American Chemical Society: Washington, DC, 1991; pp 169–179). Refolding of CAB from 5 M guanidine hydrochloride (GuHCl) was achieved at conditions ([CAB]f=10–33 μM, [GuHCl]f=1.0 M) which allowed complete recovery of active protein as well as the formation of a transiently populated dimer of the molten globule intermediate on the refolding pathway. A kinetic analysis of CAB refolding provided insight into the mechanism of the association phenomenon. Using the kinetic results, a model of the refolding with transient association was constructed. By adjusting a single variable, the dimer dissociation rate constant, the model prediction fit both the experimentally determined active protein and dimer concentrations. The model developed in this analysis should also be applicable to the refolding of proteins which have been observed to form aggregates during refolding.
An Amperometric Alcohol Sensor Based on Chemically Permeabilized Methylotrophic Microorganisms
Authors: Chen Jhy‐Chern , Naglak Thomas J. , Wang Henry Y.
Published: March 2005

Chemical permeabilization of the yeast Pichia pastoris NRRL Y‐11430 with 1.0% glycine and 0.1 % Triton X‐100 does not release intracellular alcohol oxidase but alters the cell envelope structure, allowing alcohol oxidase activity to be detected inside the cells (Naglak, 1990). An alcohol sensor based on the use of chemically permeabilized yeast cells was constructed. The sensor response time was within 2 min. The assay time was less than 30s by the initial rate method. The calibration curve for ethanol analysis was linear in the range 5–65 mg/L. The sensor response was almost unaffected by pH changes within the range pH 5–9. This alcohol sensor produced the highest sensitivity for methanol; the sensitivity for ethanol was about half that for methanol. The stability and other characteristics of this new alcohol sensor are also discussed in this work.
Highly Enantioselective Hydrolysis of (R,S)‐Phenylalanine Isopropyl Ester by Subtilisin Carlsberg. Continuous Synthesis of (S) ‐Phenylalanine in a Hollow Fiber/Liquid Membrane Reactor
Authors: Ricks Edith E. , Estrada‐Valdés Maria C. , Mclean Thomas L. , Iacobucci Guillermo A.
Published: March 2005

The resolution of racemic phenylalanine esters with esterases has been reexamined in relation to the development of a continuous process based on the use of hollow fiber/liquid membrane (SLM) reactors. The requirement for high enantioselectivity was met by phenylalanine isopropyl ester, whose (R)‐enantiomer was found refractory to the action of subtilisin Carlsberg, in water, pH 7.5, at 25 °C. The continuous feeding of (R, S)‐Phe‐O‐iPr‐HCl at 2 mm/h to 0.7 g/L subtilisin resulted in steady‐state reaction conditions that gave (S)‐Phe‐OH of 95% enantiomeric excess (ee) at the theoretical rate. The unreacted (R)‐Phe‐O‐iPr, which permeated out continuously, had 80% ee and was racemized quantitatively by heating with a salicylaldehyde catalyst in refluxing toluene. Doubling the feeding rate (4 mm/h) created excessive accumulation of (R)‐Phe‐O‐iPr in the reactor, which inhibited the enzyme through nonproductive binding, lowering the (S)‐Phe‐OH resolution rate to 68% of the theoretical rate. Similar experiments with (R, S)‐Phe‐OMe‐HCl gave (S)‐Phe‐OH of 9–48% ee, due to the enzymatic hydrolysis of (R)‐Phe‐OMe experienced during the runs. When α‐chymotrypsin was used for the continuous hydrolysis of (R, S)‐Phe‐O‐iPr, the isolated (S)‐Phe‐OH showed 67% ee, indicating that substantial hydrolysis of the (‐R)‐ester had occurred.
Activity Regeneration in Continuous Clostridium acetobutylicum Bioconversions of Glucose
Authors: Reardon Kenneth F. , Baily James E.
Published: March 2005

The regeneration of product‐forming activity by periodic nutrient feeding was studied in a continuous immobilized Clostridium acetobutylicum reactor system. Activity levels were increased by feeding ammonia and vitamins during selected intervals of the experiments; these compounds were not otherwise present in the medium. In contrast with experiments utilizing rich, complex media, regeneration with a defined medium resulted in increased rates of product formation far in excess of rates of concomitant cell growth. During one nutrient feeding phase, the increase in the butyric acid production rate was five times that of the cell growth‐rate increase, while the analogous ratio for butanol was 18. A hypothesis explaining the patterns of substrate consumption and product formation during the regeneration and deactivation periods has been developed: decreasing levels of exogenous ammonia induce the high‐affinity ammonia uptake system, causing the adenylate energy charge of the cells to increase; this in turn leads to the repression of glucose carbon flow through the EMP pathway and to the use of the acid‐forming catabolic reactions for more efficient production of ATP. Pulsewise addition of three different vitamins during one regeneration phase yielded interesting results, including a 50 % increase in butyric acid production rate that appeared to be connected with the addition of biotin. Finally, enumeration of the different morphological types of immobilized cells provided the first measurements of population dynamics in an immobilized cell system. Changes in the numbers of each cell type are consistent with the sporulation and germination processes of C. acetobutylicum observed with suspended cells.
Rheological Properties of Plant Cell Suspensions
Authors: Ballica Rabia , Ryu Dewey D. Y. , Powell Robert L. , Owen Derek
Published: March 2005

Packed cell and concentrated cell suspension cultures of Datura stramonium were exposed to oscillatory and steady shear on a Weissenberg rheogoniometer with a modified parallel plate geometry and to steady shear using other rotational viscometers. The viscoelastic properties of the cell suspensions were measured over a frequency range from 0.01 to 4 rad/s at low and high amplitudes. The applied strains were in the range of 1%‐36%. Increasing the strain caused cell damage (e.g., approximately 20% at a strain of 36% during 3 h of shearing). Cell cultures exhibited both viscoelastic and shear thinning behavior. The concentrated cell suspension cultures showed much lower dynamic viscosities and elastic moduli than the packed cell cultures. From the measured viscoelastic properties, yield stresses were calculated. The maximum calculated average yield stress was 4 × 104 dyn/cm2 for the packed cells and 325 dyn/cm2 for the concentrated cell suspension having a cell volume fraction (PCV, packed cell volume) of 0.9 mL/mL. From the steady shear experiments, yield stresses were also determined. The maximum calculated yield stress was 360 dyn/cm2 for a concentrated cell suspension (PCV = 0.9 mL/mL) and 95 dyn/cm2 for the less concentrated cell suspension (PCV = 0.6 mL/mL). It was confirmed that D. stramonium cell suspension cultures at high cell concentrations exhibit yield behavior which can be determined by either oscillatory or steady shear. The yield stress values determined in this study will be of some value to the bioprocess engineering of plant cells for high density cell cultivation and product recovery. In particular, these values will be important to determine bioreactor operating conditions such as agitation and aeration rates.
Mechanical Properties of Hydrocolloid Gels Filled with Internally Produced CO2 Gas Bubbles
Authors: Nussinovitch A. , Velez‐Silvestre R. , Peleg M.
Published: March 2005

Agar (2%), alginate (1% algin), and κ‐carrageenan (1.5%) gel specimens were prepared from mother solutions that contained 0–2.5 % sodium bicarbonate (agar and carrageenan) or calcium carbonate (alginate). Upon immersion in a citric acid bath (0–2%), the carbonate reacted with the diffusing acid to produce numerous carbon dioxide bubbles. The compressive strength and deformability of the gas‐filled gels so produced were determined using a Universal testing machine and compared with those of pure gels and gels containing the carbonate but not subjected to the process after various immersion times. While the agar and alginate gels retained considerable mechanical integrity even after several hours, the carrageenan gels disintegrated after about 2–5 h. Under similar conditions, the number of bubbles produced in the agar gels was about twice that in the alginate gels, an observation that cannot be explained solely by stoichiometric considerations.
Recombinant β‐Galactosidase Production in Serum‐Free Medium by Insect Cells in a 14‐L Airlift Bioreactor
Authors: King G. A. , Daugulis A. J. , Faulkner P. , Goosen M. F. A.
Published: March 2005

Spodoptera frugiperda (Sf9) insect cells were successfully cultured in serum‐free medium in a 14‐L airlift bioreactor. Cell densities as high as 1 × 107 cells/mL were achieved with specific growth rates of approximately 0.0286 h−1 (doubling time of 24 h). This system was also used to demonstrate the expression of a reported gene, β‐galactosidase (β‐gal), when cells were infected with a recombinant baculovirus. Approximately 0.33 mg of β‐gal/mL (i.e., 104 000 units/mL) of medium were obtained at the 14‐L scale, while about 0.95 mg of β‐gal/mL (i.e., 285 000 units/mL) of medium were obtained in small‐scale shaker flasks. The difference was attributed to a suboptimal infection in the large scale. Specific oxygen consumption rates decreased from 5.58 × 10−17 mol O2cell·s in early exponential growth to 3.13 × 10−17 mol (O2cell·s at 3 days post‐infection.
STM and AFM in Biotechnology
Authors: Haggerty Lorie , Lenhoff Abraham M.
Published: March 2005

Scanning tunneling microscopy (STM) and atomic force microscopy (AFM) are capable of providing atomic‐scale images, as well as insights into functional characteristics, of surfaces. Biological materials can be studied by depositing them on appropriate substrates, and samples have been imaged with varying degrees of resolution. In this review, the principles of STM and AFM are summarized, and practical aspects of implementing them for studies relevant to biotechnology are discussed, with the emphasis on investigations of biological macromolecules such as DNA and proteins. Items covered include samples, surfaces, medium, tips, and typical operating conditions. Finally, prospects are discussed for potential future applications to biotechnology.
The Monod Constant for Growth for Candida utilis on Ammonium in Continuous and Batch Cultures
Authors: Cooney Michael J. , Mcdonald Karen A.
Published: March 2005

Monod constants for growth on ammonium were determined for Candida utilis under different growth conditions. Results indicate that values for Ks differed in batch and in continuous cultures. The addition of the vitamin thiamine, however, had no effect on Ks. By contrast, the maximum specific growth rate did not vary between batch and continuous cultures but was affected by the addition of the vitamin thiamine.