A preliminary study of a biosensor based on flow injection of the recognition element

A biosensor based on flow injection of the recognition element has been developed. As a model a pH-transducer was used, and urease was chosen as the recognition element. The pH-transducer was immersed in an internal flow-through chamber which was in contact with the sample solution via a semi-permeable membrane. The recognition element, urease, was injected into the buffer solution passing through the biosensor. The enzyme catalysed the hydrolysis of urea and the concomitant increase in pH was recorded. The biosensor response time was about three minutes at a constant flow rate of 0·05 ml/min. The linear range of the calibration curve of the biosensor was 0–5 mM. The observed detection limit was approximately 0.1 mM. The sample throughput was 6–12 per hour. The pH-response of the biosensor, for a sample solution containing urea (3·26 mM), showed a reproducibility (r.s.d) of 28% (n = 5) and a repeatability (r.s.d.) of 8% (n = 5). Operation at elevated temperatures (up to 50°C) was demonstrated. The presence of glucose (28 mM), acetone (6·7 mM), citric acid (0·2 mM) or sodium acetate (0·6 mM) in the sample solution did not interfere with the sensor response. A lowering of the biosensor response which was observed in the presence of copper ions (due to urease inhibition) could be completely eliminated by adding EDTA to the urease solution. Thus, this work demonstrates a new type of biosensors, based on SIRE-technology (Sensors with Injectable Recognition Elements), which show high accuracy and stability, quick response and high sample throughput. These features suggest the suitability of the system for automation. Such sensors should readily be combined with other enzymes or enzyme systems. The enzyme (urease) cost per analysis (injection) for the biosensor was estimated to be approximately US$0·02. This could be substantially reduced by further optimisation and miniaturisation.     

Fatty acid composition of Goniobasis virginica, Physa sp. and Viviparus malleatus (Mollusca: Gastropoda) from Lake Musconetcong, New Jersey

Gas—liquid chromatographic studies were done to determine the fatty acid composition of Goniobasis virginica Physa sp., and Viviparus malleatus (Mollusca: Gastropoda), from Lake Musconetcong, NJ, U.S.A. Palmitic acid was the predominant saturated fatty acid, followed by stearic acid, in all three molluscan species. The chief monoenes were 16:n−7, 18:1n−7; 20:1n−11+9, and 22:1n−11+13, which together accounted for all monoene fatty acids and one-quarter of the total fatty acids. Considerable amounts of linolenic acid (2.7−4.1%) and arachidonic acid (7.8−12%) were found in all three species. The percentage composition of docosahexaenoic acid (22:6n−3) was low compared to that of eicosapentaenoic acid (22:5n−3). Non-methylene interrupted dienes (20:2 NMID), characteristic of marine molluscs, ranged from 2 to 3% in the three species of freshwater snails.     

Emulsifying behaviour of gum arabic. Part 2: Effect of the gum molecular weight on the emulsion droplet-size distribution

The effect of the molecular weight of the gum arabic sample on droplet-size distributions of n-hexadecane-in-water emulsions (1% wt gum, 10% vol. oil) has been investigated at neutral pH. A high-molecular-weight fraction (0·87% nitrogen) corresponding to 10% of a natural gum (0·38% N) gives initially slightly larger droplets but better emulsion stability than the low-molecular-weight fraction (0·35% N) corresponding to the residual 90% of the original gum. Samples of a different gum arabic (0·35% N) subjected to different degrees of controlled degradation give decreasing emulsion stability with reduction in weight-average molecular weight from 3·1 × 10⁵ to 2·2 × 10⁵Da.     

Structure of potato starch pastes in the ageing process by the measurement of their dynamic moduli

The effect of ageing on the rheological properties of potato starch pastes was investigated by measurement of dynamic viscoelasticity. The value of the storage modulus (G′) increased rapidly with time for the first few hours, but attained equilibrium after a long period of ageing. The relation between the concentration and the storage modulus showed a c^1·6 dependence. The Avrami exponent (n) obtained in this study was 0·66 and deviated significantly from the value (n = 1), corresponding to rod-like growth of crystal. The rate of storage modulus increase decreased with increasing concentrations up to 10%, and at higher concentrations little or no increase was observed. In addition, the rate of storage modulus increase increased dramatically when the NaCl concentration was raised to levels of 0·01 or above. An even greater increase in rate occurs on addition of AlCl₃.     

Pretreatment of beet molasses to increase pullulan production

Pretreatment of beet molasses with cation exchange resin, sulphuric acid, tricalcium phosphate, potassium ferrocyanide, and ethylenediaminetetraacetic acid and disodium salt (EDTA) to increase the production of pullulan was investigated. Among the above techniques used for the removal of heavy metals, sulphuric acid treatment gave better results regarding polysaccharide concentration, polysaccharide yield, and sugar utilization. Aureobasidium pullulans grown on beet molasses produced a mixture of pullulan and other polysaccharides. The pullulan content of the crude polysaccharide was 30–35%. The addition of nutrients improved the production of polysaccharide. A maximum polysaccharide concentration (32·0±1·0 g litre⁻¹) was achieved in molasses solution (70 g litre ¹ initial sugar concentration, pH 6·5–7·5) treated with sulphuric acid and supplemented with K₂HPO₄ 0·5%, -glutamic acid 1%, olive oil 2·5% and Tween 80 0·5%. In this case, the highest values of biomass dry weight (33·8±1·0 g litre⁻¹), polysaccharide yield (63·5±2·5%), and sugar utilization (97·5±1·5%) were obtained at pH 6·5, 3·5, and 4·5–7·5, respectively.     

The crystal and molecular structures of lithium trimethylenediaminetetraacetatoferrate(III) trihydrate Li[Fe(trdta)]·3H2O and sodium ethylenediamine-N,N′-diacetato-N,N′-di-3-propionatoferrate(II) pentahydrate Na[Fe(eddda)]·5H2O

The crystal structures of Li[Fe(trtda)]·3H₂O and Na[Fe(eddda)]·5H₂O (trtda = trimethylenediaminetetraacetate and eddda = ethylenediamine-N,N′-diacetate-N,N′-di-3-propionate) have been determined by single crystal X-ray diffraction techniques. The former crystal was monoclinic with the space group P2₁/n,a = 17.775(3),b = 10.261(1),c = 8.883(2)Å, β = 95.86(4)° and Z = 4. The latter was also monoclinic with the space group P2₁/n,a = 6.894(2),b = 20.710(6),c = 13.966(3)Å, β = 101.44(2)° and Z = 4. Both complex anions were found to adopt an octahedral six-coordinated structure with all of six ligand atoms of trdta⁴⁻ or eddda⁴⁻ coordinated to the Fe(III) ion, unlike the corresponding edta⁴⁻ complex which is usually seven-coordinate with the seventh coordination site occupied by H₂O. Of the three geometrical isomers possible for the eddda complex, the trans(O₅) isomer was actually found in the latter crystal. Factors determining the structural types of metal–edta complexes are discussed in detail.     

Colocalization of NO and VIP in neurons of the submucous plexus in the rat intestine

Since very few previous studies have carried out the quantitative analysis for the colocalization of nitric oxide (NO) and vasoactive intestinal peptide (VIP) in the submucous neurons in the rat digestive tract, we applied in vivo treatment of colchicine to enhance the immunoreactivity and examined the colocalization of NO synthase (nNOS) and VIP in neurons of the submucous plexus throughout the rat digestive tract. The density of nNOS-containing neurons in the submucous plexus in the stomach corpus (103±25 cells/cm², n=3) and that in the antrum (157±9 cells/cm², n=3) were significantly lower than those in small and large intestine. However no difference was detected in the cell density among duodenum (1967±188 cells/cm², n=3), jejunum (2640±140 cells/cm², n=3), ileum (2070±42 cells/cm², n=3), proximal colon (2243±138 cells/cm², n=3) and distal colon (2633±376 cells/cm², n=3). The proportion of nNOS-immunoreactive (IR), nNOS/VIP-IR and VIP-IR neurons to the total number of submucous neurons was examined. nNOS/VIP-IR neurons comprised 45–55% of total number of submucous neurons from the duodenum to the proximal colon, however those comprised 66.4±5.1% in the distal colon. The results showed that the dense distribution of nNOS-containing neurons was found in the submucous plexus throughout the small and large intestine, and large population of submucous neurons co-stored nNOS and VIP.     

Effects of the type of meniscal lesion on knee function

The relationship between the type of meniscal lesion (bucket-handle, flap or degenerative tears) and preoperative knee function (5.1±6.2 days before arthroscopic meniscectomy) was studied in 35 patients. Patients with bucket-handle tears (group B, n=12) had large knee extension work deficits during maximal voluntary contractions at 30° and 180°/s (Kin-Com dynamometer). These work deficits were accompanied by proportional decreases in the prime mover or agonist-EMG levels (VM: vastus medialis and/or VL: vastus lateralis). These patients had smaller deficits during flexion movements at 30°/s. Moreover, a larger number of negative clinical signs and symptoms (pain at rest and during tests, locking, thigh atrophy, extension and flexion movement deficits larger than 10°) were found with an equal or a higher prevalence in group B than in the other groups. Patients with flap tears (group F, n=15) had deficits in work and agonist-EMG activity (VM and VL) only during the extension tests. In contrast, patients with degenerative tears (group D, n=8) had a work deficit and a concomitant decrease in the EMG level of the medial gastrocnemius (MG), only during the flexion test at 30°/s. Comparable mean knee function scores, as measured by the Lysholm and Gillquist questionnaire [35], were obtained for the three groups of patients, suggesting that this measure was not sensitive enough to discern functional differences related to types of meniscal tears. The results of this study have demonstrated a link between the type of meniscal lesion and the consequent preoperative knee joint disability profile as defined by comparison with the sound leg. These results emphasize the need to consider meniscal lesion type and an individual's preoperative strength deficit when group comparisons of patients are made or the effects of therapy (arthroscopic surgery and rehabilitation) are evaluated.     

A computational formula for heat influx in animal experiments

1. 1.The bahavioural paradigm in which cold-exposed animals can work for pulses of infrared radiation has been extensively used in the literature, but a formula to calculate the amount of heat obtained has not been advanced.

2. 2.This paper describes a computational formula for heat influx in rats: E = 3.64 · 10⁻⁶ · n · d · I · M^0.6 where E is heat influx (kJ), n is number of rewards, d is reward duration (sec), I is irradiance (mW/cm²), and M is body mass (g).

The relationship between 8-oxo-7,8-dihydro-2'-deoxyguanosine level and extent of cytosine methylation in leukocytes DNA of healthy subjects and in patients with colon adenomas and carcinomas

It has been known for a long time that DNA hypomethylation occurs in many human cancers and precancerous conditions. However, the mechanisms of hypomethylation are largely unknown. It is possible that endogenous 8-oxo-7,8-dihydroguanine (8-oxoGua) level may be linked to aberrant DNA methylation of adjacent cytosine and in this way influences carcinogenesis. Therefore, the aim of the present study was to assess a possible link between 8-oxo-7,8-dihydro-2′-deoxyguanosine (8-oxodG) background level and 5-methylcytosine content in DNA from human leukocytes of healthy subjects (n = 105) as well as in patients with colon adenomas (n = 39) and carcinomas (n = 50).

Our results demonstrated statistically significant negative correlation between background level of 8-oxodG and 5-methylcytosine content in DNA isolated from leukocytes of healthy donors (r = −0.3436, p = 0.0003). The mean content of 5-methylcytosine was significantly lower, while 8-oxodG level was significantly higher in leukocytes DNA of patients with colon adenomas and carcinomas in comparison with healthy subjects. The mean values for 5-methylcytosine were: 3.59 ± 0.173% (healthy subjects), 3.38 ± 0.128% (patients with adenomas), 3.40 ± 0.208% (colon cancer patients). The mean values of 8-oxodG in DNA were, respectively: 4.67 ± 1.276, 5.72 ± 1.787, 5.76 ± 1.884 8-oxodG per 10⁶ dG molecules. DNA from affected tissue (colon) suffered from significant, about 10% reduction in cytosine methylation in comparison with leukocytes of the paired subjects.

Our work provides the first in vivo evidence suggesting that increased levels of 8-oxodG in DNA may lead to carcinogenesis not only via mispair/mutagenic potential of the modified base but also through its ability to influence gene expression by affecting DNA methylation.     

Ovarian follicular wave synchronization and pregnancy rate after fixed-time natural mating in llamas

The study was designed to compare the efficacy of treatments intended to induce follicular wave synchronization among llamas (Experiment 1), and to determine the effect of these treatments on pregnancy rates after fixed-time natural mating (Experiment 2). In Experiment 1, llamas were treated with: (1) saline (control, n=20); (2) estradiol and progesterone (E/P, n=20); (3) LH (LH, n=20); or (4) transvaginal ultrasound-guided follicle ablation (FA, n=20). The ovarian response was monitored daily by transrectal ultrasonography. The intervals from treatment to follicular wave emergence and to the day on which the new dominant follicle reached ≥7 mm, respectively, did not differ between the LH (2.1±0.3 days and 5.2±0.5 days, respectively) and FA groups (2.3±0.3 days and 5.0±0.5 days), but both were shorter (P<0.05) and less variable (P<0.01) than in the control group (5.5±1.0 days and 8.4±2.0 days), while the E/P group (4.5±0.8 days and 7.7±0.5 days) was intermediate. In Experiment 2, llamas at unknown stages of follicular development were assigned randomly to control, E/P, and LH groups (n=30 per group). A single, fixed-time natural mating was permitted 10–12 days after treatment. Ovulation rates did not differ among groups (control, 93%; E/P, 90%; LH, 90%; P=0.99), but the pregnancy rate was higher (P<0.05) for synchronized llamas (LH and E/P groups combined, 41/54) than for non-synchronized llamas (control group, 15/28). In conclusion, LH and FA treatments were most effective for inducing follicular wave synchronization, while E/P treatment was intermediate. Synchronization treatments did not influence ovulation rate subsequent to fixed-time natural mating, but a higher pregnancy rate in synchronized than non-synchronized llamas warrants critical evaluation of the effects of follicular status on the developmental competence of the contained oocyte.     

Capsaicin-evoked bradycardia in anesthetized guinea pigs is mediated by endogenous tachykinins

The present study was done to characterize the effects of endogenous tachykinins on heart rate in urethane-anesthetized guinea pigs. Intravenous injection of capsaicin (32 nmol/kg) was used to evoke release of tachykinins and calcitonin gene-related peptide (CGRP) from cardiac sensory nerve fibers. Such injections caused a brief decrease in heart rate (− 37 ± 7 beats/min, n = 6) that was followed by a more prolonged increase (+ 44 ± 10 beats/min). Blood pressure was lowered by − 11 ± 2 mmHg. Bilateral vagotomy did not affect the chronotropic or depressor responses to capsaicin, but atropine (1 µmol/kg) nearly abolished the bradycardic response (− 8 ± 3 beats/min, n = 7). Combined blockade of NK₂ and NK₃ receptors, with SR48968 and SR14801 respectively, also caused a significant reduction of capsaicin-evoked bradycardia (− 14 ± 3 beats/min, n = 4) but did not affect bradycardia evoked by vagal nerve stimulation. Blockade of CGRP receptors eliminated capsaicin-evoked tachycardia and prolonged the capsaicin-evoked bradycardia. These findings suggest that capsaicin-evoked bradycardia in the anesthetized guinea pig is mediated by tachykinins that stimulate cardiac cholinergic neurons. This effect appears to be truncated by the positive chronotropic action of CGRP that is also released from cardiac afferents by capsaicin.     

Assessment of DNA damage and its modulation by dietary and genetic factors in smokers using the Comet assay: a biomarker model

Methods are needed to assess exposure to genotoxins in humans and to improve understanding of dietary cancer prevention. The Comet assay was used to detect smoking-related exposures and dietary modulations in target tissues. Buccal scrapings, blood and faeces were collected from 38 healthy male volunteers (smokers and non-smokers) during a dietary intervention study with bread supplemented with prebiotics±antioxidants. GSTM1-genotype was determined with PCR. Buccal and peripheral lymphocytes were analysed for DNA damage using the Comet assay. Genotoxicity of faecal water (FW) was assayed in human colon HT29 clone 19A cells. 'Tail intensity' (TI) was used as a quantitative indicator of DNA damage in the Comet assay. Intervention with bread reduced DNA damage in lymphocytes of smokers (8.3±1.7% TI versus 10.2±4.1% TI, n=19), but not of non-smokers (8.6±2.8% TI versus 8.3±2.7% TI, n=15). Faecal water genotoxicity was reduced only in non-smokers (9.4±2.9% TI versus 18.9±13.1% TI, n=15) but not in smokers (15.5±10.7% TI versus 20.4±14.1% TI, n=13). The Comet assay was efficient in the detection of both smoking-related exposure (buccal cells) and efficacy of dietary intervention (faecal samples). Smokers and non-smokers profited differently from the intervention with prebiotic bread±antioxidants. Stratification of data by genotype enhanced specificity/sensitivity of the intervention effects and contributed important information on the role of susceptibility.     

Different effects of ATP on the contractile activity of mice diaphragmatic and skeletal muscles

Apart from acetyl-choline (Ach), adenosine-5′-trisphosphate (ATP) is thought to play a role in neuromuscular function, however little information is available on its cellular physiology. As such, effects of ATP and adenosine on contractility of mice diaphragmatic and skeletal muscles (m. extensor digitorum longa—MEDL) have been investigated in in vitro experiments. Application of carbacholine (CCh) in vitro in different concentrations led to pronounced muscle contractions, varying from 9.15 ± 4.76 to 513.13 ± 15.4 mg and from 44.65 ± 5.01 to 101.46 ± 9.11 mg for diaphragm and MEDL, respectively. Two hundred micromolars of CCh in both muscles caused the contraction with the 65% (diaphragm) to 75% (MEDL) of maximal contraction force—this concentration was thus used in further experiments. It was found that application of ATP (100 μM) increased the force of diaphragmatic contraction caused by CCh (200 μM) from 335.2 ± 51.4 mg (n = 21) in controls to 426.5 ± 47.8 mg (n = 10; P < 0.05), but decreased the contractions of MEDL of CCh from 76.6 ± 6.5 mg (n = 26) in control to 40.2 ± 9.0 mg (n = 8; P < 0.05). Application of adenosine (100 μM) had no effect on CCh-induced contractions of these muscles.

Resting membrane potential (MP) measurements using sharp electrodes were done at 10, 20 and 30 min after the application of ATP and adenosine. Diaphragm showed depolarization from 75 ± 0.6 down to 63.2 ± 1.05, 57.2 ± 0.96 and 53.6 ± 1.1 mV after 10, 20 and 30 min of exposition, respectively (20 fibers from 4 muscles each, P < 0.05 in all three cases). Adenosine showed no effect on diaphragmatic MP. Both agents were ineffective in case of MEDL.

The effects of ATP in both tissues were abolished by suramin (100 μM), a P2-receptor antagonist, and chelerythrin (50 μM), a specific protein-kinase C (PKC) inhibitor, but were not affected by 1H-[1,2,4]-oxadiazolo-[4,3-]-quinoxalin-1-one (ODQ, 1 μM), a guanylyl-cyclase inhibitor, or by adenosine-3,5-monophosphothioate (Rp-cAMP, 1 μM), a protein-kinase A (PKA) inhibitor.

Besides the action on contractile activity, ATP (100 μM) led to a significant (P < 0.001) depolarization of diaphragm muscle fibers from 74.5 ± 2.3 down to 64 ± 2.1, 58.2 ± 2.2 and 54.3 ± 2.4 mV after 10, 20 and 30 min of incubation, respectively. Incubation of MEDL with the same ATP concentration showed no significant change of MP.

Denervation of the muscles for 28 days led to a decrease of CCh-induced contractions of diaphragm down to 171.1 ± 34.5 mg (n = 11, P < 0.05), but increased the contractile force of MEDL up to 723.9 ± 82.3 mg (n = 9, P < 0.01). Application of ATP elevated the contractility of denervated diaphragm caused by CCh up to normal values (311.1 ± 79.7 mg, n = 6, P > 0.05 versus control), but did not significantly affect of contractility of MEDL, which became 848.1 ± 62.7 mg (n = 6).

These results show that the effects of ATP on both diaphragmatic and skeletal muscles are mediated through P2Y receptors coupled to chelerytrin-sensitive protein-kinase C.     

Heifers sired by bulls with either high or low expected progeny differences (EPDs) for marbling do not differ in age at puberty

Selection for carcass traits such as deposition of intramuscular fat (marbling), which is a primary trait used as an indicator of meat quality, may incur additional effects such as younger age at puberty. Therefore, a trial was conducted for 2 years to determine age of puberty for two groups of crossbred heifers (n = 124) sired by purebred Angus bulls selected for either low (Low, n = 6) or high (High, n = 6) expected progeny differences (EPDs) for marbling. Our working hypothesis was that age at puberty would be reduced in heifers sired by bulls with high EPDs for marbling in comparison to those heifers sired by bulls with low EPDs for marbling. Puberty was determined by collecting weekly blood samples and assaying for concentrations of progesterone. Mean age at puberty for heifers during the study did not differ (392 ± 5 and 387 ± 5 days, Low and High respectively). Of the 124 heifers, 79% reached puberty within the 145 days allotted for the study. We conclude that use of sires that produce daughters with increased ability to marble does not result in decreased age at puberty.     

Influence of classification levels of ram sexual activity on spring breeding ewes

Ram lambs (7–8 months old) and mature rams (19–20 months old) were used to evaluate the effect of classification levels of male sexual performance on reproductive performance of ewes during spring breeding. In Exp. 1, sexually active ram lambs with high (1.8±0.3; n=5) and low (0.9±0.2; n=5) sexual performance scores (HP and LP; mean±S.E.M.) were used in single sire breeding pens. Ewes (n=305) were stratified by age and assigned to 10 pens for 34 days starting in late March. For Exps. 2 and 3, two replicates were conducted for 2 years with sexually active mature rams in a single sire mating scheme. For Exp. 2, HP rams (n=5) averaged 3.6±0.2 ejaculations and LP rams (n=7) 1.8±0.2 ejaculations for sexual performance scores based on nine, 30 min serving capacity tests (SCT). Polypay ewes (n=152 to 153 per year) were stratified by age and assigned to pens each year for 34–38 days starting in late March for years 1 and 2. For Exp. 3, HP rams (n=6) averaged 3.7±0.1 ejaculations and LP rams (n=10) 2.3±0.1 ejaculations for sexual performance scores based on 18, 30 min SCT. Polypay ewes (n=229 in year 3 and n=244 in year 4) were stratified by age and assigned to pens each year for 34 days starting in late March. In Exp. 1, lambing rates for ewes bred to HP versus LP ram lambs did not differ (65.8 versus 53.0; P=0.20). Prolificacy tended (P=0.06) to be increased by 0.1 lambs in ewes bred by LP ram lambs. Total number of lambs born per ewe present at lambing, and lambing distribution were not altered by HP and LP ram lambs. In Exp. 2, lambing rates for fall-lambing ewes bred to mature HP or LP rams did not differ (58.1 versus 60.1; P=0.78). In Exp. 3, lambing rates for fall-lambing ewes bred to mature HP or LP rams did not differ (74.3 versus 69.0; P=0.35). There was no difference (P>0.10) between years for Exp. 2 or Exp. 3, and mature HP and LP rams did not affect the other reproductive variables monitored. Analyses of the combined data for Exps. 2 and 3 indicated only a year difference (P<0.001) in lambing rates and total lambs born. Present studies indicate that different sexual performance classifications for ram lambs and mature rams did not alter lambing rates or distribution of lambing of Polypays bred in late March to April. These results indicate that HP and LP, sexually active, Polypay rams and ram lambs with average to high quality semen can provide a source of rams for spring breeding Polypays in ambient conditions and that there was no advantage to using HP over sexually active LP ram lambs or rams.     

Effect of organic or inorganic trace mineral supplementation on follicular response, ovulation, and embryo production in superovulated Angus heifers

We determined whether source of trace mineral supplementation prior to embryo collection affected embryo production and quality. Angus half-sibling heifers (n = 20) originating from a common herd were assigned to three treatment groups using a 3 × 3 latin square design replicated in time (3×) and space (6× complete and 1× incomplete): (1) heifers received no added mineral to their diet (control; n = 53); (2) heifers received a commercially available organic mineral supplement (organic; n = 52); or (3) heifers received an all inorganic mineral supplement (inorganic; n = 55). All heifers had ad libitum access to hay and were fed a supplement containing corn and soybean meal. Treatments were initiated 23 days prior to embryo recovery. Heifers were given a 45-day adaptation period of no mineral supplementation before initiating a new treatment. Ovarian structures were evaluated using transrectal ultrasonography to determine the presence and number of follicles and CL on each ovary. The mean number of recovered ova/embryos was similar among treatments (4.1 ± 0.7, 3.8 ± 0.7, and 3.3 ± 0.7 for control, inorganic, and organic treatments, respectively), the number of unfertilized oocytes was greater (P < 0.05) for inorganic (2.3 ± 0.5) and control (1.6 ± 0.5) treated heifers than organic (0.4 ± 0.4) treated heifers. No differences among treatments existed for the number of degenerate or transferable embryos, but individual heifer influenced the total number of embryos/ova, unfertilized ova, and transferable embryos recovered. We conclude that heifer accounted for the greatest differences in embryo production and quality. Source of trace mineral supplementation did not significantly alter embryo number or quality in superovulated purebred Angus heifers fed a well-balanced diet, meeting all trace mineral requirements.     

Chromosome Spreading Induced by Vegetable Oils

Seeds soaked in the oil extracted from castor beans (Ricinus communis) for 2 hr were germinated in petri dishes on moist filter papers. Root tips were fixed in acetic alcohol (1:3) at 10-14°C, for 24 hr, washed successively with 70% alcohol (15 min) and water (10 min), hydrolysed in 1 N HCl at 60°C for 15 min and stained in leucobasic fuchsin for 30 min. The stained tip was squashed under a cover glass in a drop of acetocarmine and sealed with paraffin wax. The slides were made permanent by separating the cover glass in a mixture of acetic acid and n-butyl alcohol (1:1), passing through 2 changes of n-butyl alcohol and mounting in balsam. Such a method leads to contraction and spreading of chromosomes, without affecting either the clarity of the constriction regions or the anaphase separation of chromosomes.     

Studies on isolation and characterization of starch from oat (Avena nuda) grains

Starch from AC Hill oat grains (Avena nuda) was isolated and some of the characteristics determined. The yield of starch was 23·4% on a whole grain basis. The shape of the granule was polyhedral to irregular, with granules 6–10 μm in diameter. Lipids were extracted by acid hydrolysis and by selective solvent extraction with chloroform-methanol 2:1 v/v (CM) at ambient temperature, followed by n-propanol-water 3:1 v/v (PW) at 90–100°C. The acid hydrolyzed extracts which represented the total starch lipids (TSL) was 1·13%. The free lipids in the CM extract (1% TSL) was 6·2%, whereas the free and bound lipids in the PW extracts was 93.0%. Neutral lipids formed the major lipid class in the CM and PW extracts. The monoacyl lipid content in both CM and PW extracts was 61·0%. The total amylose content was 19·4%, of which 13·9% was complexed by native lipids. X-ray diffraction was of the ‘A’ type. Oat starch differed from wheat starch in showing a higher swelling factor, decreased amylose leaching, coleaching of a branched starch component and amylose during the pasting process, higher peak viscosity and set-back, low gel rigidity, greater susceptibility towards acid hydrolysis, greater resistance to -amylase action and a higher freeze-thaw stability. Furthermore, in comparison to wheat starch, the amylose chains of oat starch appear to be more loosely arranged in the amorphous regions, whereas in crystalline regions, oat starch chains are more compactly packed. Lipid removal from oat and wheat starches decreased their swelling factor, peak viscosity, set-back, gelatinization temperatures, freeze-thaw stability and paste clarity (at pH > 4·0), and increased their thermal stability, amylose leaching, enthalpy of gelatinization, susceptibility towards -amylase and paste clarity (at pH < 4·0). The results also showed that the properties of AC Hill oat starch is not representative of oat starch in general.     

Xenobiotic acyl-CoA formation: evidence of kinetically distinct hepatic microsomal long-chain fatty acid and nafenopin-CoA ligases

Multiplicity of hepatic microsomal coenzyme A ligases catalyzing acyl-CoA thioester formation is an important factor for consideration in relation to the metabolism of xenobiotic carboxylic acids. In this study the kinetic characteristics of rat hepatic microsomal nafenopin-CoA ligase were studied and compared with those of long-chain fatty acid (palmitoyl) CoA ligase. The high affinity component of palmitoyl-CoA formation was inhibited by nafenopin (K_i 53 μM) and ciprofibrate (K_i 1000 μM). Analagous to palmitoyl-CoA, nafenopin-CoA formation was catalyzed by an apparent high affinity low capacity isoform (K_m 6 ± 2.5 μM, (V_max 0.33 ± 0.12 nmol/mg per min) which was inhibited competitively by palmitic acid (mean K_i 1.7 μM, n = 5) and R-ibuprofen (mean K_i 10.8 μM, n = 5) whilst ciprofibrate and clofibric acid were ineffective as inhibitors. The intrinsic metabolic clearance of nafenopin to nafenopin-CoA (V_max/K_m 0.057 ± 0.011 nmol/mg/min ± M) was similar to that reported recently for the formation of ibuprofenyl-CoA by rat liver microsomes. Evidence of both a substantial difference between the K_m and K_i for nafenopin and lack of commonality with regard to xenobiotic inhibitors suggests that the high affinity microsomal nafenopin-CoA and long-chain fatty acid-CoA ligases are kinetically distinct. Thus until the current ‘long-chain like’ xenobiotic-CoA ligases are fully characterised in terms of substrate specificity, inhibitor profile, etc, it will be impossible to rationalize (and possibly predict) the metabolism and hence toxicity of xenobiotic carboxylic acids forming acyl-CoA thioester intermediates.