Incubation Flasks Research Articles

A novel flow-based procedure for automation of respirometric assays in soils

A flow-based strategy involving a gas-diffusion sampling probe was proposed for evaluating the respiration rate in soils. The amount of CO2 collected after a pre-defined time interval was proportional to the free CO2 released by the soil ecosystem. The 500-mL incubation flasks typically used for soil respirometric assays were adapted and a special cover was designed for connecting a tubular gas diffusion membrane, a fan, and a septum for adding the CO2(g) standards required for calibration. The method relied on the pH-dependent absorbance variations resulting from the CO2 collection. A 1.3mmolL−1 bromothymol blue solution (pH 7.0) acted as both acceptor and carrier streams. In order to widen the dynamical working range to 0.003–0.2mmol CO2, two analytical curves were obtained, each related to a different time interval for the CO2 collection. Kinetic curves related to CO2 release by the soil samples were straightforwardly attained. Repeatability and detection limit were estimated as 2.0% and 0.001mmol CO2 (n=10), and accuracy was assessed in relation to a recommended titrimetric procedure.

Diuron tolerance and potential degradation by pelagic microbiomes in the Great Barrier Reef lagoon.

Diuron is a herbicide commonly used in agricultural areas where excess application causes it to leach into rivers, reach sensitive marine environments like the Great Barrier Reef (GBR) lagoon and pose risks to marine life. To investigate the impact of diuron on whole prokaryotic communities that underpin the marine food web and are integral to coral reef health, GBR lagoon water was incubated with diuron at environmentally-relevant concentration (8 µg/L), and sequenced at specific time points over the following year. 16S rRNA gene amplicon profiling revealed no significant short- or long-term effect of diuron on microbiome structure. The relative abundance of prokaryotic phototrophs was not significantly altered by diuron, which suggests that they were largely tolerant at this concentration. Assembly of a metagenome derived from waters sampled at a similar location in the GBR lagoon did not reveal the presence of mutations in the cyanobacterial photosystem that could explain diuron tolerance. However, resident phages displayed several variants of this gene and could potentially play a role in tolerance acquisition. Slow biodegradation of diuron was reported in the incubation flasks, but no correlation with the relative abundance of heterotrophs was evident. Analysis of metagenomic reads supports the hypothesis that previously uncharacterized hydrolases carried by low-abundance species may mediate herbicide degradation in the GBR lagoon. Overall, this study offers evidence that pelagic phototrophs of the GBR lagoon may be more tolerant of diuron than other tropical organisms, and that heterotrophs in the microbial seed bank may have the potential to degrade diuron and alleviate local anthropogenic stresses to inshore GBR ecosystems.

Microalgae on display: a microfluidic pixel-based irradiance assay for photosynthetic growth.

Microalgal biofuel is an emerging sustainable energy resource. Photosynthetic growth is heavily dependent on irradiance, therefore photobioreactor design optimization requires comprehensive screening of irradiance variables, such as intensity, time variance and spectral composition. Here we present a microfluidic irradiance assay which leverages liquid crystal display technology to provide multiplexed screening of irradiance conditions on growth. An array of 238 microreactors are operated in parallel with identical chemical environments. The approach is demonstrated by performing three irradiance assays. The first assay evaluates the effect of intensity on growth, quantifying saturating intensity. The second assay quantifies the influence of time-varied intensity and the threshold frequency for growth. Lastly, the coupled influence of red-blue spectral composition and intensity is assessed. Each multiplexed assay is completed within three days. In contrast, completing the same number of experiments using conventional incubation flasks would require several years. Not only does our approach enable more rapid screening, but the short optical path avoids self-shading issues inherent to flask based systems.

Relationship between soil properties and the bias of N2O reduction by acetylene inhibition technique for analyzing soil denitrification potential

The acetylene inhibition technique (AIT) has been widely used to measure soil denitrification potential (SDP), but has been criticized also for underestimating the actual SDP due to limitations of the AIT. Possible effects of soil properties on the bias of the AIT-derived SDP have not been thoroughly investigated yet. The study presented here therefore aimed at quantifying the relationships between soil texture and nutrient contents and the bias of AIT-derived SDP. A total of 26 soils with a wide range of clay and nutrient contents were incubated according to the standard procedure of AIT for assaying SDP. Incubation flasks were made anaerobic by gas substitution with pure helium (99.999%). Changes in the N2 and N2O concentrations in the He headspace were measured using gas chromatography. The emission rates of N2O and N2 were calculated to quantify the bias of the AIT-derived SDP (expressed as the percentage of the actual SDP that was not accounted for by AIT). The results showed that the bias ranged from 8 to 98%. The bias was negatively correlated (P < 0.05) with the clay, silt, organic matter and nutrient contents of the soils. These results indicate that the bias of the AIT-derived SDP was higher in low-fertility soils than in fertile soils. We recommend that soil properties are taken into account when interpreting results of AIT-derived SDP values.

Effect of propionic and lactic acids on in vitro ruminal bacteria growth

The objective of this work was to evaluate the effect of the levels of lactic and propionic acids on in vitro fermentation of ruminal microorganisms. In experiment 1, the levels, in a total of 12 were the following: addition of 0 (control 1), 50, 100, 150, 200 and 250 mM of lactic acid and 0 (control 2), 50, 100, 150, 200 and 250 mM of propionic acid, respectively, in incubation flasks, which contained ruminal inoculum, glucose and synthetic culture medium, with two repetitions for each combination. In experiment 2, the combinations, in a total of 4, were the following: presence of 12 and 24 mM of propionic acid and 0 mg of glucose, respectively; presence of 12 and 24 mM of propionic acid and 40 mg of glucose, respectively, to the incubation flasks which contained ruminal inoculum, with or without glucose and in synthetic culture medium with two repetitions each. There was no effect on the specific growth velocity of ruminal microorganisms in the presence of lactic acid or propionic acid. However, when there were greater concentrations of these acids in the media, there was a longer lag phase in the microorganism phase. Acid propionic at the concentration of 24 mM inhibited the production of acid acetic and butyric acid in a media with glucose. Despite of not being used as a source of energy by the ruminal microorganisms, propionic acid affects their metabolism. Lactic and propionic acids inhibit growth of some ruminal microorganisms at elevated concentrations.

Whole Body Deletion of AMP-activated Protein Kinase β2 Reduces Muscle AMPK Activity and Exercise Capacity

AMP-activated protein kinase (AMPK) β subunits (β1 and β2) provide scaffolds for binding α and γ subunits and contain a carbohydrate-binding module important for regulating enzyme activity. We generated C57Bl/6 mice with germline deletion of AMPK β2 (β2 KO) and examined AMPK expression and activity, exercise capacity, metabolic control during muscle contractions, aminoimidazole carboxamide ribonucleotide (AICAR) sensitivity, and susceptibility to obesity-induced insulin resistance. We find that β2 KO mice are viable and breed normally. β2 KO mice had a reduction in skeletal muscle AMPK α1 and α2 expression despite up-regulation of the β1 isoform. Heart AMPK α2 expression was also reduced but this did not affect resting AMPK α1 or α2 activities. AMPK α1 and α2 activities were not changed in liver, fat, or hypothalamus. AICAR-stimulated glucose uptake but not fatty acid oxidation was impaired in β2 KO mice. During treadmill running β2 KO mice had reduced maximal and endurance exercise capacity, which was associated with lower muscle and heart AMPK activity and reduced levels of muscle and liver glycogen. Reductions in exercise capacity of β2 KO mice were not due to lower muscle mitochondrial content or defects in contraction-stimulated glucose uptake or fatty acid oxidation. When challenged with a high-fat diet β2 KO mice gained more weight and were more susceptible to the development of hyperinsulinemia and glucose intolerance. In summary these data show that deletion of AMPK β2 reduces AMPK activity in skeletal muscle resulting in impaired exercise capacity and the worsening of diet-induced obesity and glucose intolerance.

Effect of In Vitro Docosahexaenoic Acid Supplementation to Marine Algae-Adapted and Unadapted Rumen Inoculum on the Biohydrogenation of Unsaturated Fatty Acids in Freeze-Dried Grass

The objective of this study was to examine the ruminal biohydrogenation of linoleic (18:2n-6) and linolenic (18:3n-3) acid during in vitro incubations with rumen inoculum from dairy cattle adapted or not to marine algae and with or without additional in vitro docosahexaenoic acid (DHA, 22:6n-3) supplementation. Treatments were incubated in 100-mL flasks containing 400mg of freeze-dried grass, 5mL of strained ruminal fluid, and 20mL of phosphate buffer. Ruminal fluid was collected just before the morning feeding from 3 cows receiving a control diet (49% ryegrass silage, 39% corn silage, 1% straw, and 11% concentrate, fresh-weight basis) supplemented with marine algae for 21 d (adapted rumen fluid, aRF) or from the same cows receiving the control diet only for 14 d after marine algae supplementation was stopped (unadapted rumen fluid, uRF). In half of the incubation flasks, pure DHA (5mg) was added as an oil-ethanol solution (100mL). Incubations were carried out during 0, 0.5, 1, 2, 4, 6, and 24h. After 24h, in vitro addition of DHA resulted in greater amounts (mg/incubation) of 18:3n-3 (0.23, 0.43, 0.26, and 0.34 for aRF, aRF+DHA, uRF, and uRF+DHA), 18:2n-6 (0.14, 0.22, 0.15, and 0.20 for aRF, aRF+DHA, uRF, and uRF+DHA) and trans-11, cis-15-18:2 (0.27, 2.40, 0.06, and 2.21 for aRF, aRF+DHA, uRF, and uRF+DHA), whereas no effect of inoculum source was observed. Trans-11-18:1 accumulated after 24h when aRF was incubated irrespective of in vitro DHA supplementation, whereas in incubations with uRF, accumulation of trans-11-18:1 only occurred when DHA was added (6.40, 4.35, 1.06, and 3.91 for aRF, aRF+DHA, uRF, and uRF+DHA). The increased amounts of trans-11-18:1 were due to the strong inhibition of the reduction to 18:0 because no 18:0 was formed when trans-11-18:1 accumulated after 24h. The results of the current experiment shows hydrogenation of trans-11, cis-15-18:2 occurred in the absence of in vitro DHA only, whereas substantial hydrogenation of trans-11-18:1 to 18:0 only took place in incubations without DHA and with unadapted rumen inoculum, confirming the higher sensitivity of the latter process to DHA.

Mixing layer running incubator (MIRI): an instrument for incubating samples while moving vertically in the mixing layer

Research on the impact of natural radiation on aquatic biota requires novel instruments and methods enabling realistic tests of the effects of changing radiation on communities. This article describes an instrument devised to expose samples maintained in incubation flasks to the variable conditions experienced by the water masses in motion in the mixing layer. This apparatus can be useful to measure the biological processes occurring in the photic zone of aquatic ecosystems avoiding the bias due to the incubation at fixed depths. Here an example of its use is presented, comparing the effect of photosynthetically active radiation (PAR) and ultraviolet radiation (UV-R) in samples incubated at fixed depths and using MIRI (MIxing layer Running Incubator).

Selective tachykinin NK 3-receptor agonists stimulate in vitro exocrine pancreatic secretion in the guinea pig

The tachykinins, including substance P, neurokinin A and neurokinin B, are a mammalian peptide family that have documented motor, sensory and circulatory neurotransmitter functions in the gut. Little is known about their action on the exocrine pancreas. In this study we investigated the effects of PG-KII, a natural NK 3-tachykinin receptor agonist, and senktide, a synthetic NK 3-tachykinin receptor agonist, on amylase release from isolated pancreatic lobules of the guinea pig in comparison with the secretagogues carbachol, caerulein and substance P and the depolarizing agent KCl. When added to incubation flasks at various concentrations (from 10 −10 to 10 −6 M), PG-KII and senktide both caused a dose-dependent increase in amylase release from pancreatic lobules. PG-KII and senktide elicited a lower maximal response (7.5±0.8 and 8.1±0.6% of the total lobular amylase content) than carbachol (34.4±3.9%), caerulein (26.5±2.8%) and KCl (22.5±3.8%). Whereas atropine left PG-KII and senktide-stimulated secretion unaffected, the non peptide NK 3 receptor antagonist SR 142801 significantly reduced the stimulant effect of PG-KII and senktide. PG-KII (10 −7 M) also slightly though significantly increased the response to lower concentrations of caerulein (10 −11 and 10 −10 M) and carbachol (10 −7 and 10 −6 M). These findings show that PG-KII and senktide are weak stimulants of exocrine pancreatic secretion that act directly on the acinar cells through NK 3 receptors, without cholinergic involvement. We suggest also that the tachykininergic NK 3 receptor system cooperates with the other known secretagogues in the control of pancreatic exocrine secretion.

Aerobic Metabolism of Flupropacil in Sandy Loam Soil

The aerobic soil metabolism of [ 14 C]flupropacil (isopropyl 2-chloro-5-(1,2,3,6-tetrahydro-3-methyl-2,6-dioxo-4-trifluoromethylpyrimidin-1-yl)benzoate) was determined in microbially active, sieved (2-mm) sandy loam soil with a soil moisture content of 75% at 1/3 bar. The soil was treated with [ 14 C]flupropacil at 0.5 mg kg -1 (twice the field use rate) and placed in incubation flasks connected to a series of traps (50 g litre -1 NaOH, 0.5 M H 2 SO 4 , ethylene glycol) and incubated at 25(±1)°C. Soil was sampled at 0, 3, 9, 20, 30, 48, 76, 120, 181 and 238 days of aerobic incubation. Volatiles were collected once every two weeks and on the day of soil sampling. Flupropacil metabolized with a half-life of 79 days under aerobic conditions. The major metabolite was flupropacil acid which accounted for up to 69.1% of the initially applied radioactivity at Day 238. Each of the two minor metabolites detected at the end of the study accounted for less than 0.5%. One of the minor metabolites was identified as C4242 acid (2-chloro-5-(1,2,3,6-tetrahydro-2,6-dioxo-4-trifluoromethylpyrimidin-1-yl)benzoic acid). Only a negligible portion (less than 0.3%) of the applied flupropacil was mineralized to [ 14 C]carbon dioxide. Extractable radioactivity ranged from 78.9% to 95.5%, with bound residues accounting for 3.2%-23.4%. The material balance ranged from 91.6% to 104.4%.

Formation of Organic Cadmium by Marine Microorganisms

This research was designed with a view to finding out whether or not there is a microbial or spontaneous transformation of inorganic cadmium into a highly toxic organic derivative of this metal by microorganism obtained from the Eastern Mediterranean marine environment. Sterile and nonsterile marine bottom sediment samples were incubated at ambient temperature for different time intervals, under aerobic and anaerobic conditions, with different concentrations of CdCl 2, and the medium, the atmosphere in the incubation flasks, and the sediments were assayed for their organic cadmium contents. The results were: (a) There was microbial growth in all systems containing up to 250 μg cadmium per milliliter of growth medium; however, with higher concentrations a complete growth-inhibition was observed. (b) Organic cadmium was found in the bottom sediments, at all cadmium levels permitting considerable microbial growth, but not in the systems′ atmospheres or in the liquid growth media. (c) Under aerobic conditions higher levels of organic cadmium were found than in the anaerobic systems. (d) The microorganisms occurring in the different experimental systems were isolated and identified. The predominating species in all systems were bacteria.

Remineralization of organic matter and degradation of the organic fraction of suspended solids in the River Danube

AME Aquatic Microbial Ecology Contact the journal Facebook Twitter RSS Mailing List Subscribe to our mailing list via Mailchimp HomeLatest VolumeAbout the JournalEditorsSpecials AME 09:279-288 (1995) - DOI: https://doi.org/10.3354/ame009279 Remineralization of organic matter and degradation of the organic fraction of suspended solids in the River Danube Hoch B, Berger B, Kavka G, Herndl GJ To determine the role of microbes in the degradation of organic matter in the River Danube, the biochemical oxygen demand (BOD) was followed over 2 and 5 d from July to December 1993, and the fluctuations of the microbial community were monitored in incubation flasks over the incubation periods of 2 and 5 d from January to December 1993. In the BOD flasks, bacterial abundance increased by up to 63% of the initial abundance during the first 15 h of incubation followed by an increase in flagellates, reaching their maximum about 35 h after starting the incubation. After 60 h, bacterial abundance declined to 15% of the initial density. This rapid fluctuation indicates that the BOD method routinely used to monitor water quality may be inappropriate for measuring the degradability of organic matter since the confined bacterial community is efficiently controlled by flagellates. The role and degradability of the organic coating of suspended solids was evaluated by Alcian blue staining and transmission light microscopy. Degradation experiments revealed that the number of particles decreased by up to 50% during the incubation period indicating rapid microbial utilization of this particulate matter. River Danube . Large river . Particle . Bacteria . Flagellates . BOD . Seston . Degradation Full text in pdf format PreviousNextExport citation RSS - Facebook - Tweet - linkedIn Cited by Published in AME Vol. 09, No. 3. Publication date: December 21, 1995 Print ISSN: 0948-3055; Online ISSN: 1616-1564 Copyright © 1995 Inter-Research.

A perifusion loop-gap resonator NMR probe for aerobic cell suspensions.

NMR studies of aerobic cell metabolism require that spectral data of high sensitivity and resolution be acquired from samples maintained under suitable conditions. The loop-gap resonator 31P NMR probe described here was designed for investigations of highly aerobic-dependent renal proximal tubules in an environment similar to those of conventional incubation flasks. The inherently higher sensitivity of the loop-gap resonator design and associated circuitry made it possible to obtain adequate spectra in short periods of time with small amounts of sample. Cellular physiological properties of perifused aerobic renal proximal tubule cell suspensions in the loop-gap resonator probe were found to be similar to equivalent tissue samples incubated in conventional flasks. Furthermore, the loop-gap perifusion probe was found to provide useful 31P NMR spectra of the kidney tubule system after acquisition times as short as 3 min.

Evaluation of Batch Culture Approaches That Can Be Used to Screen Release Rates of Ruminal Buffers

Our objectives were to evaluate in vitro characteristics of ruminal fluid from several diets and from several substrates that can be used to evaluate release rates of ruminal buffers. Ruminal fluid was collected from three cows fed diets of concentrate and sorghum silage in three ratios: 70:30, 60:40, and 50:50 (DM basis). Ruminal fluid was incubated in a shaking water bath with either purified corn starch, the same grain mix that was fed to the cow that provided the ruminal fluid for incubation, or the TMR fed to the cow that provided the ruminal fluid. In addition to this substrate, incubation flasks received either .5g of a 2:1 mixture of NaHCO3 and MgO or no buffer. A flask representing each substrate and buffer combination was removed every hour for 5h and analyzed for pH, buffering capacity, buffer value index, and VFA content. Each of the substrates yielded temporal alterations in ruminal fluid acid-base status similar to those observed previously in vivo. However, because ruminal fluid acidity tended to develop more rapidly with TMR as a substrate, we recommend that TMR be used as the substrate in batch culture to evaluate release rates of ruminal buffers. Addition of the NaHCO3 and MgO yielded an increase in ruminal fluid pH similar to that observed previously in vivo. Ruminal fluid acidity was highest for fluid from cows consuming the 70:30 diet; however, the high gas content of this fluid prevented accurate volume measurement for the incubations. Based on handling characteristics and temporal acid generation, we suggest that donor cows be fed a 60:40 grain to forage ratio to provide ruminal fluid for batch culture incubation; a combination of 75ml of this fluid, .5g of this TMR, and .5g of the test buffer provided a model that was acceptable for rapidly screening the release rates of ruminal buffers in vitro.

Ethylene Inhibition of Ammonium Oxidation in Soil

AbstractSmall quantities of ethylene (C2H4) are produced by bacteria and fungi in soils. Ethylene is produced when nitrogenase activity is measured by acetylene (C2H2) reduction. Acetylene has been used as a nitrification inhibitor and has been shown to produce C2H4 in flooded rice soils. Whether C2H4 affects the oxidation of NH4 by the soil's nitrifying organisms and at what concentrations was the focus of this study. A Weld clay loam surface soil (fine, montmorillonitic, mesic Aridic Paleustoll) was treated with (NH4)2SO4, subjected to various partial pressures of C2H4 and incubated at 30 °C. At designated incubation times, the atmospheres of the incubation flasks were analyzed for O2, CO2, and C2H4 by gas chromatography (GC) and the soil was analyzed for NH4, and NO3 plus NO2 by steam‐distillation procedures. At a C2H4 partial pressure of 0.017 kPa, NH4 oxidation was similar to untreated samples. At a C2H4 partial pressure of 0.17 kPa, NH4 oxidation was partially inhibited and, at a C2H4 partial pressure of 4.55 kPa, inhibition was nearly complete.

Methodological Modifications for Accurate and Efficient Determination of Contaminant Biodegradation in Unsaturated Calcareous Soils

Many techniques for quantifying microbial biodegradation of C-labeled compounds use soil-water slurries and trap mineralization-derived CO(2) in solution wells suspended within the incubation flasks. These methods are not satisfactory for studies of arid-region soils that are highly calcareous and unsaturated because (i) slurries do not simulate unsaturated conditions and (ii) the amount of CO(2) released from calcareous soils exceeds the capacity of the suspended well. This report describes simple, inexpensive methodological modifications for quantifying microbial degradation of [C]benzene and 1,2-dichloro[U-C]ethane in calcareous soils under unsaturated conditions. Soils at 50% water holding capacity were incubated with labeled contaminants for periods up to 10 weeks, followed by acidification of the soil and trapping of the evolved CO(2) in a separate container of 2 N NaOH. The CO(2) was transferred from the incubation flask to the trap solution by a gas transfer shunt containing activated charcoal to remove any volatilized labeled organics. The amount of CO(2) in the trap solution was measured by scintillation counting (disintegrations per minute). The method was tested by using two regional unamended surface soils, a sandy aridisol and a clay-rich riparian soil. The results demonstrated that both [C]benzene and 1,2-dichloro[U-C]ethane were mineralized to release substantial amounts of CO(2) within 10 weeks. Levels of mineralization varied with contaminant type, soil type, and aeration status (anaerobic vs. aerobic); no significant degradation was observed in abiotic control samples. Methodological refinements of this technique resulted in total CO(2) recovery efficiency of approximately 90%.

Leucopoiesis versus concentration of cytokines in diffusion chamber cultures of mouse bone marrow cells: clues to the physiological roles of growth factors.

Physiological mechanisms that regulate formation of neutrophil granulocytes, macrophages and their precursor cells were studied with the diffusion chamber (DC) technique. DC inoculated with mouse bone marrow cells were implanted intraperitoneally into host mice. When these in vivo cultures had been established and their marrow populations were expanding (2-day cultures), the DC were transferred to different environments: new, normal mice, lethally irradiated mice, or incubation flasks with optimal concentrations of growth factors. Culture development during the following final culture period was correlated to the concentration of some select candidate growth regulators in DC. After 3 d the cellularity of DC in irradiated hosts had increased significantly more than in the normal hosts. A difference was detectable already after 1-2 d when preculturing was omitted. The increased growth appeared to take place at several stages of cell maturation and not only at the progenitor cell level. Colony stimulating factor(s) for granulocyte and macrophage progenitors, as well as cytokines stimulating the bone marrow-derived cell line, 14 M.1, were present in DC fluid (DCF) at higher concentrations in irradiated than in normal mice throughout the final culture period. On the other hand, DCF concentrations of tumour necrosis-like factor (that may either induce CSF secretion or directly inhibit myelopoiesis), were not significantly different in irradiated compared with normal DC hosts. The cytokines detected in the DC may at least in part stem from inflammatory cells accumulating around the chambers. This animal model should be useful in further investigation of the highly complex regulatory network governing formation of white blood cells in the intact organism.

Increases in the basal secretion rate of follicle-stimulating hormone (FSH) accompany age-associated changes in serum FSH levels on estrus.

This laboratory has recently reported that by 5-6 months of age, alterations in the secretion and production of follicle-stimulating hormone (FSH) occur in virgin female rats which precedes the age-related disruption of estrous cycles and attenuation of preovulatory gonadotropin surges. Specifically, circulating immunoreactive FSH levels are higher on estrus in rats 5 months and older compared to levels measured in 2- to 3-month-old rats. Therefore, the present study was conducted to explore a possible mechanism for this age-related increase in FSH levels. At 1400 hr on proestrus, estrus and diestrus-1, groups (n = 6-12 rats/group) of 3- and 7-month-old, cyclic rats were decapitated, trunk blood was collected, and anterior pituitary glands were bisected and placed in incubation flasks containing 1 ml media (medium 199). Following a 30-min preincubation period, hemipituitary fragments were incubated for an additional 2 hr. Media and serum FSH levels were quantified by RIA. Levels of FSH were twofold higher in the serum of 7-month-old rats than 3-month-old rats on estrus. Similarly, the basal secretion rate (BSR) of FSH (expressed as ng FSH/ml/2 hr) was significantly (P less than 0.05) higher from incubated hemipituitary fragments of 7-month-old estrous rats than from fragments obtained from younger estrous rats (7 month: 1637 ng/ml/2 hr vs 3 months: 1253 ng/ml/2 hr). Neither the serum FSH levels nor the BSR of FSH differed between age groups on proestrus or diestrus-1. These results show that age-associated increases in circulating FSH levels on estrus may be attributed to an enhanced basal secretion of FSH from the pituitary gland.

Dinitrogen fixation in a montane forest sere determined by 15N 2 assimilation and in situ acetylene-reduction methods

Annual N input rates in a Rocky Mountain forest sere were determined by 15N 2 assimilation in an aerobic atmosphere using Swaby-type electrolytic O 2-producing incubation flasks, and by in situ enclosures employing C 2H 2 reduction methods. It was determined that the input in a subalpine meadow was 0.33, in aspen 0.52 and in spruce sites 0.32 kg N ha −1 yr −1, respectively, by combined non-symbiotic and associative-symbiotic fixation. The in situ C 2H 2 reduction values equated with the 15N 2 fixation values if a ratio of 7.5 C 2H 2 reduced to 1 N 2 fixed was used. Symbiotic fixation by a lupine in the aspen site added 0.63 kg N ha −1 yr −1. The associate-symbiotic fixation contributed 14% of the N input in the meadow, 30% in the aspen, and nil in the coniferous stands estimated by the C 2H 2 reduction method in situ. The results supported the conclusions that the N 2 fixation, similar to other microbiological characteristics of the aspen and conifer sites, agrees with the accepted definition of a successional forest sere.

Cyanobacterial carotenoids: their roles in maintaining optimal photosynthetic production among aquatic bloom forming genera.

Photoprotective and photosynthetic roles of carotenoid pigments (xanthophylls and β-carotene) were examined in the major bloom forming blue-green algal (cyanobacterial) genera, Anabaena, Aphanizomenon and Microcystis. Since these genera often reside as scums in surface waters, attention was given to the ability of carotenoids to counter potential photooxidation due to maximum near U.V. and visible radiation as well as O2 supersaturation, characterizing surface waters supporting blooms. In U.V.-transparent quartz incubation flasks it was shown that inhibition of carotenoid synthesis by diphenylamine led to rapid photooxidation among the above genera. When carotenoid synthesis was allowed to proceed, a high degree of resistance to photooxidation resulted. Prolonged exposure to near U.V. irradiation led to enhanced carotenoid synthesis relative to chlorophyll a, which extended viability. Carotenoid enhancement also increased chlorophyll a-specific photosynthetic O2 production. It is concluded that enhanced carotenoid synthesis observed during blooms serves at least two ecological functions, i) providing photoprotection and ii) increasing photosynthetic performance of surface cyanobacterial populations.