Selected ion monitoring/isotope dilution mass spectrometric determination of 1-aminocyclopropane-1-carboxylic acid levels in ripening tomato fruit

A new method is described for the quantitation of 1-aminocyclopropane-1-carboxylic acid (ACC), the immediate precursor of ethylene in plants. [2,2,3,3,-2H4]ACC has been synthesized and used as an internal standard for selected ion monitoring/isotope dilution quantitation of this compound in ripening tomato fruit. These data are compared with those derived from the widely used indirect oxidative ACC assay (which underestimated the ACC levels by between two- and fourfold). The greater accuracy, sensitivity (100X), and specificity of the mass spectrometric method will be of considerable benefit to those interested in factors which control ACC and ultimately ethylene levels since it is believed that ACC synthesis and its oxidative metabolism to ethylene are the key points at which ethylene biosynthesis is regulated.     

Turnover rates of different collagen types measured by isotope ratio mass spectrometry.

The rates of collagen turnover in different tissues have been estimated in growing rats previously exposed to gaseous 18O2. The abundance of the stable isotope was measured using isotope ratio mass spectrometry following combustion of isolated collagen-derived hydroxyproline. Using this method, problems of label reutilization associated with radiolabelling methods are avoided. In general the results confirm the slow turnover rates with half-lives of total collagen in skin, muscle and gut of 74, 45 and 244 d, respectively. The use of cyanogen bromide digests of whole tissues followed by isolation of collagen type-specific peptides has allowed the comparison of turnover rates of collagen types I and III, indicating that collagen type III is turned over more rapidly than type I.     

A review of gastric processing in decapod crustaceans

This article reviews the mechanical processes associated with digestion in decapod crustaceans. The decapod crustacean gut is essentially an internal tube that is divided into three functional areas, the foregut, midgut, and hindgut. The foregut houses the gastric mill apparatus which functions in mastication (cutting and grinding) of the ingested food. The processed food passes into the pyloric region of the foregut which controls movement of digesta into the midgut region and hepatopancreas where intracellular digestion takes place. The movements of the foregut muscles and gastric mill are controlled via nerves from the stomatogastric ganglion. Contraction rates of the gastric mill and foregut muscles can be influenced by environmental factors such as salinity, temperature, and oxygen levels. Gut contraction rates depend on the magnitude of the environmental perturbation and the physiological ability of each species. The subsequent transit of the digesta from the foregut into the midgut and through the hindgut has been followed in a wide variety of crustaceans. Transit rates are commonly used as a measure of food processing rates and are keys in understanding strategies of adaptation to trophic conditions. Transit times vary from as little as 30 min in small copepods to over 150 h in larger lobsters. Transit times can be influenced by the size and the type of the meal, the size and activity level of an animal and changes in environmental temperature, salinity and oxygen tension. Ultimately, changes in transit times influence digestive efficiency (the amount of nutrients absorbed across the gut wall). Digestive efficiencies tend to be high for carnivorous crustaceans, but somewhat lower for those that consume plant material. A slowing of the transit rate allows more time for nutrient absorption but this may be confounded by changes in the environment, which may reduce the energy available for active transport processes. Given the large number of articles already published on the stomatogastric ganglion and its control mechanisms, this area will continue to be of interest to scientists. There is also a push towards studying animals in a more natural environment or even in the field and investigation of the energetic costs of the components of digestion under varying biotic and environmental conditions will undoubtedly be an area that expands in the future.     

Respiratory and digestive responses of postprandial Dungeness crabs, Cancer magister, and blue crabs, Callinectes sapidus, during hyposaline exposure

Respiratory responses and gastric processing were examined during hyposaline exposure in two crab species of differing osmoregulatory ability. The efficient osmoregulator, Callinectes sapidus, displayed an immediate increase in oxygen uptake when exposed to low salinity in isolation. In contrast, the weak osmoregulator, Cancer magister, showed no change in oxygen uptake upon acute exposure (<6 h), but slight increases in oxygen uptake tended to occur over longer time scales (12–24 h). These changes were likely attributable to an increase in avoidance activity after 6 h hyposaline exposure. Following feeding in 100% SW, oxygen uptake doubled for both species and remained elevated for 15 h. When postprandial crabs were exposed to low salinities, C. sapidus were able to sum the demands of osmoregulation and digestion. Thus, gastric processes continued unabated in low salinity. Conversely, postprandial C. magister prioritized responses to low salinity over those of digestion, resulting in a decrease in oxygen uptake when exposed to low salinity. This decrease in oxygen uptake corresponded to a reduction in the rate of contraction of the pyloric stomach and a subsequent doubling of gastric evacuation time. The current study is one of the few to illustrate how summation or prioritization of competing physiological systems is manifested in digestive processes.     

Quantifying factors influencing American lobster (Homarus americanus) predation on the invasive green crab (Carcinus maenas) in Newfoundland,Canada

The European green crab (Carcinus maenas) is a newly invasive species in Newfoundland, where it has likely been present for ≤15 years. The green crab has been found in stomach contents of American lobster (Homarus americanus) in New England and Nova Scotia, Canada, but predation on this species has not yet been quantified in Newfoundland. We conducted feeding experiments to determine whether lobsters from Newfoundland were as likely as those from Nova Scotia (which have coexisted with green crabs for >60 years) to recognize and prey upon this new species. We also performed experiments to determine whether green crabs reach a size refuge from predation and whether factors including starvation, availability of alternate food sources, or habitat complexity would influence the probability of lobster attacking or feeding on green crabs. In our trials, lobster origin had no significant effect on crab predation; lobsters, irrespective of origin, were more likely to consume small (<40 mm carapace width [CW]) and medium (40–65 mm CW) crabs than larger (>65 mm CW) ones. Nevertheless, even small lobsters (73–76 mm carapace length, 300 g) were able to kill and consume the largest green crabs (78 mm CW, 100 g). Green crabs were less likely to be attacked or eaten when an alternative food source was present, suggesting that the lobsters were preying on the crabs, rather than simply killing them in a dispute over territory. The addition of a shelter provided a refuge for the green crabs; however, the crabs were only able to avoid being injured or eaten if this shelter was structurally complex. The green crab is slowly spreading westward around the island of Newfoundland, and so its long‐term effects, interactions with other organisms, and contribution to the diet of Newfoundland lobsters remain to be seen.     

A year in the life of a Dungeness crab: methodology for determining microhabitat conditions experienced by large decapod crustaceans in estuaries

This study presents a methodology for combining archival data storage tags (DSTs) and ultrasonic transmitters to investigate the microhabitat conditions of adult Cancer magister (Dana), inhabiting an estuary. The temperature, salinity and depth experienced by free-ranging Dungeness crabs were recorded at 10-min intervals for periods ranging from 1 week to 8 months. Crabs were tracked using a hydrophone, and tags were recovered via concentrated trapping or returned by recreational fishers for a reward. These methods led to a return rate of 50%. Representative CTD tag data showed that the conditions recorded at fixed stations within the estuary were not reflective of those experienced by free-ranging crabs, but rather crabs were able to orientate and avoid low salinity within the estuary. The prevalence of low salinity exposure was linked to times of increased food availability within the estuary, suggesting that crabs were entering the shallows of the estuary to forage. The techniques used in this study demonstrate that DSTs are a viable means of determining the microhabitat conditions of crustaceans inhabiting highly variable environments.     

Submerged growth of the cultivated mushroom,Agaricus bisporus

Agaricus bisporus grew well in submerged culture in a medium containing malt extract, phosphate, and casein. Moderate growth occurred in defined media containing glucose, asparagine, phenylalanine, vitamins and minerals. Other amino acids did not stimulate growth. Growth was stimulated by vegetable oils, partly due to utilization of the oils, and partly to a more complex mechanism. Oleates had the same effect as vegetable oils; palmitates a lesser one. In shaken flasks maximum yield was reached after 22–24 days and in stirred and aerated fermentors after 8–10 days. Besides dry weight of mycelium, laccase activity was determined. The latter determination is suitable for a rapid estimation of the growth in routine experiments. The flavour of the mycelium was like that of mushrooms but weaker. It was strongest in standing liquid cultures and on solid media. The mycelium grown in submerged culture was suitable as spawn for mushroom culture. Presented at the First International Mycological Congress, Exeter, 7–16 September 1971, and at the Meeting of the Netherlands Society for Microbiology, Rotterdam, 8 December 1971. We thank the Mushroom Experiment Station, Horst, the Netherlands, for kindly supplying the compost for fructification experiments and Mr. P. Arntz, M.Sc., for advice in the fermentor work. F. IJ. Dijkstra is indebted to the Royal Netherlands Fermentation Industries (Gist-Brocades), Delft, for a research grant.     

Measurement of fractionated water loss and CO2 production using triply labelled water

The doubly labelled water (DLW) method for measuring CO2 production has recently been the subject of much interest since no other technique gives integrated values for CO2 production over long periods by free-living subjects. The importance of evaporative water loss and fractionation factors to the calculation of CO2 production using this technique is described. Present methods of estimating evaporative water loss and the use of fractionation factors are summarized together with a discussion of their limitations. A novel technique is proposed whereby water labelled with three isotopes can be used to measure evaporative water loss and CO2 production in completely free-living subjects, and the feasibility of the method is tested in simulations using experimental data. This technique has three advantages over existing methods of estimating evaporative water loss: (1) it can be used in completely free-living subjects without any additional experimental procedures (e.g. water-balance studies or physical trapping of water vapour); (2) it gives a direct estimate of fractionated evaporative water loss in each subject, since non-fractionated water lost as vapour is automatically compensated for; and (3) the routes of water loss do not have to be known. The appropriate calculations are presented together with a discussion of the difficulties of measuring oxygen-17 by mass spectrometry. It is estimated that the maximum theoretical error on calculated CO2 production is +/- 0.3%. Practical ways of achieving this theoretical level of accuracy are suggested. We conclude that the proposed technique will allow correction for evaporative water loss to be made more exactly, thereby increasing the accuracy of the heavy water technique for measuring CO2 production in free-living subjects.     

Cytokinin oxidase fromZea mays kernels andVinca rosea crown-gall tissue

Cytokinin oxidase has been partially purified from matureZea mays kernels and fromVinca rosea corwn-gall tissue. The molecular weights of the two enzymes, determined by gel filtration, are very different: 94,400 (±10%) forZ. mays and 25,100 (±10%) forV. rosea. Specificity studies have been performed using a large number of synthetic and naturally occurring cytokinins. Only a small number of these compounds serve as substrates and both enzymes exhibit similar substrate specificity. In agreement with other workers, a Δ² double bond in the N⁶ side chain is essential for activity. The presence of glucosyl or ribosyl groups in the 7-or 9-position or an alanyl group in the 9-position of the purine moiety have little effect on their susceptibility to cytokinin oxidase, but O-glucosyl derivatives are resistant to oxidation. The relevance of these enzyme systems to studies on cytokinin metabolism and to the endogenous cytokinins is discussed.     

Biosynthesis of tropane ester alkaloids in Datura

Datura meteloides plants were fed via the roots with [1″,2′-¹⁴C]tigloyl hygroline and as a control, [2′-¹⁴C]hygrine. After a week the alkaloids were isolated and degraded. Despite hydrolysis of the putative precursor it was possible, by label ratio, to show that esterification occurs after, and not before, the tropane ring has been synthesized. Hygroline is proposed as a possible intermediate.