Endogenous levels of oxygen, carbon dioxide and ethylene in stems of Norway spruce trees during one growing season

The carbon dioxide and ethylene concentrations in tomato fruit (Lycopersicon esculentum cv. Castelmart) and their stage of ripeness (characteristic external color changes) were periodically measured in fruit attached to and detached from the plant. An external collection apparatus was attached to the surface of individual tomato fruit to permit non‐destructive sampling of internal gases. The concentration of carbon dioxide and ethylene in the collection apparatus reached 95% of the concentration in the fruit after 8 h. Gas samples were collected every 24 h. A characteristic climacteric surge in carbon dioxide (2‐fold) and ethylene (10‐fold) concentration occurred coincident with ripening of detached tomato fruit. Fruit attached to the plant exhibited a climacteric rise in ethylene (20‐fold) concentration during ripening, but only a linear increase in carbon dioxide concentration. The carbon dioxide concentration increases in attached fruit during ripening, but the increase is a continuation of the linear increase seen in both attached and detached fruit before ripening and does not exhibit the characteristic pattern normally associated with ripening climacteric fruit. In tomato fruit, it appears that a respiratory climacteric per se, which has been considered intrinsic to the ripening of certain fruit, may not be necessary for the ripening of “climacteric” fruit at all, but instead may be an artifact of using harvested fruit.

Our initial study of sagebrush and grasshopper responses to elevated and historical carbon dioxide atmospheres is complete and has been accepted for publication. The study on Biomass Allocation Patterns of Defoliated Sagebrush Grown Under Two Levels of Carbon Dioxide has completed and the manuscript has been submitted for publication. We have completed the study of plant growth under two nutrient and carbon dioxide regimes and grasshopper feeding responses. The study of a specialist feeding caterpillar, the cabbage butterfly, and a mustard hostplant has recently been completed. We were able to identify the principal allelochemicals of the mustard plants, butenyl and pentenyl isothiocyanates, by combined gas chromatography and mass spectrometry. Measurement of these chemicals has been a critical component of this study since these compounds contain nitrogen and sulphur and act as a feeding stimulant to the caterpillar. This insect responds to elevated carbon dioxide by consuming more leaves and we can now say that this is not due to a change in the feeding stimulants. Reduced leaf protein content is a critical factor for even specialist feeding insect herbivores under elevated carbon dioxide conditions. The study on Grasshopper Population Responses to Enriched Carbon Dioxide Concentration is currently in progress at the Duke University Phytotron. We have changed hostplant species in order to complement the investigations of carbon dioxide effects on tallgrass prairie. Specifically, we are using big bluestem, Andropogon geradii, as the host plant to feed to the grasshoppers. This experiment will be completed in July 1990.

Mild intermittent hypoxia protocols are characterized by a few (e.g. 12 episodes) brief episodes (e.g. 2 min) of mild hypoxia (e.g. 85–87 % oxygen saturation) interspersed with short recovery periods (Mateika & Sandhu 2011; Puri et al., 2021). In animal models, exposure to this stimulus leads to the initiation of long-term facilitation (LTF), which is a term used to characterize sustained increases in motor neuron, nerve or muscle activity initiated in animals following exposure to mild intermittent hypoxia. The sustained increases in motoneuronal activity have been linked to downstream increases in ventilation, upper airway patency and limb muscle function in animal models (Mateika et al., 2015). Some of the neuronal pathways and cellular mechanisms responsible for the sustained increase in activity have been determined (Mitchell & Baker 2022). Many studies completed over the past two decades have attempted to initiate LTF of ventilation, upper airway muscle or limb muscle activity in humans (Mateika & Sandhu 2011; Puri et al., 2021). However, the initiation of this phenomenon, or the magnitude of the phenomenon when it manifests, has been variable when comparisons are made across studies (see supplements in Mateika & Sandhu 2011; Puri et al., 2021). Potential reasons for this variability include differences in the number and duration of episodes that formulate mild intermittent hypoxia protocols, along with differences in the intensity of hypoxia considered to fall within the mild range (Mateika & Sandhu 2011; Puri et al., 2021). Likewise, some protocols have employed a combination of intermittent hypoxia and hypercapnia (see section below on 'Potential Mechanisms that Link Carbon Dioxide to the Initiation and Manifestation of LTF' for further discussion of this point) rather than intermittent hypoxia alone. Moreover, different human models (i.e. healthy humans, humans living with obstructive sleep apnoea (OSA), humans living with spinal cord injury) have been used to explore this phenomenon (Mateika & Sandhu 2011; Puri et al., 2021). These experimental design variations could be responsible for the variability that has been reported. On the other hand, we propose that one of the primary keys to the initiation and subsequent manifestation of LTF in humans is the presence of sustained levels of carbon dioxide during and following exposure to mild intermittent hypoxia. We and others established two decades ago that ventilatory LTF (vLTF) (Harris et al., 2006; Jordan et al., 2002; Khodadadeh et al., 2006; Mateika et al., 2004) and LTF of upper airway muscle activity (Harris et al., 2006) does not manifest in healthy humans (Harris et al., 2006; Jordan et al., 2002; Mateika et al., 2004) or humans with sleep apnoea (Khodadadeh et al., 2006) following exposure to mild intermittent hypoxia, when carbon dioxide levels are uncontrolled and hypocapnia ensues. Subsequent to these findings, ongoing studies provided additional insight by showing that sustained increases in respiratory and limb muscle function in humans, often presented in the framework of spinal cord injury, are frequently not evident following exposure to mild intermittent hypoxia in the presence of uncontrolled carbon dioxide levels (Gandevia & Butler 2024). Given the inaugural findings, these latter results are not surprising because common neural pathways and cellular mechanisms have been proposed to be involved in the initiation of LTF of respiratory and limb motor neurons based on work completed in animal models. To confirm the important role that carbon dioxide has in the initiation of LTF in humans, we took two approaches. The first approach was to show that even though vLTF was not evident during recovery from mild intermittent hypoxia in the presence of uncontrolled carbon dioxide, this phenomenon became evident in the presence of progressively increasing carbon dioxide levels (Khodadadeh et al., 2006; Mateika et al., 2004). Specifically, participants completed rebreathing studies before and following exposure to mild intermittent hypoxia that was unaccompanied by the manifestation of LTF (Khodadadeh et al., 2006; Mateika et al., 2004). During the rebreathing tests, hypoxia was sustained at 50 mmHg and carbon dioxide slowly increased over time. We showed that the ventilatory response to hypoxia at carbon dioxide levels 3 and 6 Torr above the recruitment threshold was elevated during the rebreathing tests completed after compared to before exposure to intermittent hypoxia (Khodadadeh et al., 2006; Mateika et al., 2004). This difference was not evident at carbon dioxide levels that demarcated the recruitment threshold (i.e. 0 Torr above the recruitment threshold) (Khodadadeh et al., 2006; Mateika et al., 2004). We suggested that this increased ventilatory response was due to the initiation of LTF. In subsequent experiments, we demonstrated that LTF of ventilation and genioglossus muscle activity was clearly evident when carbon dioxide levels were sustained above baseline (i.e. 5 mmHg above baseline in this study) throughout and following exposure to mild intermittent hypoxia, but was not evident if we reduced carbon dioxide levels back to baseline levels (Harris et al., 2006). Once this finding was established, we showed that exposure to acute mild intermittent hypoxia in the presence of sustained hypercapnia ( MI H C O 2 ${\mathrm{MI}}{{{\mathrm{H}}}_{{\mathrm{C}}{{{\mathrm{O}}}_2}}}$ ) resulted in vLTF in healthy men and women (Wadhwa et al., 2008) and that the magnitude of the response was independent of sex. We also showed that vLTF was initiated in individuals with OSA (Gerst et al., 2011; Syed et al., 2013) and that the magnitude of the response was greater compared to healthy individuals (Syed et al., 2013). Likewise, exposure to MI H C O 2 ${\mathrm{MI}}{{{\mathrm{H}}}_{{\mathrm{C}}{{{\mathrm{O}}}_2}}}$ lead to the initiation of LTF in individuals with spinal cord injury (Tester et al., 2014). Moreover, we initiated vLTF during wakefulness and sleep in humans (Syed et al., 2013) and showed that the magnitude of the response was greater during wakefulness. We also showed that repeated daily exposure to MI H C O 2 ${\mathrm{MI}}{{{\mathrm{H}}}_{{\mathrm{C}}{{{\mathrm{O}}}_2}}}$ enhanced the magnitude of LTF in humans (Gerst et al., 2011) and was greater in the evening compared to the morning in individuals with obstructive sleep apnoea (Gerst et al., 2011). In the studies completed over the years, the degree to which carbon dioxide was sustained above baseline ranged from 2–5 mmHg (Panza et al., 2023). In all cases, vLTF was evident, although the magnitude of the response varied (Panza et al., 2023) (see section below on 'Potential Mechanisms that Link Carbon Dioxide to the Initiation and Manifestation of LTF' for further discussion of this point). The mechanistic role(s) that sustained elevated levels of carbon dioxide have in the initiation and manifestation of LTF following exposure to MI H C O 2 ${\mathrm{MI}}{{{\mathrm{H}}}_{{\mathrm{C}}{{{\mathrm{O}}}_2}}}$ is not fully established, although there are a number of possibilities. Spinal motoneurons associated with movement and breathing are innervated in part by similar medullary neuronal pathways (e.g. raphe neurons) that are activated by peripheral chemoreflexes that respond to mild intermittent hypoxia. Thus, the inability to initiate LTF in spinal motoneurons could be a result of peripheral chemoreflex feedback that is insufficient to activate downstream mechanisms. Our recent findings support this suggestion (Panza et al., 2023). We showed in humans (n = 124) that the magnitude of LTF is predicted in part by the sensitivity of the hypoxic ventilatory response, which reflects peripheral chemoreflex sensitivity (Panza et al., 2023). It is well established that increasing the level of sustained carbon dioxide for a given level of hypoxia is associated with an increase in the hypoxic ventilatory response (Duffin & Mateika 2013). Given this relationship, the input from the peripheral chemoreceptors known to initiate LTF will be severely diminished in the presence of hypocapnia. Thus, maintaining carbon dioxide levels in the presence of mild intermittent hypoxia serves to ensure that an adequate stimulus originating from the peripheral chemoreceptors exists to initiate LTF. It is also possible that LTF is initiated but does not manifest because uncontrolled carbon dioxide is reduced significantly below a well-defined threshold (referred to as the apnoeic threshold during sleep and the recruitment threshold during wakefulness) leading to the cessation of breathing. The cessation of breathing via reductions in carbon dioxide is easily induced during sleep using artificial ventilation and is evident in some individuals during wakefulness following hyperventilation. Consequently, if carbon dioxide is significantly reduced during the application of mild intermittent hypoxia and the reduction in carbon dioxide endures after exposure, LTF might not be evident in measures of ventilation because of the withdrawal of this powerful stimulus, even though the phenomenon was initiated (Mateika & Narwani 2009). This possibility is particularly applicable to spinal motoneurons that innervate respiratory muscles and receive inputs from medullary respiratory neurons and other neuronal groups (i.e. raphe neurons) that have a role in initiating LTF. Thus, maintaining levels of carbon dioxide may impact both the initiation and manifestation of LTF. Lastly, sustained elevated levels of carbon dioxide, independent of mild intermittent hypoxia, could activate distinct neuronal and cellular pathways that independently initiate LTF (Mateika et al., 2018). Work completed using animal models has indicated that separate neuronal pathways that offset each other are activated by intermittent hypoxia and intermittent hypercapnia (Kinkead et al., 2001). On the other hand, to our knowledge, pathways activated by sustained levels of carbon dioxide in the context of initiating LTF have not been discovered (Mateika et al., 2018). Nonetheless, studies in humans have revealed that sustained levels of hypercapnia can lead to ventilatory drift which suggests that novel mechanisms may be activated by this stimulus (Harris et al., 2006). However, there is also evidence to suggest that sustained hypercapnia is not solely responsible for the magnitude of LTF. We have shown that the magnitude of the drift in ventilation recorded in response to sustained hypercapnia is much smaller than the response recorded when humans are exposed to MI H C O 2 ${\mathrm{MI}}{{{\mathrm{H}}}_{{\mathrm{C}}{{{\mathrm{O}}}_2}}}$ (Gerst et al., 2011). Thus, it is possible that MI H C O 2 ${\mathrm{MI}}{{{\mathrm{H}}}_{{\mathrm{C}}{{{\mathrm{O}}}_2}}}$ initiates LTF via the interaction of separate mechanistic pathways (Mateika et al., 2018). If so, allowing carbon dioxide levels to decrease in an uncontrolled manner would result in the removal of a stimulus that is capable of initiating LTF independently. Given that sustained levels of carbon dioxide appears to have an important role in initiating vLTF in healthy humans (Harris et al., 2006; Wadhwa et al., 2008) and humans living with sleep apnoea (Syed et al., 2013) or spinal cord injury (Tester et al., 2014), MI H C O 2 ${\mathrm{MI}}{{{\mathrm{H}}}_{{\mathrm{C}}{{{\mathrm{O}}}_2}}}$ may ultimately prove to be an effective therapeutic modality that has a multipronged effect on numerous physiological systems including respiratory and cardiovascular outcomes in individuals living with sleep apnoea (Puri et al., 2021). We have begun to explore this possibility and, to date, our work has focused on outcome measures linked to upper airway patency and cardiovascular function. We showed that acute and repeated daily exposure to MI H C O 2 ${\mathrm{MI}}{{{\mathrm{H}}}_{{\mathrm{C}}{{{\mathrm{O}}}_2}}}$ improves upper airway patency (El-Chami et al., 2017; Panza et al., 2022). Reduced critical closing pressure measures indicated that upper airway patency improves following exposure to MI H C O 2 ${\mathrm{MI}}{{{\mathrm{H}}}_{{\mathrm{C}}{{{\mathrm{O}}}_2}}}$ (Panza et al., 2022). In addition, we showed that continuous positive airway pressure (CPAP) required to treat sleep apnoea was reduced as a consequence of improved upper airway patency and as a result treatment adherence improved (Panza et al., 2022). Presently, we are exploring whether exposure to MI H C O 2 ${\mathrm{MI}}{{{\mathrm{H}}}_{{\mathrm{C}}{{{\mathrm{O}}}_2}}}$ also dampens responses to tactile pressure and auditory noise, which are stimuli often experienced when patients are treated with CPAP. If our preliminary findings are correct, these additional modifications might increase the arousal threshold to these stimuli, which could contribute to improving treatment adherence. Given the changes in the critical closing pressure that indicate improvement in upper airway patency we expected that acute exposure to MI H C O 2 ${\mathrm{MI}}{{{\mathrm{H}}}_{{\mathrm{C}}{{{\mathrm{O}}}_2}}}$ would be coupled to decreases in the apnoea/hypopnea index (Syed et al., 2013). This was not the case and we postulated that other forms of plasticity (i.e. progressive augmentation) initiated by MI H C O 2 ${\mathrm{MI}}{{{\mathrm{H}}}_{{\mathrm{C}}{{{\mathrm{O}}}_2}}}$ may override any benefits associated with improved airway patency in the absence of CPAP treatment (Mateika & Narwani 2009). We are presently exploring whether timing of exposure and length of exposure might alter the balance between these forms of plasticity and their impact on apnoea severity. Improvement in treatment adherence could have important implications for modifying cardiovascular outcome measures in individuals with sleep apnoea. Moreover, MI H C O 2 ${\mathrm{MI}}{{{\mathrm{H}}}_{{\mathrm{C}}{{{\mathrm{O}}}_2}}}$ might have direct effects on cardiovascular outcomes independent of improved adherence. To date, we have shown that repeated daily exposure to MI H C O 2 ${\mathrm{MI}}{{{\mathrm{H}}}_{{\mathrm{C}}{{{\mathrm{O}}}_2}}}$ results in a significant decrease in blood pressure in OSA patients treated nightly with CPAP (Panza et al., 2022). We are following up these studies by exploring whether this decrease is evident in the OSA population independent of nightly treatment with CPAP. Likewise, we are exploring whether blood pressure modifications are sustained for long periods of time. Our preliminary data suggests that this is the case because reductions in blood pressure following treatment with MI H C O 2 ${\mathrm{MI}}{{{\mathrm{H}}}_{{\mathrm{C}}{{{\mathrm{O}}}_2}}}$ are sustained for up to 8 weeks in most individuals. There are a number of additional studies to be performed in the coming years. The mechanisms underlying the impact of MI H C O 2 ${\mathrm{MI}}{{{\mathrm{H}}}_{{\mathrm{C}}{{{\mathrm{O}}}_2}}}$ on select outcomes measures are of interest, particularly in the context of teasing out the roles of MIH and sustained carbon dioxide individually and combined. It is also of interest to explore whether sustaining elevated carbon dioxide levels is necessary to attain maximum therapeutic value for all outcome measures or whether this requirement is limited to specific physiologic responses. In addition, studies designed to determine the efficacy of different doses of MI H C O 2 ${\mathrm{MI}}{{{\mathrm{H}}}_{{\mathrm{C}}{{{\mathrm{O}}}_2}}}$ , at the same time as considering the concurrent impact of hypoxic sensitivity on efficacy, comprise an important next step. In the context of these dosing studies, the potential safety issues related to exposure to MI H C O 2 ${\mathrm{MI}}{{{\mathrm{H}}}_{{\mathrm{C}}{{{\mathrm{O}}}_2}}}$ should be further explored. It is possible that some subgroups within a given population (e.g. low vs. high hypoxic sensitivity) will not benefit from this therapy or will experience detrimental outcomes, which could be an additional explanation for the variable responses that have been reported to date. However, it should be noted that in our studies and other published studies significant adverse events linked to MI H C O 2 ${\mathrm{MI}}{{{\mathrm{H}}}_{{\mathrm{C}}{{{\mathrm{O}}}_2}}}$ have not been reported. Likewise, based on prior work the timing of administration of this stimulus and its effectiveness on outcome measures, particularly as it relates to improvement in the severity of OSA, will be an important contribution to the existing literature. Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article. No competing interests declared. J.H.M. was responsible for the conception of the work. J.H.M., R.B. and D.M.K. contributed to drafting the work or revising it critically for important intellectual content. J.H.M., R.B. and D.M.K. approved the final version of the manuscript submitted for publication and agree to be accountable for all aspects of the work by ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved. Lastly, all persons designated as authors qualify for authorship, and all those who qualify for authorship are listed. This work was supported by the United States Department of Veterans Affairs (I01CX00125, IK6CX002287) and the National Institute of Heart, Lung and Blood (R01HL085537).

The effect of changing tree species composition in favor of a greater representation of Douglas fir at the expense of Norway spruce on the carbon pool of Central European forests has not yet been investigated. Here, we compare the allocation of aboveground biomass and carbon stock in Douglas fir and spruce at the tree and stand level. At the tree level, Douglas fir accumulated, on average, 16.9% more aboveground biomass than Norway spruce. A greater amount of biomass was allocated mainly in the wood and bark of Douglas fir stem. For these biomass compartments, the difference between Douglas fir and Norway spruce was 21.1% and 60.3%, respectively. Spruce allocated more biomass in the crown, where the difference was 25.6% compared to Douglas fir. In needle biomass, Norway spruce exceeded Douglas fir by 84%. At the stand level, the analysis of model stands revealed that pure Norway spruce stands accumulated more carbon in the high and medium quality sites. As the site quality decreased, so did the differences in the amount of stored carbon. The higher carbon sink in Norway spruce stands was also confirmed in the analysis of real Norway spruce and Douglas fir stands. The difference in the carbon stock of young, medium-aged, and mature stands was 11.5%, 14.8%, and 1%, respectively. The positive balance in favor of spruce is mainly due to significantly higher numbers of trees per ha in Norway spruce stands. A positive effect of a greater representation of Douglas fir on the carbon budget of forest stands was not confirmed.

Tree mortality caused by bark beetles has increased in recent decades in both Europe and North America. In a large recent outbreak in central Sweden the bark beetle Polygraphus poligraphus was often found together with the spruce bark beetle Ips typographus in killed trees. To increase the understanding of the aggregation behavior of P. poligraphus we used solid phase microextraction (SPME) to collect volatile organic compounds (VOCs) released from single P. poligraphus males, with and without added females, colonizing Norway spruce stem sections and analyzed the sampled compounds by combined gas chromatography and mass spectrometry (GC-MS). High amounts of terpinen-4-ol, a substance found in the hindguts of P. poligraphus males in earlier studies, were released by colonizing males. The emission of both enantiomers of terpinen-4-ol was monitored by GC-MS over time as the males aged in the absence and presence of females. Single males emitted (R)-(-)-terpinen-4-ol for up to 60 days in high enantiomeric purity but the enantiomeric excess (ee) varied between males, and also for the same individual, over time from 96.3% to 99.3% ee. In the presence of females, males also emitted terpinen-4-ol for up to 50 days but now in lower amounts and with lower enantiomeric purity varying from 67.7% ee to 99.3% ee. Small quantities of other volatile compounds were emitted from the colonizing beetles including cis- and trans-4-thujanol, both of which were previously shown to be present in the hindguts of males. In earlier studies frontalin was found to attract P. poligraphus, but in our study it was not identified among emitted compounds from colonizing beetles.

The rotation lengths of intensively managed production forests may be altered to achieve a variety of goals, with correspondingly implications for biodiversity. Here we consider the potential implications of shortened rotation times for biodiversity in planted monocultures of the two most common production tree species in Sweden, Scots pine (Pinus sylvestris) and Norway spruce (Picea abies). To do so we surveyed bird, bryophyte, epiphytic lichen and vascular plant diversity in 80 and 55-year-old stands; stand ages which approximate present-day and potential future rotation lengths in this region respectively. We found clear differences in the species communities of the 55 compared to the 80-year-old stands for both understory species and epiphytes, but not for birds. Nevertheless, bird species richness was still highest in the 80-year-old Norway spruce dominated stands. Dead wood amount was also highest the 80-year-old Norway spruce stands. Highest species richness of epiphytic lichens was found in 80-year-old Scots pine stands. However, 55-year-old Scots pine stands had a higher understory species richness and diversity than the older Scots pine stands, including a larger number of open land species. The 80-year-old forest stands examined may be considered old with respect to production forest rotation lengths in Sweden but are relatively young when comparing stand ages of unmanaged natural forest stands. Nevertheless, our results indicate that shortening the rotation time of Scots pine and Norway spruce, in this part of Sweden from 80 to 55 years, could have important consequences for forest biodiversity. These consequences are primarily inferred from the likely implications from shortened rotations for lichens community composition and diversity in both Norway spruce and Scots pine stands, as well as impacts on understory plant species in Norway spruce stands.

Journal Article Levels of endogenous ethylene, carbon dioxide, and soluble pectin, and activities of pectin methylesterase and polygalacturonase in ripening tomato fruits Get access Masayoshi Sawamura, Masayoshi Sawamura Department of Plant Physiology, Agricultural UniversityWageningen, The Netherlands Search for other works by this author on: Oxford Academic Google Scholar Erik Knegt, Erik Knegt Department of Plant Physiology, Agricultural UniversityWageningen, The Netherlands Search for other works by this author on: Oxford Academic Google Scholar Johan Bruinsma Johan Bruinsma Department of Plant Physiology, Agricultural UniversityWageningen, The Netherlands Search for other works by this author on: Oxford Academic Google Scholar Plant and Cell Physiology, Volume 19, Issue 6, September 1978, Pages 1061–1069, https://doi.org/10.1093/oxfordjournals.pcp.a075672 Published: 01 September 1978 Article history Received: 16 February 1978 Published: 01 September 1978

Objectives The purpose of this study is to examine changes in carbon dioxide concentration and noise levels in classrooms, and to analyze the effects of these changes on elementary school students' attention and problem behaviors. Methods First, To measure the change of carbon dioxide and noise levels in classrooms, theses levels were measured in general classroom, specific subject classroom, and specific subject classroom with ventilation system operating. Second, to analyze the effect of carbon dioxide and noise levels on students’ attention and problem behavior, situations where both carbon dioxide and noise levels are low (cLnL), only carbon dioxide levels are high (cHnL), and only noise levels are high (cLnH), and both carbon dioxide and noise levels are high (cHnH). Results The carbon dioxide concentration in each classroom ranged from 400 to 1951.8 ppm, and the noise level ranged from 12.6 to 96.6 db(A). As a result of analyzing the effects of carbon dioxide and noise levels on attention and problem behavior, it was resulted that the higher the noise level, the negatively correlated with Work instruction comprehension, Selective attention, and Sustained attention. In addition, problem behaviors occurred the least (3 times) in the cLnL group and the most (31 times) in the cHnH group. Conclusions Carbon dioxide concentration and noise levels in the school exceeded the standard. The learning environment can affect not only students' attention and problem behavior, but also their health. To improve this, physical environment improvement is required.

Because sweetpotato [Ipomoea batatas (L.) Lam.] stem cuttings regenerate very easily and quickly, a study of their early growth and development in microgravity could be useful to an understanding of morphological changes that might occur under such conditions for crops that are propagated vegetatively. An experiment was conducted aboard a U.S. Space Shuttle to investigate the impact of microgravity on root growth, distribution of amyloplasts in the root cells, and on the concentration of soluble sugars and starch in the stems of sweetpotatoes. Twelve stem cuttings of 'Whatley/Loretan' sweetpotato (5 cm long) with three to four nodes were grown in each of two plant growth units filled with a nutrient agarose medium impregnated with a half-strength Hoagland solution. One plant growth unit was flown on Space Shuttle Colombia for 5 days, whereas the other remained on the ground as a control. The cuttings were received within 2 h postflight and, along with ground controls, processed in approximately 45 min. Adventitious roots were counted, measured, and fixed for electron microscopy and stems frozen for starch and sugar assays. Air samples were collected from the headspace of each plant growth unit for postflight determination of carbon dioxide, oxygen, and ethylene levels. All stem cuttings produced adventitious roots and growth was quite vigorous in both ground-based and flight samples and, except for a slight browning of some root tips in the flight samples, all stem cuttings appeared normal. The roots on the flight cuttings tended to grow in random directions. Also, stem cuttings grown in microgravity had more roots and greater total root length than ground-based controls. Amyloplasts in root cap cells of ground-based controls were evenly sedimented toward one end compared with a more random distribution in the flight samples. The concentration of soluble sugars, glucose, fructose, and sucrose and total starch concentration were all substantially greater in the stems of flight samples than those found in the ground-based samples. Carbon dioxide levels were 50% greater and oxygen marginally lower in the flight plants, whereas ethylene levels were similar and averaged less than 10 nL.L (-1). Despite the greater accumulation of carbohydrates in the stems, and greater root growth in the flight cuttings, overall results showed minimal differences in cell development between space flight and ground-based tissues. This suggests that the space flight environment did not adversely impact sweetpotato metabolism and that vegetative cuttings should be an acceptable approach for propagating sweetpotato plants for space applications.

The Interactive Effects of Atmospheric Carbon Dioxide and Light on Stem Elongation in Seedlings of Four Species

Hepatocellular carcinoma (HCC) is a common malignancy in the world with high morbidity and mortality rate. Identification of novel biomarkers in HCC remains impeded primarily because of the heterogeneity of the disease in clinical presentations as well as the pathophysiological variations derived from underlying conditions such as cirrhosis and steatohepatitis. The aim of this study is to search for potential metabolite biomarkers of human HCC using serum and urine metabolomics approach. Sera and urine samples were collected from patients with HCC (n = 82), benign liver tumor patients (n = 24), and healthy controls (n = 71). Metabolite profiling was performed by gas chromatography time-of-flight mass spectrometry and ultra performance liquid chromatography-quadrupole time of flight mass spectrometry in conjunction with univariate and multivariate statistical analyses. Forty three serum metabolites and 31 urinary metabolites were identified in HCC patients involving several key metabolic pathways such as bile acids, free fatty acids, glycolysis, urea cycle, and methionine metabolism. Differentially expressed metabolites in HCC subjects, such as bile acids, histidine, and inosine are of great statistical significance and high fold changes, which warrant further validation as potential biomarkers for HCC. However, alterations of several bile acids seem to be affected by the condition of liver cirrhosis and hepatitis. Quantitative measurement and comparison of seven bile acids among benign liver tumor patients with liver cirrhosis and hepatitis, HCC patients with liver cirrhosis and hepatitis, HCC patients without liver cirrhosis and hepatitis, and healthy controls revealed that the abnormal levels of glycochenodeoxycholic acid, glycocholic acid, taurocholic acid, and chenodeoxycholic acid are associated with liver cirrhosis and hepatitis. HCC patients with alpha fetoprotein values lower than 20 ng/ml was successfully differentiated from healthy controls with an accuracy of 100% using a panel of metabolite markers. Our work shows that metabolomic profiling approach is a promising screening tool for the diagnosis and stratification of HCC patients.

Identification of acetates of secondary straight-chain alcohols by gas chromatography—Mass spectrometry

In metabolomics, the purpose is to identify and quantify all the metabolites in a biological system. Combined gas chromatography and mass spectrometry (GC/MS) is one of the most commonly used techniques in metabolomics together with 1H NMR, and it has been shown that more than 300 compounds can be distinguished with GC/MS after deconvolution of overlapping peaks. To avoid having to deconvolute all analyzed samples prior to multivariate analysis of the data, we have developed a strategy for rapid comparison of nonprocessed MS data files. The method includes baseline correction, alignment, time window determinations, alternating regression, PLS-DA, and identification of retention time windows in the chromatograms that explain the differences between the samples. Use of alternating regression also gives interpretable loadings, which retain the information provided by m/z values that vary between the samples in each retention time window. The method has been applied to plant extracts derived from leaves of different developmental stages and plants subjected to small changes in day length. The data show that the new method can detect differences between the samples and that it gives results comparable to those obtained when deconvolution is applied prior to the multivariate analysis. We suggest that this method can be used for rapid comparison of large sets of GC/MS data, thereby applying time-consuming deconvolution only to parts of the chromatograms that contribute to explain the differences between the samples.

Cadmium (Cd), being one of the most toxic elements, exerts negative effects on plant growth and development. Ethylene (ET) has a prime significance in abiotic stress-induced cellular modulations and regulation of plant development. In the present study, seeds of a Cd-tolerant (NM-98) and -sensitive (NM-28) genotype were primed using 50 µM ACC (ET precursor 1-amino cyclopropane-1-carboxylic acid) and ET inhibitors (1 mM AVG/aminoethoxyvinylglycine, 0.5 mM PZA/pyrazinamide or 0.25 mM AgNO3/silver nitrate). Subsequently, these were allowed to grow in soil containing 0 or 50 µM CdCl2. After germination, ACC, AVG, PZA or AgNO3 was again applied, accordingly, as foliar spray. In NM-98, application of ACC resulted in reduced root fresh (38%) and dry (38%) weight, shoot fresh weight (39%), total phenolics (22%), catalase/CAT and peroxidase/POD activity (21% and 16%, respectively) under Cd stress. Contrarily, application of inhibitors (AVG, PZA and AgNO3), under Cd stress, improved plant biomass, photosynthetic pigments, total phenolics, total soluble proteins, activity of CAT and POD in NM-98 as compared to NM-28 genotype that exhibited opposite response. It was concluded that contrasting modulations observed in NM-98 and NM-28 towards exogenous ACC (or inhibitors) was linked with Cd-tolerance level of the two genotypes where exogenous ACC likely significantly raised endogenous ET level in NM-98 leading to disrupted antioxidants status and reduced growth. However, AVG, PZA and AgNO3 maintained ET levels and thus NM-98 exhibited less oxidative stress. Meanwhile, exogenous ACC assisted growth of NM-28 in Cd-stressed soil by modulating the antioxidative defence mechanism, possibly by maintaining endogenous ET concentrations. Further molecular-level research is recommended to investigate the underlying mechanisms associated with each component of ET biosynthesis.

The endogenous gibberellins (GAs) in leaf tissues of two day-neutral cultivars (Rapella and Selva) of strawberry (Fragaria × ananassa Duch.) were analysed using combined gas chromatography -- mass spectrometry (GC-MS). Seven of the later members of the 13-hydroxylation GA biosynthetic pathway were identified, by comparison of Kovats retention indices and mass spectral data obtained for methyl ester trimethylsilyl ether derivatives, either with data obtained from authentic compounds or literature values. GA1, GA3, GA8, GA17, GA19, GA20 and GA29 were detected in extracts of both cultivars.