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The carbon stable isotopic composition of mosses: A record of temperature variation
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Organic Geochemistry Organic Geochemistry 38 (2007) 1770–1781 www.elsevier.com/locate/orggeochem
The carbon stable isotopic composition of mosses: A record of temperature variation Grzegorz Skrzypek
a,b,*
, Adam Kału_zny b, Bronisław Wojtun´ c, Mariusz-Orion Je˛drysek b
a
b
Laboratory for Stable Isotope Geochemistry, Department of Earth and Environmental Science, The University of Texas at San Antonio, One UTSA Circle, San Antonio, 78249 TX, USA Laboratory of Isotope Geology and Geoecology, Institute of Geological Sciences, The University of Wrocław, Cybulskiego 30, 50-205 Wrocław, Poland c The Wrocław University of Environmental and Life Sciences, Department of Botany and Plant Ecology, pl. Grunwaldzki 24a, 50-363 Wrocław, Poland Received 8 October 2006; received in revised form 8 February 2007; accepted 1 May 2007 Available online 5 June 2007
Abstract Samples of two moss species, Sphagnum and Polytrichum, were collected at 19 points selected along vertical transects in the Karkonosze Mts. (739–1393 m) and the Izerskie Mts. (500–1100 m), SW Poland. Continuous automatic monitoring of temperature and humidity at the sampling points was carried out (1 h intervals) through the entire growth season (2004). A strong correlation between d13C value of plant tissue cellulose and mean air temperature was noted for both areas and sampling periods (R2 between 0.92 and 0.64). It was calculated that a 1 °C increase in air temperature during the growing season results in a 1.6& (Sphagnum) and a 1.5& (Polytrichum) decrease in d13C. In contrast, it seems that humidity shows no clear influence on the carbon isotope composition of mosses. Ó 2007 Elsevier Ltd. All rights reserved.
1. Introduction Climatic variations during the last millennium are relatively well documented (e.g. Barber, 1981; Lamb, 1985; Lipp et al., 1991; White et al., 1994; Je˛drysek et al., 2003; Barber et al., 2004a,b; Lang* Corresponding author. Present address: Laboratory for Stable Isotope Geochemistry, Department of Earth and Environmental Science, The University of Texas at San Antonio, One UTSA Circle, San Antonio, 78249 TX, USA. Tel.: +1 210 7248430; fax: +1 210 4584469. E-mail addresses: [email protected], [email protected]. wroc.pl (G. Skrzypek).
don and Barber, 2005). However, the role of our industrial civilization, as well as the scale of the changes, is still not clear. Two major questions are: ‘‘Which aspects of reconstructed past climates allow us to better understand present climatic variation?’’ and ‘‘Is it possible to quantitatively predict future climatic trends?’’ The stable isotope composition of organic matter from peat profiles may potentially bring crucial information to answer these questions (e.g. O’Leary et al., 1986; Francey and Farquhar, 1982; Farquhar et al., 1989; Aucour et al., 1994; Boutton et al., 1998; Dawson et al., 2002). Although evidence suggests that temperature
0146-6380/$ - see front matter Ó 2007 Elsevier Ltd. All rights reserved. doi:10.1016/j.orggeochem.2007.05.002
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is the dominant (primary) factor controlling d13C values in peat (Skrzypek and Je˛drysek, 2005), there is no well documented quantitative calibration defining the temperature control on the carbon isotope composition (d13C) in peat-forming plants (Skrzypek and Je˛drysek, 2001, 2005). The role of secondary factors can be so important that the primary signal can be overwhelmed. For certain habitats it is possible to exclude most of these secondary factors by properly selecting sampling points, both spatially and temporally, e.g. where and when the humidity, insolation and soil conditions are very similar, thereby permitting the exclusion of these factors. Moreover, the gametophytes of mosses do not have stomata, so excluding the so called ‘‘stomata effect’’ that is important in the case of carbon stable isotope fractionation during CO2 assimilation by higher plants (the scale of the effect depends on the humidity variation). Although temperature has an extremely important influence on fractionation, some authors have shown that precipitation (humidity) and atmospheric pollution, rather than temperature, are the most important factors controlling d13C values in vascular plants (e.g. Je˛drysek et al., 2003). Likewise, Ko¨rner et al. (1991, 1988) and Menot and Burns (2001) suggest the crucial role of altitude (atmospheric pressure and CO2 atmospheric partial pressure). However, the carbon isotopic composition of plant tissue is not a direct effect of atmospheric CO2 – plant tissue isotope fractionation, but rather results from complex processes controlled by environmental conditions during metabolism (e.g. O’Leary et al., 1986; Farquhar et al., 1989). The quantitative role of these factors in controlling isotopic fractionation is proposed in the equation defined by Farquhar et al. (1989). During plant growth, isotopic fractionation generally proceeds in two stages (Park and Epstein, 1960; O’Leary, 1981, 1986; Farquhar et al., 1989): (1) adsorption and diffusion of CO2 into the plant tissue and (2) the initial carboxylation. Plant growth results in an 18–27& (C3 plants) and 4–6& (C4 plants) 12C enrichment in the plant tissue with respect to atmospheric CO2 (O’Leary, 1981, 1986; Lajtha and Marshal, 1994), with the value dependent on the plant species and temperature during CO2 assimilation and carboxylation. Many authors, on the basis of various plant studies, have proposed significantly different Fq values, defined as the change in the d13C value of plant tissue resulting from a 1 °C change in temperature
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(Skrzypek and Je˛drysek, 2005), varying from 1 to 2.4&/°C (Smith et al., 1973; Troughton and Card, 1975; Griensted et al., 1979; O’Leary, 1981; Leavitt and Long, 1986; Lipp et al., 1991; Robertson et al., 1997). According to calibrations based on peat (Skrzypek and Je˛drysek, 2005) the Fq value is 0.6&/°C. Likewise, Menot and Burns (2001) found that, in peat-forming Sphagnum, Fq varies from 0.2 to 0.4&/°C, but the temperatures were not measured exactly at the sampling places but at meteorological stations in few kilometers distance from sampling points, at the respective altitude. Therefore, it is difficult to precisely reconstruct temperature variations based on the d13C analysis of peat profiles. To address this problem, we analyzed the d13C values for mosses, peat-forming Sphagnum and Polytrichum and continuously monitored both the temperature and humidity at the specific sampling points where the samples were collected twice during the growth season. 2. Goal and strategy The major goal was to describe the influence of temperature on the d13C value of moss tissue, the direct result of which is calculation of Fq factors [change in d13C value during a 1 °C increase in temperature (&/°C)]. The potential influence of other factors, such as humidity and habitat condition, was also analyzed. It was therefore necessary to find natural locations of mosses growing in similar habitats but with different temperature ranges during the growing season. These criteria are attained on relatively steep mountain slopes where a significantly different altitude of sampling points (e.g. 100 m) can be utilized, with little difference in horizontal distance (only a few km). Usually, an increase in altitude of 100 m results in a decrease in air temperature of ca. 0.6 °C. 3. Sampling points and methods Samples of two moss species, Sphagnum girgensohnii Russow and Polytrichum commune Hedw., were collected from northern hillsides in the Karkonosze Mts. (vertical transect 739–1393 m) and Izerskie Mts. (500–1101 m), SW Poland (Fig. 1). Details about locations of the 19 sampling/observation points are shown in Table 1 (RH = relative humidity). The two species were selected because of their common occurrence in the selected regions. Furthermore, their ecological preferences are only
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Fig. 1. Location of sampling points in two areas (SW Poland); the Izerskie Mts. are located about 30 km west of the Karkonosze Mts. The mountain slopes (IZ500–IZ11KO and KR600–KR1400) face north. The short vertical transect IZ11KO–IZ84HA is located at the southwest facing slope. Point KR700 is excluded because of data logging defects.
Table 1 d13C values of moss samples and meteorological data recorded (HOBO loggers) at the place of collection Location
Name
Altitude (m)
Average for season #1 (4 July 2004) T (°C)
RH (%)
d13C [&,VPDB], Poly
d13C [&,VPDB], Sph
Average for season #2 (4 October 2004) T (°C)
RH (%)
d13C [&,VPDB], Poly
d13C [&,VPDB], Sph
Izerskie Mts.
IZ84HA IZ85HA IZ90JJ IZ10JJ IZ11KOP IZ10SI IZ90SI IZ80SI IZ700 IZ600 IZ500
843 857 902 998 1101 1002 899 799 695 597 500
9.7a 8.7 8.1 8.0 7.7 8.5 9.1 9.6 9.7 10.1 11.1
77 81 89 83 85 80 82 80 76 76 67
26.31b 26.75 27.55 24.86 25.02 24.88 25.55 26.12 26.23 28.06 28.95
28.50c 28.03 30.56 26.21 25.03 27.51 28.62 26.00 28.73 29.67 30.94
10.8a 9.7 9.3 9.2 9.1 9.9 10.2 10.7 10.8 11.2 12.0
78 82 90 84 85 81 83 82 78 78 72
25.80b 25.82 27.18 24.81 24.79 24.63 25.90 25.89 28.02 28.45 30.07
29.36c n/a 31.12 26.60 25.93 26.96 29.14 28.31 30.42 30.10 30.34
Karkonosze Mts.
KR1400 KR1300 KR1200 KR1100 KR1000 KR900 KR800 KR700 KR600
1393 1266 1240 1091 1052 914 834 675 602
6.5 6.8 7.3 8.1 9.0 9.2 9.4 n/ad 9.7
85 85 82 82 79 81 83 n/ad 75
24.60 25.02 25.45 28.53 25.14 28.96 25.96 26.43 27.44
27.52 26.43 26.19 30.04 27.83 31.58 31.30 26.78 31.86
7.9 8.0 8.7 9.2 10.1 10.4 10.7 n/ad 10.9
85 85 80 81 81 82 84 n/ad 76
24.45 24.11 24.74 28.56 24.72 30.60 26.27 26.75 27.55
27.52 26.86 26.58 31.57 27.45 30.35 30.86 26.68 31.83
a
The temperatures are average values calculated from 1 h intervals measured for a period starting at the beginning of the growing season (April 15–19) and ending with sample collection (July 2–4 and October 15–17, 2004) for short growth ‘‘Season #1’’ and long growth ‘‘Season #2’’, respectively. b Polytrichum commune. c Sphagnum girgensohnii. d n/a – lost sample (Sph IZ85HA), damaged sensor (KR700).
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slightly different the anatomy and ecophysiology of S. girgensohnii is similar to that of other peat-forming species of the genus Sphagnum in ombrotrophic bogs or poor fens. Although in the Western Sudetes S. girgensohnii and P. commune are shade-tolerant and have similar trophic amplitude, the former prefers wet microhabitats, while the latter is typically found at dryer sites (Wojtun´, 2006). Special attention was paid to the selection of sampling points where environmental conditions were, to the furthest extent possible, similar, especially the presence of a peat layer, relatively stable soil moisture during growing season and similarity in the surrounding ecosystem. Despite these assumptions, local conditions varied according to altitude and differences in forest stand and condition. Similarity in conditions amongst the sampling sites is suggested by the close proximity of the sample locations. The furthest distance between sampling points in each transect was < 5.7 km (Izerskie Mts.) and < 4.9 km (Karkonosze Mts.; Fig. 1). However, the local microclimate may differ slightly at each location (e.g. concentration of CO2 and d13C in air). Based upon dominant vegetation type, observation points could generally be classified into three major groups: 1. A sub-alpine dwarf pine group dominated by Pinetum mugo sudeticum; this group includes only KR1400. 2. A mire group situated in poor fens. This group includes three sampling points: KR1300, IZ84HA and IZ85HA. 3. A forest group situated inside a spruce forest. This group may be further divided into two sub-groups: the first includes points KR1100, KR1200, IZ90SI, IZ90JJ, IZ10SI, IZ10JJ and IZ11KO, located in the upper forest belt, and the second points KR600, KR800, KR900, KR1000, IZ500, IZ600, IZ700 and IZ80SI in the lower forest belt. Within the forest group, points KR800 and KR1000 were different as they are located in local clearings. Two points in time define the duration of the growing season for all specimens: the normal weather-controlled start of growth and the termination of growth by sample collection. As sample collection was carried out at the same time for all samples, the variable for the length of the growing season is the elevation-controlled start of plant growth. The start of the growing season for each
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location was defined as that period when the average temperature of five consecutive days reached 5 °C. All samples were collected during two field seasons in 2004, one during July 2–4, and the other during October 15–17, resulting in a short and long growth season for comparison (season 1st July and 2nd October). Throughout the entire 2004 growing season, for all sample locations, automated measurements of temperature and humidity were collected at 60 min intervals using a U8 HOBO data logger (located about 1.5–2.0 m above ground and not further than 5 m from plant sampling points). The length of the growing season varied for different points depending on altitude, but the variation was < 15 days (start date between April 15 and 29). In some places, the real growth period can be shorter, because of longer snow cover. After botanical identification, the whole batches of individual moss species were washed, dried under vacuum and homogenized (2–3 g dry matter). The cellulose was extracted following the modified technique developed by Epstein et al. (1976). This procedure consisted of three major steps (e.g. Skrzypek et al., 2007): (1) multiple chemical treatments: soak in 1:1 C6H6–MeOH (24 h), rinse with HCl (4%), boil with water (1 h), heat (70 °C) with CH3CO2H and NaCl, soak in 17% NaOH (1 h) and CH3CO2H for 10 min; (2) nitrification: dry samples were nitrated using 100% HNO3 (48 h); (3) dissolution and precipitation of nitrocellulose: dissolution in acetone and, after centrifugation and decantation, the nitrocellulose was precipitated from the solution by quickly adding redistilled water. After each chemical treatment, samples were washed with redistilled water and after each step were vacuum dried. Stable isotope analysis was carried out using an off-line preparation system technique for maximum precision. About 3–5 mg of pure nitrocellulose obtained from the plant materials were combusted with CuO wire in a sealed quartz tube under vacuum at 900 °C (Je˛drysek and Skrzypek, 2005). The CO2 produced was cryogenically purified off-line and introduced into an isotope ratio mass spectrometer (IRMS Finnigan-Mat Delta E/dual inlet) for stable carbon isotope ratio analysis. The d13C values were normalized against NBS22 and USGS24 international standards and are reported relative to the Vienna Pee Dee Belemnite (VPDB) scale, with ± 0.05& analytical precision. The d13CVPDB value is defined as the relative difference, in parts per thousand (&), between the isotope ratio of the sample and the international standard (value determined
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by International Atomic Energy Authority, IAEA, Vienna). Least squares regression analysis was performed with d13C as the response variable and RH or T as the predictor variable. The relationship was described with a simple linear model, y = ax + b, where x = d13C and y = T or RH, respectively. Traditional statistical tests assume that the relationship in question is true and uses data to detect differences. Because we wished to use our data to look for similarities, we also calculated probability (P) associated with a Student’s paired t-test, with a two-tailed distribution. 4. Results and discussion The d13C values of nitrocellulose extracted from moss tissue varied from 31.86 to 25.03& for Sphagnum (Sph) and from 30.60 to 24.11& for Polytrichum (Poly) (Table 1). No significant differences were observed between the same moss species
collected in the Izerskie and Karkonosze Mts. (P-values range from 0.3 to 0.6, paired t-test, twotailed). However, it was observed that the d13C values for Polytrichum were more positive than those for Sphagnum. The average difference observed was about 2.21&, but varied over a wide range, depending on location and sampling season (s.d. 1.36&). The mean air temperature (T) during long growth season (#2) varied according to altitude as follows: from 9.1 to 12.0 °C for the Izerskie (altitude 500–1101 m) and 7.9 to 10.9 °C for the Karkonosze Mts. (602–1394 m). The relative humidity (RH) varied within a narrow range: for the Izerskie Mts. 72% to 90% (mean 82%) and for the Karkonosze 76–85% (mean 82%; Table 1). Very high and relatively constant RH at all sampling points was due to frequent rain (total mean annual precipitation in these regions is 1400 mm/year). A very high negative correlation exists between the temperature and humidity of the air overall (R2 varies from 0.86 to 0.63), which is interpreted to be the
Fig. 2. Correlation between temperature and humidity (T–RH) observed in two regions the Karkonosze Mts. and Izerskie Mts. The outliers excluded include IZ90JJ for the linear regression analyses (Izery, graphs a and b) and KR800 (Karkonosze, c and d).
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effect of cloud cover and the resultant lower insolation (Fig. 2). However, the observed humidity (measured 1.5 m above ground) is likely to be lower than the humidity directly at ground level where the moss layers occurred. A limited number of previous studies have investigated the relationship between plant carbon stable isotope composition and altitude (Ko¨rner et al., 1988, 1991; Menot and Burns, 2001). In general, d13C changed with altitude for three Sphagnum species (n = 10, 8, 6), observed by Menot and Burns (2001), but correlations were weak (R2 = 0.25, 0.48, 0.17). No clear correlation with air temperature or humidity was reported. In our study, variability in d13C value with respect to temperature and humidity of air was analyzed separately for species, location, and season. 4.1. d13C/temperature relationship A strong correlation between d13C value of moss tissue cellulose and temperature exists for both areas and sampling periods (Fig. 3). The linear regression equations and the R2 coefficients were calculated for the respective data pairs T (°C) and d13C (&). R2 ranged from 0.76 to 0.92 for the Izerskie Mts. and from 0.64 to 0.87 for the Karkonosze Mts. (Table 2). The relationship between d13C value and temperature was described with a simple linear regression model T = a(d13C) + b, where slope (a) represents Fq value (&/°C). The calculated Fq value (slope) differed slightly for each moss genus depending on location and season [Sphagnum: 1.78 to 1.56&/°C (s.d. 0.12); Polytrichum: 1.75 to 1.15&/°C (s.d. 0.31)]. The average value of d13C decreases approximately 1.62& for Sphagnum and 1.46& for Polytrichum for each 1 °C increase in air temperature during the growing season, which corresponds to a +1& increase for each 0.62 °C and 0.72 °C decrease, respectively. The Fq value did not vary significantly, and no significant differences between d13C of moss species collected during different sampling seasons were found (P-value from 0.06 to 0.95, unpaired t-test, two-tailed). This observation was confirmed for both moss genera by the high correlation between d13C value for moss pairs collected during July and October; R2 varies between 0.80 and 0.95 (Fig. 4). These high R2 values suggest that the same mechanism defines the carbon stable isotope fractionation for both seasons and moss species. How-
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ever, in the case of Polytrichum, some difference was observed between seasons. For the first season, Fq was 1.15 and 1.20&/°C and for the second season, 1.75 and 1.68&/°C for Polytrichum and Sphagnum, respectively. Different averages of air temperature occurred during shorter (#1) and longer (#2) seasons; therefore the d13C of plant tissue differs, respectively. The d13C value of moss at a place where the temperature of vegetation season was lower (e.g. at higher altitude) was more positive than where the temperature was higher. This difference, defined as Fq (&/°C), and calculated for each species, varied slightly depending on season, location and environmental conditions. The variation in Fq value was narrower for Sphagnum (s.d. 0.12) than Polytrichum (0.31; Table 2). The Fq standard deviation can be considered as an error in the calculations, as a result of the minor influence of local parameters as well as differences between the field-recorded temperature and actual temperature during photosynthesis. Thus, the average value calculated for each species (for two seasons and two locations) can be used as the most reliable. On the other hand, we observed only slight differences between the species (Fq 1.6 and 1.5&/°C), these probably being due to small differences in physiology and environmental preference. Data for two sampling points in Karkonosze, KR1000 (1052, both species and periods) and KR800 (834 m, Polytrichum only, both periods) and two points from Izerskie IZ90JJ (902 m, both species and periods) and IZ80SI (799 m, Sphagnum, first period only) do not match the general linear regressions trend for T–d13C (Fig. 3). These points were excluded from the calculation of linear regression equations and R2 (open symbols in all figures), based on multiple point diagnostics of outliers (e.g. Rousseeuw and Leroy, 2003). The points, which do not match the general T–d13C trend (Fig. 3), lie on the regression line for d13C–d13C (July–October; Fig. 4a–d). This fact suggests that the offset in these four points is not a random variation and may result from the specific location relative to local conditions. However, no specific conditions have been observed that can explain the inconsistency. Points KR1000 and IZ80SI are located in clearings within local spring mires, whereas IZ90JJ and KR800 are located in spruce forest where the surrounding floor is covered with spruce litter. The d13C values of plants collected at IZ80SI, KR800 and KR1000 are significantly more positive than one would
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Fig. 3. Correlation between temperature (T) and stable carbon isotope composition (d13C) of cellulose extracted as nitrocellulose from mosses (Polytrichum and Sphagnum) from the Izerskie Mts. and Karkonosze Mts., SW Poland. The temperatures are average values calculated from 1 h intervals measured for a period starting at the beginning of the growing season (April 2004) and ending with sample collection (July or October 2004). Points IZ90JJ (graphs a–d), IZ80SI (a) KR1000 (e–h), and KR800 (g and h) are excluded from calculations of R2 and linear regression analyses (open symbols).
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Table 2 Regression analysisa of temperature (T, °C) and relative humidity (RH, %) changes in carbon stable isotope discrimination (d13C, &) Species
Location/sampling period
n
T = a(d13C) + b a
RH = a(d13C) + b b
R
2
a
R2
b
Polytrichum
Izerskie/1 Izerskie/2 Karkonosze/1 Karkonosze/2 Average (s.d.)
10 10 6 6
1.15 1.75 1.20 1.68 1.45 (0.31)
15.64 8.22 17.10 11.25
0.76 0.77 0.68 0.64
0.23 0.40 0.30 0.26 0.30 (0.07)
44.41 58.39 51.00 47.36
0.73 0.77 0.33 0.27
Sphagnum
Izerskie/1 Izerskie/2 Karkonosze/1 Karkonosze/2 Average (s.d.)
9 9 7 7
1.56 1.64 1.78 1.49 1.62 (0.12)
13.79 11.46 14.74 15.35
0.92 0.80 0.87 0.69
0.29 0.33 0.46 0.33 0.35 (0.07)
51.26 54.70 67.26 56.27
0.79 0.64 0.38 0.23
a The relation was described with a simple linear model y = ax + b, where x = d13C and y = T or RH, respectively. The value ‘‘a’’ in the linear equation represents Fq factor d13C/T (&/°C) or d13C/RH (&/%).
Fig. 4. Correlation of d13CJuly–d13COctober pairs for Sphagnum and Polytrichum collected at each sampling point during July and October seasons.
expect on the basis of the general trend. One possible explanation, for an effect like this, may lie in the observation that samples IZ80SI and KR1000 are located in the area surrounded by watered mires and, as a result, would not have been exposed to conditions of drought, which could cause a tempo-
rary pause in assimilation and a shift in d13C value (in less watered locations). However, this is speculative as the HOBO data loggers recorded no periods of drought. The environmental conditions at point IZ90JJ are similar to those at KR800. However, at this location the d13C value is more negative than
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Fig. 5. Correlation between relative humidity (RH) and carbon stable isotope composition (d13C) of cellulose extracted as nitrocellulose from mosses (Polytrichum and Sphagnum) from the Izerskie Mts. and Karkonosze Mts., SW Poland. The temperatures are average values calculated from 1 h intervals measured for a period starting at the beginning of the growing season (April 2004) and ending with sample collection (July or October 2004). Points (open symbols) IZ90JJ (graphs a–d), KR1000 (graphs e–g), and KR900 (graph h) are excluded from calculations of R2 and linear regression analyses.
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expected (for Sphagnum and Polytrichum, both periods). This can be explained by the snow cover being retained for a significantly longer time at this location than at the others. Similar effects may occur when most of the assimilated CO2 is of soil origin (d13C is usually much more negative than atmospheric CO2; Szaran et al., 2005). Two of these points (IZ90JJ and KR800) are also outliers from the regression line T–RH, which may confirm unusual conditions at those locations (Fig. 2). No simple explanation of these outliers can be offered at this point. 4.2. d13C/relative humidity relationship The strength of the relationship between d13C value and relative humidity was different for the Izerskie and Karkonosze Mts (Fig. 5). The correlation for the Izerskie Mts. is almost as strong as the correlation between temperature and d13C, with R2 ranging from 0.79 to 0.64 when point IZ90JJ was excluded. The correlation for the Karkonosze Mts. was weak, and the highest values of R2 (0.23–0.38) were observed when points KR1000 (Fig. 5 a–c) and KR900 (Fig. 5d) were excluded. Despite these weak correlations, the slope of the regression relationships for both species is relatively consistent at 0.30&/% (s.d. 0.07) and 0.35&/% (s.d. 0.07) (Table 2). In our opinion, the RH–d13C relationship is inconclusive because of the very high correlation between relative humidity and temperature (Fig. 2). In the analyzed system d13C–RH–T, the strongest correlations were observed for T–RH and T–d13C. Indeed, it was not surprising that correlation of d13C–RH was also observed. Overcast conditions and rain reduce insolation, causing a rapid decrease in temperature but an increase air humidity. This may explain the good T–RH correlation. Along the transects, humidity (measured about 1.5 m above ground) was high and varied only slightly from about 75% to 89% (at one location 67%) during the first season and from 72% to 90% during the second season. At ground level, higher and more stable RH may be expected due to soil/ peat evaporation. Based on high air RH, we could assume that no water stress conditions occurred during the entire growth season. A certain minimal humidity is necessary to maintain the biological processes of mosses; extreme drought may cause a temporary stop in the assimilation. Therefore, this
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period would not be recorded in the isotopic signature via the d13C value of plant tissue because no tissue would be formed. A freezing period could also temporarily stop the assimilation process. On the other hand, Bryophytes have developed the ability to be poikilohydric plants. That means that they take up water over the whole surface of the shoots from various sources. Moreover, a characteristic feature of Bryophytes is desiccation tolerance. In practice, most mosses carry substantial amounts of external water, which can vary widely without affecting the water status of the cells (Proctor, 2000). Sphagnum can keep the capitula wet by capillary water transport and maintain a high rate of photosynthesis (Rydin and Jeglum, 2006). Polytrichum stem possesses, in addition, a well developed internal water conducting system, which transfers water from the base to the actively photosynthesizing leaves at the apex (Proctor, 1982, 2000). Shoots of Polytrichum commune showed ability to photosynthesize in a wide range (55–90%) of air relative humidity (Penny and Bayfield, 1982). Results of other studies suggest that humidity and d13C are related by about 0.01 to 0.2&/% for vascular plants (Menot and Burns, 2001). Non-vascular plants were not studied in detail. Humidity can control the d13C of plants mainly due to the stomata isotope fractionation effect, but this does not occur in the case of gametophyte mosses. Therefore, due to their lack of stomata, mosses should not be directly influenced by humidity. 5. Conclusions The computed Fq factors illustrate that the 1.6& and 1.5& d13C decrease for Sphagnum and Polytrichum, respectively, corresponds to a 1 °C increase in air temperature in the growing season. Air temperature in the growing season is probably the major factor controlling the stable carbon isotopic composition of peat-forming plant tissue. However, variation in other local environmental parameters may play a minor role, which, in the case of bogs, can be negligible because of the usually high water level and high RH that are required to sustain bog forming plant vegetation. Thus, under conditions encountered along the altitude transects we considered, changes in relative humidity seem to have had minor influence on carbon isotopic fractionation. Observed correlations of d13C and RH are the consequence of high correlation for T–RH and T–d13C.
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These results are significant for the re-analysis of peat core profiles in studies that investigate variation in palaeotemperature, especially given the good preservation of the original isotopic signatures of peat-forming plants (Skrzypek and Je˛drysek, 2005). Acknowledgements The authors are grateful to Dr. J.K. Haschenburger and Dr. S. Birnbaum for their critical reading of the text and valuable remarks, S. Bruder for text corrections and E. Fisher and an anonymous reviewer for constructive reviews. The study was realized as scientific Grant No. 2P04G 004 26 (KBN), supported from 2022/W/ING/05 and 1017/S/ING/05-IX funds. Associate Editor—I.D. Bull
References Aucour, A.M., Hillaire-Marcel, C., Bonnefille, R., 1994. Late Quaternary biomass changes from 13C measurements in a highland peat bog from Equatorial Africa (Burundi). Ternary Research 41, 225–233. Barber, K.E., 1981. Peat Stratigraphy and Climatic Change. A.A. Balkema, Rotterdam. Barber, K.E., Chambers, F.M., Maddy, D., 2004b. Late Holocene climatic history of northern Germany and Denmark: peat macrofossil investigations at Dosenmoor, SchleswigHolstein, and Svanemose, Jutland. Boreas 33, 132–144. Barber, K., Zolitschka, B., Tarasov, P., Lotter, A.F., 2004a. Atlantic to Urals – the Holocene climatic record of MidLatitude Europe. In: Past Climate Variability through Europe and Africa. Springer, Dordrecht, pp. 417–442. Boutton, T.W., Archer, S.R., Midwood, A.J., Zitzer, S.F., Bol, R., 1998. d13C values of soil organic carbon and their use in documenting vegetation change in a subtropical savanna ecosystem. Geoderma 82, 5–42. Dawson, T.E., Mambelli, S., Plamboeck, A.H., Templer, P.H., Tu, K.P., 2002. Stable isotopes in plant ecology. Annual Review of Ecology Systematics 33, 507–559. Epstein, S., Yapp, C.J., Hall, J.H., 1976. Determination of the D/ H ratios of nonexchangeable hydrogen in cellulose extracted from aquatic and land plants. Earth and Planetary Science Letters 30, 241–251. Farquhar, G.D., Ehleringer, J.R., Hubic, K.T., 1989. Carbon isotope discrimination and photosynthesis. Annual Review of Plant Physiology and Plant Molecular Biology 40, 503– 537. Francey, R.J., Farquhar, G.D., 1982. An explanation of 13C/12C variations in tree rings. Nature 297, 28–31. Griensted, M.J., Wilson, A.T., Ferguson, C.W., 1979. 13C/12C ratio variations in Pinus longaeva (Bristelone pine) cellulose during the last Millennium. Earth and Planetary Sciences Letters 42, 251–253. Je˛drysek, M.O., Krapiec, M., Skrzypek, G., Kału_zny, A., 2003. Air-pollution effect and paleotemperature scale versus d13C
records in tree rings and in a peat core (Southern Poland). Water, Air and Soil Pollution 145, 359–375. Je˛drysek, M.O., Skrzypek, G., 2005. Hydrogen, carbon and sulphur isotope ratios in peat: the role of diagenesis and water regimes in reconstruction of past climates. Environmental Chemistry Letters 2, 179–183. Ko¨rner, Ch., Farquhar, G.D., Roksandic, Z., 1988. A global survey of carbon isotope discrimination in plants from high altitude. Oecologia (Berlin) 74, 623–632. Ko¨rner, Ch., Farquhar, G.D., Wong, S.C., 1991. Carbon isotope discrimination by plants follows latitudinal and altitudinal trends. Oecologia (Berlin) 88, 30–40. Lajtha, K., Marshal, J.D., 1994. Sources of variation in the stable isotopic composition of plants. In: Lajtha, K., Michener, R.H. (Eds.), Stable Isotopes in Ecology and Environmental Sciences. Blackwell Scientific Publication, Oxford, pp. 1–21. Lamb, H.H., 1985. Climate, History and the Modern World. Menthuen, London. Langdon, P.G., Barber, K.E., 2005. The climate of Scotland over the last 5000 years inferred from multi-proxy peatland records: inter-site correlations and regional variability. Journal of Quaternary Science 20, 546–566. Leavitt, S.W., Long, A., 1986. Stable-carbon isotope variability in tree foliage and wood. Ecology 67, 1002–1010. Lipp, J., Trimborn, P., Fritz, P., Moser, H., Becker, B., Frenzel, B., 1991. Stable isotopes in tree ring cellulose and climatic change. Tellus 44B, 322–330. Menot, G., Burns, S.J., 2001. Carbon isotopes in ombrogenic peat bog plants as climatic indicators: calibration from an altitudinal transect in Switzerland. Organic Geochemistry 32, 233–245. O’Leary, M.H., 1981. Carbon isotope fractionation in plants. Phytochemistry 20, 553–567. O’Leary, M.H., Treichel, I., Rooney, M., 1986. Short-term measurement of carbon isotope fractionation in plants. Plant Physiology 80, 578–582. Park, R., Epstein, S., 1960. Carbon isotope fractionation during photosynthesis. Geochimica Cosmochimica Acta 21, 110–126. Penny, M.G., Bayfield, N.G., 1982. Photosynthesis in desiccated shoots of Polytrichum. New Phytologist 91, 637–645. Proctor, M.C.F., 1982. Physiological ecology: water relations, light and temperature responses carbon balance. In: Smith, A.J.E. (Ed.), Bryophyte Ecology. Chapman & Hall, London, pp. 333–381. Proctor, M.C.F., 2000. Physiological ecology. In: Shaw, A.J., Goffinet, B. (Eds.), Bryophyte Biology. Cambridge University Press, pp. 225–247. Robertson, I., Rolfe, J., Switsur, V.R., Carter, A.H.C., Hall, M.A., Barker, A.C., Waterhouse, J.S., 1997. Signal strength and climate relationships in 13C/12C ratios of tree ring cellulose from oak in southwest Finland. Geophysical Research Letters 24, 1487–1490. Rousseeuw, P.J., Leroy, A.M., 2003. Robust Regression and Outlier Detection. John Wiley and Sons Inc., Hoboken. Rydin, H., Jeglum, J., 2006. The Biology of Peatlands. Oxford University Press, p. 343. Skrzypek, G., Je˛drysek, M.O., 2001. Conservation of organic matter in peat: d13C and dD in peat profiles from ‘‘Suche Bagno’’ (NE Poland). Mineralogical Society of Poland, Special Papers 18, pp. 195–198. Skrzypek, G., Kału_zny, A., Je˛drysek, M.O., 2007. Carbon stable isotope analyses of mosses – comparisons of bulk organic
G. Skrzypek et al. / Organic Geochemistry 38 (2007) 1770–1781 matter and extracted nitrocellulose. Journal of the American Society for Mass Spectrometry, in press. Skrzypek, G., Je˛drysek, M.O., 2005. 13C/12C ratio in peat cores: record of past climates. In: Lichtfouse, E., Schwarzbauer, J., Robert, D. (Eds.), Environmental Chemistry – Green Chemistry and Pollutants in Ecosystems. Springer, Berlin, pp. 65–73. Smith, B.N., Herath, H.M., Chase, J.B., 1973. Effect of growth temperature on carbon isotopic ratios in barley, pea and rape. Plant and Cell Physiology 14, 177–182. Szaran, J., Dudziak, A., Trembaczowski, A., Niezgoda, H., Halas, S., 2005. Diurnal variations and vertical distribution of
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d13C, and concentration of atmospheric and soil CO2 in a meadow site, SE Poland. Geological Quarterly 49, 135–144. Troughton, J.H., Card, K.A., 1975. Temperature effects on the carbon-isotope ratio of C3, C4 and crasssulacean-acid metabolism (CAM) plants. Planta 123, 185–190. White, J.W.C., Ciais, P., Figge, R.A., Kenny, R., Markgraf, A., 1994. A high-resolution record of atmospheric CO2 content from carbon isotopes in peat. Nature 367, 153– 156. Wojtun´, B., 2006. Peat Mosses (Sphagnaceae) in Mires of the Sudetes Mountains (SW Poland): A Floristic and Ecological Study. Agricultural University of Wrocław, p. 225.
Organic Geochemistry Organic Geochemistry 38 (2007) 1770–1781 www.elsevier.com/locate/orggeochem
The carbon stable isotopic composition of mosses: A record of temperature variation Grzegorz Skrzypek
a,b,*
, Adam Kału_zny b, Bronisław Wojtun´ c, Mariusz-Orion Je˛drysek b
a
b
Laboratory for Stable Isotope Geochemistry, Department of Earth and Environmental Science, The University of Texas at San Antonio, One UTSA Circle, San Antonio, 78249 TX, USA Laboratory of Isotope Geology and Geoecology, Institute of Geological Sciences, The University of Wrocław, Cybulskiego 30, 50-205 Wrocław, Poland c The Wrocław University of Environmental and Life Sciences, Department of Botany and Plant Ecology, pl. Grunwaldzki 24a, 50-363 Wrocław, Poland Received 8 October 2006; received in revised form 8 February 2007; accepted 1 May 2007 Available online 5 June 2007
Abstract Samples of two moss species, Sphagnum and Polytrichum, were collected at 19 points selected along vertical transects in the Karkonosze Mts. (739–1393 m) and the Izerskie Mts. (500–1100 m), SW Poland. Continuous automatic monitoring of temperature and humidity at the sampling points was carried out (1 h intervals) through the entire growth season (2004). A strong correlation between d13C value of plant tissue cellulose and mean air temperature was noted for both areas and sampling periods (R2 between 0.92 and 0.64). It was calculated that a 1 °C increase in air temperature during the growing season results in a 1.6& (Sphagnum) and a 1.5& (Polytrichum) decrease in d13C. In contrast, it seems that humidity shows no clear influence on the carbon isotope composition of mosses. Ó 2007 Elsevier Ltd. All rights reserved.
1. Introduction Climatic variations during the last millennium are relatively well documented (e.g. Barber, 1981; Lamb, 1985; Lipp et al., 1991; White et al., 1994; Je˛drysek et al., 2003; Barber et al., 2004a,b; Lang* Corresponding author. Present address: Laboratory for Stable Isotope Geochemistry, Department of Earth and Environmental Science, The University of Texas at San Antonio, One UTSA Circle, San Antonio, 78249 TX, USA. Tel.: +1 210 7248430; fax: +1 210 4584469. E-mail addresses: [email protected], [email protected]. wroc.pl (G. Skrzypek).
don and Barber, 2005). However, the role of our industrial civilization, as well as the scale of the changes, is still not clear. Two major questions are: ‘‘Which aspects of reconstructed past climates allow us to better understand present climatic variation?’’ and ‘‘Is it possible to quantitatively predict future climatic trends?’’ The stable isotope composition of organic matter from peat profiles may potentially bring crucial information to answer these questions (e.g. O’Leary et al., 1986; Francey and Farquhar, 1982; Farquhar et al., 1989; Aucour et al., 1994; Boutton et al., 1998; Dawson et al., 2002). Although evidence suggests that temperature
0146-6380/$ - see front matter Ó 2007 Elsevier Ltd. All rights reserved. doi:10.1016/j.orggeochem.2007.05.002
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is the dominant (primary) factor controlling d13C values in peat (Skrzypek and Je˛drysek, 2005), there is no well documented quantitative calibration defining the temperature control on the carbon isotope composition (d13C) in peat-forming plants (Skrzypek and Je˛drysek, 2001, 2005). The role of secondary factors can be so important that the primary signal can be overwhelmed. For certain habitats it is possible to exclude most of these secondary factors by properly selecting sampling points, both spatially and temporally, e.g. where and when the humidity, insolation and soil conditions are very similar, thereby permitting the exclusion of these factors. Moreover, the gametophytes of mosses do not have stomata, so excluding the so called ‘‘stomata effect’’ that is important in the case of carbon stable isotope fractionation during CO2 assimilation by higher plants (the scale of the effect depends on the humidity variation). Although temperature has an extremely important influence on fractionation, some authors have shown that precipitation (humidity) and atmospheric pollution, rather than temperature, are the most important factors controlling d13C values in vascular plants (e.g. Je˛drysek et al., 2003). Likewise, Ko¨rner et al. (1991, 1988) and Menot and Burns (2001) suggest the crucial role of altitude (atmospheric pressure and CO2 atmospheric partial pressure). However, the carbon isotopic composition of plant tissue is not a direct effect of atmospheric CO2 – plant tissue isotope fractionation, but rather results from complex processes controlled by environmental conditions during metabolism (e.g. O’Leary et al., 1986; Farquhar et al., 1989). The quantitative role of these factors in controlling isotopic fractionation is proposed in the equation defined by Farquhar et al. (1989). During plant growth, isotopic fractionation generally proceeds in two stages (Park and Epstein, 1960; O’Leary, 1981, 1986; Farquhar et al., 1989): (1) adsorption and diffusion of CO2 into the plant tissue and (2) the initial carboxylation. Plant growth results in an 18–27& (C3 plants) and 4–6& (C4 plants) 12C enrichment in the plant tissue with respect to atmospheric CO2 (O’Leary, 1981, 1986; Lajtha and Marshal, 1994), with the value dependent on the plant species and temperature during CO2 assimilation and carboxylation. Many authors, on the basis of various plant studies, have proposed significantly different Fq values, defined as the change in the d13C value of plant tissue resulting from a 1 °C change in temperature
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(Skrzypek and Je˛drysek, 2005), varying from 1 to 2.4&/°C (Smith et al., 1973; Troughton and Card, 1975; Griensted et al., 1979; O’Leary, 1981; Leavitt and Long, 1986; Lipp et al., 1991; Robertson et al., 1997). According to calibrations based on peat (Skrzypek and Je˛drysek, 2005) the Fq value is 0.6&/°C. Likewise, Menot and Burns (2001) found that, in peat-forming Sphagnum, Fq varies from 0.2 to 0.4&/°C, but the temperatures were not measured exactly at the sampling places but at meteorological stations in few kilometers distance from sampling points, at the respective altitude. Therefore, it is difficult to precisely reconstruct temperature variations based on the d13C analysis of peat profiles. To address this problem, we analyzed the d13C values for mosses, peat-forming Sphagnum and Polytrichum and continuously monitored both the temperature and humidity at the specific sampling points where the samples were collected twice during the growth season. 2. Goal and strategy The major goal was to describe the influence of temperature on the d13C value of moss tissue, the direct result of which is calculation of Fq factors [change in d13C value during a 1 °C increase in temperature (&/°C)]. The potential influence of other factors, such as humidity and habitat condition, was also analyzed. It was therefore necessary to find natural locations of mosses growing in similar habitats but with different temperature ranges during the growing season. These criteria are attained on relatively steep mountain slopes where a significantly different altitude of sampling points (e.g. 100 m) can be utilized, with little difference in horizontal distance (only a few km). Usually, an increase in altitude of 100 m results in a decrease in air temperature of ca. 0.6 °C. 3. Sampling points and methods Samples of two moss species, Sphagnum girgensohnii Russow and Polytrichum commune Hedw., were collected from northern hillsides in the Karkonosze Mts. (vertical transect 739–1393 m) and Izerskie Mts. (500–1101 m), SW Poland (Fig. 1). Details about locations of the 19 sampling/observation points are shown in Table 1 (RH = relative humidity). The two species were selected because of their common occurrence in the selected regions. Furthermore, their ecological preferences are only
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Fig. 1. Location of sampling points in two areas (SW Poland); the Izerskie Mts. are located about 30 km west of the Karkonosze Mts. The mountain slopes (IZ500–IZ11KO and KR600–KR1400) face north. The short vertical transect IZ11KO–IZ84HA is located at the southwest facing slope. Point KR700 is excluded because of data logging defects.
Table 1 d13C values of moss samples and meteorological data recorded (HOBO loggers) at the place of collection Location
Name
Altitude (m)
Average for season #1 (4 July 2004) T (°C)
RH (%)
d13C [&,VPDB], Poly
d13C [&,VPDB], Sph
Average for season #2 (4 October 2004) T (°C)
RH (%)
d13C [&,VPDB], Poly
d13C [&,VPDB], Sph
Izerskie Mts.
IZ84HA IZ85HA IZ90JJ IZ10JJ IZ11KOP IZ10SI IZ90SI IZ80SI IZ700 IZ600 IZ500
843 857 902 998 1101 1002 899 799 695 597 500
9.7a 8.7 8.1 8.0 7.7 8.5 9.1 9.6 9.7 10.1 11.1
77 81 89 83 85 80 82 80 76 76 67
26.31b 26.75 27.55 24.86 25.02 24.88 25.55 26.12 26.23 28.06 28.95
28.50c 28.03 30.56 26.21 25.03 27.51 28.62 26.00 28.73 29.67 30.94
10.8a 9.7 9.3 9.2 9.1 9.9 10.2 10.7 10.8 11.2 12.0
78 82 90 84 85 81 83 82 78 78 72
25.80b 25.82 27.18 24.81 24.79 24.63 25.90 25.89 28.02 28.45 30.07
29.36c n/a 31.12 26.60 25.93 26.96 29.14 28.31 30.42 30.10 30.34
Karkonosze Mts.
KR1400 KR1300 KR1200 KR1100 KR1000 KR900 KR800 KR700 KR600
1393 1266 1240 1091 1052 914 834 675 602
6.5 6.8 7.3 8.1 9.0 9.2 9.4 n/ad 9.7
85 85 82 82 79 81 83 n/ad 75
24.60 25.02 25.45 28.53 25.14 28.96 25.96 26.43 27.44
27.52 26.43 26.19 30.04 27.83 31.58 31.30 26.78 31.86
7.9 8.0 8.7 9.2 10.1 10.4 10.7 n/ad 10.9
85 85 80 81 81 82 84 n/ad 76
24.45 24.11 24.74 28.56 24.72 30.60 26.27 26.75 27.55
27.52 26.86 26.58 31.57 27.45 30.35 30.86 26.68 31.83
a
The temperatures are average values calculated from 1 h intervals measured for a period starting at the beginning of the growing season (April 15–19) and ending with sample collection (July 2–4 and October 15–17, 2004) for short growth ‘‘Season #1’’ and long growth ‘‘Season #2’’, respectively. b Polytrichum commune. c Sphagnum girgensohnii. d n/a – lost sample (Sph IZ85HA), damaged sensor (KR700).
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slightly different the anatomy and ecophysiology of S. girgensohnii is similar to that of other peat-forming species of the genus Sphagnum in ombrotrophic bogs or poor fens. Although in the Western Sudetes S. girgensohnii and P. commune are shade-tolerant and have similar trophic amplitude, the former prefers wet microhabitats, while the latter is typically found at dryer sites (Wojtun´, 2006). Special attention was paid to the selection of sampling points where environmental conditions were, to the furthest extent possible, similar, especially the presence of a peat layer, relatively stable soil moisture during growing season and similarity in the surrounding ecosystem. Despite these assumptions, local conditions varied according to altitude and differences in forest stand and condition. Similarity in conditions amongst the sampling sites is suggested by the close proximity of the sample locations. The furthest distance between sampling points in each transect was < 5.7 km (Izerskie Mts.) and < 4.9 km (Karkonosze Mts.; Fig. 1). However, the local microclimate may differ slightly at each location (e.g. concentration of CO2 and d13C in air). Based upon dominant vegetation type, observation points could generally be classified into three major groups: 1. A sub-alpine dwarf pine group dominated by Pinetum mugo sudeticum; this group includes only KR1400. 2. A mire group situated in poor fens. This group includes three sampling points: KR1300, IZ84HA and IZ85HA. 3. A forest group situated inside a spruce forest. This group may be further divided into two sub-groups: the first includes points KR1100, KR1200, IZ90SI, IZ90JJ, IZ10SI, IZ10JJ and IZ11KO, located in the upper forest belt, and the second points KR600, KR800, KR900, KR1000, IZ500, IZ600, IZ700 and IZ80SI in the lower forest belt. Within the forest group, points KR800 and KR1000 were different as they are located in local clearings. Two points in time define the duration of the growing season for all specimens: the normal weather-controlled start of growth and the termination of growth by sample collection. As sample collection was carried out at the same time for all samples, the variable for the length of the growing season is the elevation-controlled start of plant growth. The start of the growing season for each
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location was defined as that period when the average temperature of five consecutive days reached 5 °C. All samples were collected during two field seasons in 2004, one during July 2–4, and the other during October 15–17, resulting in a short and long growth season for comparison (season 1st July and 2nd October). Throughout the entire 2004 growing season, for all sample locations, automated measurements of temperature and humidity were collected at 60 min intervals using a U8 HOBO data logger (located about 1.5–2.0 m above ground and not further than 5 m from plant sampling points). The length of the growing season varied for different points depending on altitude, but the variation was < 15 days (start date between April 15 and 29). In some places, the real growth period can be shorter, because of longer snow cover. After botanical identification, the whole batches of individual moss species were washed, dried under vacuum and homogenized (2–3 g dry matter). The cellulose was extracted following the modified technique developed by Epstein et al. (1976). This procedure consisted of three major steps (e.g. Skrzypek et al., 2007): (1) multiple chemical treatments: soak in 1:1 C6H6–MeOH (24 h), rinse with HCl (4%), boil with water (1 h), heat (70 °C) with CH3CO2H and NaCl, soak in 17% NaOH (1 h) and CH3CO2H for 10 min; (2) nitrification: dry samples were nitrated using 100% HNO3 (48 h); (3) dissolution and precipitation of nitrocellulose: dissolution in acetone and, after centrifugation and decantation, the nitrocellulose was precipitated from the solution by quickly adding redistilled water. After each chemical treatment, samples were washed with redistilled water and after each step were vacuum dried. Stable isotope analysis was carried out using an off-line preparation system technique for maximum precision. About 3–5 mg of pure nitrocellulose obtained from the plant materials were combusted with CuO wire in a sealed quartz tube under vacuum at 900 °C (Je˛drysek and Skrzypek, 2005). The CO2 produced was cryogenically purified off-line and introduced into an isotope ratio mass spectrometer (IRMS Finnigan-Mat Delta E/dual inlet) for stable carbon isotope ratio analysis. The d13C values were normalized against NBS22 and USGS24 international standards and are reported relative to the Vienna Pee Dee Belemnite (VPDB) scale, with ± 0.05& analytical precision. The d13CVPDB value is defined as the relative difference, in parts per thousand (&), between the isotope ratio of the sample and the international standard (value determined
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by International Atomic Energy Authority, IAEA, Vienna). Least squares regression analysis was performed with d13C as the response variable and RH or T as the predictor variable. The relationship was described with a simple linear model, y = ax + b, where x = d13C and y = T or RH, respectively. Traditional statistical tests assume that the relationship in question is true and uses data to detect differences. Because we wished to use our data to look for similarities, we also calculated probability (P) associated with a Student’s paired t-test, with a two-tailed distribution. 4. Results and discussion The d13C values of nitrocellulose extracted from moss tissue varied from 31.86 to 25.03& for Sphagnum (Sph) and from 30.60 to 24.11& for Polytrichum (Poly) (Table 1). No significant differences were observed between the same moss species
collected in the Izerskie and Karkonosze Mts. (P-values range from 0.3 to 0.6, paired t-test, twotailed). However, it was observed that the d13C values for Polytrichum were more positive than those for Sphagnum. The average difference observed was about 2.21&, but varied over a wide range, depending on location and sampling season (s.d. 1.36&). The mean air temperature (T) during long growth season (#2) varied according to altitude as follows: from 9.1 to 12.0 °C for the Izerskie (altitude 500–1101 m) and 7.9 to 10.9 °C for the Karkonosze Mts. (602–1394 m). The relative humidity (RH) varied within a narrow range: for the Izerskie Mts. 72% to 90% (mean 82%) and for the Karkonosze 76–85% (mean 82%; Table 1). Very high and relatively constant RH at all sampling points was due to frequent rain (total mean annual precipitation in these regions is 1400 mm/year). A very high negative correlation exists between the temperature and humidity of the air overall (R2 varies from 0.86 to 0.63), which is interpreted to be the
Fig. 2. Correlation between temperature and humidity (T–RH) observed in two regions the Karkonosze Mts. and Izerskie Mts. The outliers excluded include IZ90JJ for the linear regression analyses (Izery, graphs a and b) and KR800 (Karkonosze, c and d).
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effect of cloud cover and the resultant lower insolation (Fig. 2). However, the observed humidity (measured 1.5 m above ground) is likely to be lower than the humidity directly at ground level where the moss layers occurred. A limited number of previous studies have investigated the relationship between plant carbon stable isotope composition and altitude (Ko¨rner et al., 1988, 1991; Menot and Burns, 2001). In general, d13C changed with altitude for three Sphagnum species (n = 10, 8, 6), observed by Menot and Burns (2001), but correlations were weak (R2 = 0.25, 0.48, 0.17). No clear correlation with air temperature or humidity was reported. In our study, variability in d13C value with respect to temperature and humidity of air was analyzed separately for species, location, and season. 4.1. d13C/temperature relationship A strong correlation between d13C value of moss tissue cellulose and temperature exists for both areas and sampling periods (Fig. 3). The linear regression equations and the R2 coefficients were calculated for the respective data pairs T (°C) and d13C (&). R2 ranged from 0.76 to 0.92 for the Izerskie Mts. and from 0.64 to 0.87 for the Karkonosze Mts. (Table 2). The relationship between d13C value and temperature was described with a simple linear regression model T = a(d13C) + b, where slope (a) represents Fq value (&/°C). The calculated Fq value (slope) differed slightly for each moss genus depending on location and season [Sphagnum: 1.78 to 1.56&/°C (s.d. 0.12); Polytrichum: 1.75 to 1.15&/°C (s.d. 0.31)]. The average value of d13C decreases approximately 1.62& for Sphagnum and 1.46& for Polytrichum for each 1 °C increase in air temperature during the growing season, which corresponds to a +1& increase for each 0.62 °C and 0.72 °C decrease, respectively. The Fq value did not vary significantly, and no significant differences between d13C of moss species collected during different sampling seasons were found (P-value from 0.06 to 0.95, unpaired t-test, two-tailed). This observation was confirmed for both moss genera by the high correlation between d13C value for moss pairs collected during July and October; R2 varies between 0.80 and 0.95 (Fig. 4). These high R2 values suggest that the same mechanism defines the carbon stable isotope fractionation for both seasons and moss species. How-
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ever, in the case of Polytrichum, some difference was observed between seasons. For the first season, Fq was 1.15 and 1.20&/°C and for the second season, 1.75 and 1.68&/°C for Polytrichum and Sphagnum, respectively. Different averages of air temperature occurred during shorter (#1) and longer (#2) seasons; therefore the d13C of plant tissue differs, respectively. The d13C value of moss at a place where the temperature of vegetation season was lower (e.g. at higher altitude) was more positive than where the temperature was higher. This difference, defined as Fq (&/°C), and calculated for each species, varied slightly depending on season, location and environmental conditions. The variation in Fq value was narrower for Sphagnum (s.d. 0.12) than Polytrichum (0.31; Table 2). The Fq standard deviation can be considered as an error in the calculations, as a result of the minor influence of local parameters as well as differences between the field-recorded temperature and actual temperature during photosynthesis. Thus, the average value calculated for each species (for two seasons and two locations) can be used as the most reliable. On the other hand, we observed only slight differences between the species (Fq 1.6 and 1.5&/°C), these probably being due to small differences in physiology and environmental preference. Data for two sampling points in Karkonosze, KR1000 (1052, both species and periods) and KR800 (834 m, Polytrichum only, both periods) and two points from Izerskie IZ90JJ (902 m, both species and periods) and IZ80SI (799 m, Sphagnum, first period only) do not match the general linear regressions trend for T–d13C (Fig. 3). These points were excluded from the calculation of linear regression equations and R2 (open symbols in all figures), based on multiple point diagnostics of outliers (e.g. Rousseeuw and Leroy, 2003). The points, which do not match the general T–d13C trend (Fig. 3), lie on the regression line for d13C–d13C (July–October; Fig. 4a–d). This fact suggests that the offset in these four points is not a random variation and may result from the specific location relative to local conditions. However, no specific conditions have been observed that can explain the inconsistency. Points KR1000 and IZ80SI are located in clearings within local spring mires, whereas IZ90JJ and KR800 are located in spruce forest where the surrounding floor is covered with spruce litter. The d13C values of plants collected at IZ80SI, KR800 and KR1000 are significantly more positive than one would
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Fig. 3. Correlation between temperature (T) and stable carbon isotope composition (d13C) of cellulose extracted as nitrocellulose from mosses (Polytrichum and Sphagnum) from the Izerskie Mts. and Karkonosze Mts., SW Poland. The temperatures are average values calculated from 1 h intervals measured for a period starting at the beginning of the growing season (April 2004) and ending with sample collection (July or October 2004). Points IZ90JJ (graphs a–d), IZ80SI (a) KR1000 (e–h), and KR800 (g and h) are excluded from calculations of R2 and linear regression analyses (open symbols).
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Table 2 Regression analysisa of temperature (T, °C) and relative humidity (RH, %) changes in carbon stable isotope discrimination (d13C, &) Species
Location/sampling period
n
T = a(d13C) + b a
RH = a(d13C) + b b
R
2
a
R2
b
Polytrichum
Izerskie/1 Izerskie/2 Karkonosze/1 Karkonosze/2 Average (s.d.)
10 10 6 6
1.15 1.75 1.20 1.68 1.45 (0.31)
15.64 8.22 17.10 11.25
0.76 0.77 0.68 0.64
0.23 0.40 0.30 0.26 0.30 (0.07)
44.41 58.39 51.00 47.36
0.73 0.77 0.33 0.27
Sphagnum
Izerskie/1 Izerskie/2 Karkonosze/1 Karkonosze/2 Average (s.d.)
9 9 7 7
1.56 1.64 1.78 1.49 1.62 (0.12)
13.79 11.46 14.74 15.35
0.92 0.80 0.87 0.69
0.29 0.33 0.46 0.33 0.35 (0.07)
51.26 54.70 67.26 56.27
0.79 0.64 0.38 0.23
a The relation was described with a simple linear model y = ax + b, where x = d13C and y = T or RH, respectively. The value ‘‘a’’ in the linear equation represents Fq factor d13C/T (&/°C) or d13C/RH (&/%).
Fig. 4. Correlation of d13CJuly–d13COctober pairs for Sphagnum and Polytrichum collected at each sampling point during July and October seasons.
expect on the basis of the general trend. One possible explanation, for an effect like this, may lie in the observation that samples IZ80SI and KR1000 are located in the area surrounded by watered mires and, as a result, would not have been exposed to conditions of drought, which could cause a tempo-
rary pause in assimilation and a shift in d13C value (in less watered locations). However, this is speculative as the HOBO data loggers recorded no periods of drought. The environmental conditions at point IZ90JJ are similar to those at KR800. However, at this location the d13C value is more negative than
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Fig. 5. Correlation between relative humidity (RH) and carbon stable isotope composition (d13C) of cellulose extracted as nitrocellulose from mosses (Polytrichum and Sphagnum) from the Izerskie Mts. and Karkonosze Mts., SW Poland. The temperatures are average values calculated from 1 h intervals measured for a period starting at the beginning of the growing season (April 2004) and ending with sample collection (July or October 2004). Points (open symbols) IZ90JJ (graphs a–d), KR1000 (graphs e–g), and KR900 (graph h) are excluded from calculations of R2 and linear regression analyses.
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expected (for Sphagnum and Polytrichum, both periods). This can be explained by the snow cover being retained for a significantly longer time at this location than at the others. Similar effects may occur when most of the assimilated CO2 is of soil origin (d13C is usually much more negative than atmospheric CO2; Szaran et al., 2005). Two of these points (IZ90JJ and KR800) are also outliers from the regression line T–RH, which may confirm unusual conditions at those locations (Fig. 2). No simple explanation of these outliers can be offered at this point. 4.2. d13C/relative humidity relationship The strength of the relationship between d13C value and relative humidity was different for the Izerskie and Karkonosze Mts (Fig. 5). The correlation for the Izerskie Mts. is almost as strong as the correlation between temperature and d13C, with R2 ranging from 0.79 to 0.64 when point IZ90JJ was excluded. The correlation for the Karkonosze Mts. was weak, and the highest values of R2 (0.23–0.38) were observed when points KR1000 (Fig. 5 a–c) and KR900 (Fig. 5d) were excluded. Despite these weak correlations, the slope of the regression relationships for both species is relatively consistent at 0.30&/% (s.d. 0.07) and 0.35&/% (s.d. 0.07) (Table 2). In our opinion, the RH–d13C relationship is inconclusive because of the very high correlation between relative humidity and temperature (Fig. 2). In the analyzed system d13C–RH–T, the strongest correlations were observed for T–RH and T–d13C. Indeed, it was not surprising that correlation of d13C–RH was also observed. Overcast conditions and rain reduce insolation, causing a rapid decrease in temperature but an increase air humidity. This may explain the good T–RH correlation. Along the transects, humidity (measured about 1.5 m above ground) was high and varied only slightly from about 75% to 89% (at one location 67%) during the first season and from 72% to 90% during the second season. At ground level, higher and more stable RH may be expected due to soil/ peat evaporation. Based on high air RH, we could assume that no water stress conditions occurred during the entire growth season. A certain minimal humidity is necessary to maintain the biological processes of mosses; extreme drought may cause a temporary stop in the assimilation. Therefore, this
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period would not be recorded in the isotopic signature via the d13C value of plant tissue because no tissue would be formed. A freezing period could also temporarily stop the assimilation process. On the other hand, Bryophytes have developed the ability to be poikilohydric plants. That means that they take up water over the whole surface of the shoots from various sources. Moreover, a characteristic feature of Bryophytes is desiccation tolerance. In practice, most mosses carry substantial amounts of external water, which can vary widely without affecting the water status of the cells (Proctor, 2000). Sphagnum can keep the capitula wet by capillary water transport and maintain a high rate of photosynthesis (Rydin and Jeglum, 2006). Polytrichum stem possesses, in addition, a well developed internal water conducting system, which transfers water from the base to the actively photosynthesizing leaves at the apex (Proctor, 1982, 2000). Shoots of Polytrichum commune showed ability to photosynthesize in a wide range (55–90%) of air relative humidity (Penny and Bayfield, 1982). Results of other studies suggest that humidity and d13C are related by about 0.01 to 0.2&/% for vascular plants (Menot and Burns, 2001). Non-vascular plants were not studied in detail. Humidity can control the d13C of plants mainly due to the stomata isotope fractionation effect, but this does not occur in the case of gametophyte mosses. Therefore, due to their lack of stomata, mosses should not be directly influenced by humidity. 5. Conclusions The computed Fq factors illustrate that the 1.6& and 1.5& d13C decrease for Sphagnum and Polytrichum, respectively, corresponds to a 1 °C increase in air temperature in the growing season. Air temperature in the growing season is probably the major factor controlling the stable carbon isotopic composition of peat-forming plant tissue. However, variation in other local environmental parameters may play a minor role, which, in the case of bogs, can be negligible because of the usually high water level and high RH that are required to sustain bog forming plant vegetation. Thus, under conditions encountered along the altitude transects we considered, changes in relative humidity seem to have had minor influence on carbon isotopic fractionation. Observed correlations of d13C and RH are the consequence of high correlation for T–RH and T–d13C.
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These results are significant for the re-analysis of peat core profiles in studies that investigate variation in palaeotemperature, especially given the good preservation of the original isotopic signatures of peat-forming plants (Skrzypek and Je˛drysek, 2005). Acknowledgements The authors are grateful to Dr. J.K. Haschenburger and Dr. S. Birnbaum for their critical reading of the text and valuable remarks, S. Bruder for text corrections and E. Fisher and an anonymous reviewer for constructive reviews. The study was realized as scientific Grant No. 2P04G 004 26 (KBN), supported from 2022/W/ING/05 and 1017/S/ING/05-IX funds. Associate Editor—I.D. Bull
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