IAWA Journal, Vol.

26 (1), 2005: 121–136

CARBON AND OXYGEN ISOTOPE DENDROCHRONOLOGY IN

SUB-FOSSIL BOG OAK TREE RINGS – A PRELIMINARY STUDY

U. Sass-Klaassen1,2, I. Poole 3,4, T. Wils 5, G. Helle 5, G.H. Schleser 6 &

P.F. van Bergen 3

SUMMARY

Isotope dendroclimatology is a relatively new field investigating en-

vironmental factors that control the radial growth of trees. Tree-ring
series of sub-fossil bog oaks can be dated from sites across northwest
Europe indicating that the environmental change(s) were regional rather
than local. Bog oaks show characteristic periods of suppressed growth
thought to have resulted from changes in the hydrological status of bogs
towards either wetter or drier conditions. This study investigates relative
changes in stable carbon (δ13 C) and oxygen (δ18 O) isotope content in
phases of suppressed and normal growth in three bog oaks dated as c.
200 BC to 150 AD from Zwolle, eastern Netherlands. Bog oaks show
no clear relationship between tree-ring width and isotopic composition
although one tree exhibited relatively depleted values of 13C and 18O with
suppressed growth. Suppressed ring growth is characterised by the forma-
tion of earlywood only, possibly as a result of hydrologic alterations that
limited the formation of latewood, which would otherwise have locked
up a detectable signal in stable isotopic shift.
Key words: Stable isotopes, carbon, oxygen, dendrochronology, sub-
fossil bog oak, palaeoclimate.

INTRODUCTION

ʻBog oaksʼ are sub-fossil oaks that have been preserved in peat environments for up to
many thousands of years. These oaks grew under sub-optimal site conditions either in the
vicinity of – or, during drier periods, directly on the peat itself. Within the Netherlands,
Germany and Ireland more than 200 such sites yield sub-fossil oak material, often in

1) Netherlands Centre for Dendrochronology, RING, PO Box 1600, 3800 BP Amersfoort, The
Netherlands.
2) Forest Ecology and Forest Management Group, Wageningen University, PO Box 342, 6700
AH Wageningen, The Netherlands [E-mail: ute.sassklaassen@wur.nl].
3) Geochemistry, Earth Sciences, Utrecht University, PO Box 80021, 3508 TA Utrecht, The Nether-
lands.
4) Palaeontological Museum, Oslo University, PO Box 1172 Blindern, N-0318 Oslo, Norway.
5) Physical Geography, Utrecht University, PO Box 80115, 3508 TC Utrecht, The Netherlands.
6) Research Centre Jülich GmbH, Institute Sedimentary Systems, ICG-V, D-52425 Jülich, Ger-
many.
Associate Editor: Thomas Yanosky
122 IAWA Journal, Vol. 26 (1), 2005

association with species characteristic of wetland woods such as alder (Alnus glutinosa),
birch (Betula pubescens), willow (Salix spp.) and ash (Fraxinus exelsior) (Kooistra
et al. 2005). The sites were characterised by high ground-water levels and, in many loca-
tions, recurrent inundation from nearby river systems (Leuschner 1992; Kooistra et al.
2005). Consequently, changes in hydrology had a major impact on the growth of the
trees and the population dynamics of these ancient woodlands (Leuschner et al. 2002;
Leuschner & Sass-Klaassen 2003; Sass-Klaassen & Hanraets 2005).
Tree-ring chronologies provide a unique palaeoenvironmental archive since den-
drochronological methods can precisely date rings to an annual resolution. Ultra-long
tree-ring chronologies from bog oaks across northwest Europe extend from 6069 BC to
the tenth century AD (Pilcher et al. 1984; Jansma 1996; Spurk et al. 1998; Leuschner
et al. 2002). These indicate that most of the oaks were slow growing with characteristic
alternating phases of normal and suppressed growth (Fig. 1). These chronologies enable
the analysis of changing environments, including climate and hydrology, and provide
a valuable base for climate reconstruction through the Holocene (e.g. Leuschner et al.
2002; Spurk et al. 2002; Mayr et al. 2003). Since bog oaks originally grew and sub-
sequently became preserved in wetland environments, it has been hypothesised that
variations in tree growth were caused predominantly by hydrological shifts in the
environment towards either wetter or drier conditions (Pilcher et al. 1996). Surplus
water during growing periods may result in lowered water potentials (Ewe & Sternberg
2002) and possible root damage (Kozlowski 1997; Crawford et al. 2003) resulting in
water-stress. This scenario has been assumed to have had a major impact on the growth
of bog oaks (Leuschner et al. 2002; Sass-Klaassen et al. 2004). Indeed, recent results
from an excavation of wood in Zwolle-Stadshagen (The Netherlands) suggest that epi-
sodes of high ground-water level and/or frequent inundations limited radial growth and
most likely provided the trigger for the characteristic long-term growth suppressions
observed in bog oaks (Kooistra et al. 2005; Sass-Klaassen & Hanraets 2005). However,
it has not been possible to use tree-ring patterns to determine the precise environmental
factors causing growth suppressions because there are no analogous sites with living
oaks in Europe today. Thus the assessment of the climate/hydrology-growth relation-
ship of oaks in wetland woods remains speculative (Sass-Klaassen 2004).
Isotope dendroclimatology is a rapidly expanding field of investigation focusing
on relative changes in stable isotope content of wood, namely carbon, oxygen1) and
hydrogen and has been applied to sub-fossil and fossil material (e.g. Frielingsdorf
1993; Lipp et al. 1996; Mayr et al. 2003). Changes in stable isotope compositions in
plant material may occur as a result of changes in climate and environmental condi-
tions, such as temperature, relative humidity, light intensity and soil hydrology (Saurer
& Siegenthaler 1989; Dupouey et al. 1993; Switsur et al. 1996; Saurer et al. 1997a;

1) The δ13C and δ18 O are defined as: (Rs / Rref - 1) × 1000 ‰ in which Rs is 13C/ 12 C and 18 O/16 O
in the wood sample and R ref is 13 C / 12 C and 18 O / 16 O of a reference standard. For carbon iso-
topes the reference standard is a fossil belemnite from the Pee Dee Formation (VPDB), Upper
Cretaceous, South Carolina, USA. For oxygen isotopes the reference standard is the Standard
Mean Ocean Water (VSMOW).
Sass-Klaassen et al. — Carbon and oxygen isotope dendrochronology 123

Fig. 1. Photomicrograph of the transverse surface of a typical bog oak from Zwolle, The Nether-
lands, showing alternating phases of normal and suppressed growth. Note that only earlywood is
present during periods of growth suppression. — Scale bar = 1 cm.

Switsur & Waterhouse 1998; Schleser et al. 1999b; Leavitt 2001). Some studies have
shown that isotopic signals have proved to be more sensitive than tree-ring width in
determining environmental, and in particular climatic, conditions (e.g. Saurer et al.
1995; Loader et al. 2003; Helle & Schleser 2004a).
Water is often a limiting environmental constraint and losses from the plant via
evapotranspiration must be kept to a minimum. However, a surplus of water (such as
in boggy environments) can also result in physiological drought and cause stomatal
closure (Kozlowski 1984; Ewe & Sternberg 2002). Stomatal movement is an efficient
mechanism that regulates water loss (either as a result of shortage or excess) from the
leaf surface whilst optimising carbon acquisition from the atmosphere. When stomata
apertures are extremely narrow, the CO2 influx becomes restricted and Rubisco, the
enzyme responsible for carbon fixation, reduces its discrimination against 13 C from
the carbon dioxide pool within the intercellular cavities. This, in turn, affects the rela-
tive 13 C/ 12 C ratio of the organic entities metabolised from the photosynthetic end
product (i.e. glucose) at that time (Farquhar et al. 1989). Hence, plants that have expe-
rienced water stress normally show less negative δ13 C values relative to non-stressed
plants.
Oxygen isotope ratios of plant organic matter, particularly cellulose of tree rings,
may provide additional insights into palaeoclimatic reconstructions. The oxygen isotope
ratio in plant cellulose normally preserves a signal relating to the fractionation between
source water and cellulose; i.e. the isotopic signature of leaf and/or xylem water used
for cellulose synthesis and the isotope composition of precipitation, which is mainly
temperature dependent (e.g. Sternberg et al. 1986; Rozanski et al. 1992; Sternberg et al.
2003). Soil water as well as water vapour exchanged between plant and atmosphere,
are major sources for the 18 O signature of leaf water which in turn determines the δ18 O
signature of cellulose (Saurer et al. 1997a, b; Anderson et al. 1998, 2002). An additional
18 O imprint may be associated with evapotranspiration from the leaves, thus providing
124 IAWA Journal, Vol. 26 (1), 2005

potential proxy information regarding environmental temperature and/or humidity in

which the tree grew (e.g. Förstel 1978; Saurer et al. 1997a; Anderson et al. 1998; Bar-
bour et al. 2002; Saurer 2003). During evapotranspiration H216 O is preferentially lost
from the leaf. Therefore, the δ18 O signature of the plant can provide information relat-
ing to the intensity of transpiration (Scheidegger et al. 2000) such that an increase in
transpiration results in relatively enriched δ18 O values in plant material and vice versa
(Roden & Ehleringer 1999; Waterhouse et al. 2002).
If the soil is permanently saturated, oxygen deficiency in the roots may occur. Oxygen
deficiency lowers the osmotic potential in the roots which in turn hinders hydraulic
conductivity through the tree (Everard & Drew 1987). The tree uses the same strategy
as outlined above in trying to minimize water loss, i.e. by closing its stomata. Since
carbon and oxygen originate from different sources (i.e. carbon dioxide and water,
respectively), the combination of these isotopes might provide a clearer understanding
of the physiological factors affecting tree growth (e.g. Edwards et al. 2000). It must
be noted, however, that biochemical reactions encompassing additional fractionation
steps along the path from source carbon and oxygen to final organic moiety (i.e. cellu-
lose, lignin etc.), can potentially bias environmental assumptions (Roden & Ehleringer
1999).
The aims of this study were to use variations in stable carbon and oxygen isotopes
of tree rings to investigate potential hydrologic changes in bogs and to try and obtain
a better understanding of the physiological processes underlying tree growth. It was
hoped that both these aims would aid dendrochronological interpretations from bog
oaks. This study was not concerned with the actual isotopic value but rather with rela-
tive shifts relating to environmental change that may manifest itself in normal versus
suppressed growth assuming that fractionation associated with biochemical pathways
remained constant.
As with all isotope studies using plant fossils and subfossils, issues associated with
chemical taphonomy (Robertson et al. 1995; van Bergen & Poole 2002) first need to
be determined. Bulk stable isotope compositions are simply the mean average of all
preserved molecules or moieties, i.e. cellulose or lignin. However, each moiety has
its own individual isotopic signature and the preferential loss of certain moieties over
others can greatly affect the resultant isotopic composition (van Bergen & Poole 2002).
Therefore, it is important to first determine the molecular composition of the material
under study to ensure that similarly composed material is being compared.
With the aforementioned in mind, the carbon and oxygen stable isotope compo-
sitions along with possible chemical taphonomical effects were studied and com-
pared with growth ring data in three Dutch sub-fossil bog oaks. It was hypothesized
that the isotope proxies would yield consistent relationships with growth ring varia-
tions in the bog oaks. Moreover, the combined data were used to obtain further insights
into possible regional scale environmental factor(s). In particular, wetter conditions
during the growing season were expected to result in severe physiological stress (similar
to that experienced with drought stress) and thus 13 C and 18 O enrichment of organic
matter.
Sass-Klaassen et al. — Carbon and oxygen isotope dendrochronology 125

3.0
Z74

2.5

2.0

1.5

1.0

0.5

0.0
3.0
Z25
2.5
Tree-ring width (mm)

2.0

1.5

1.0

0.5

0.0
3.0
Z162

2.5

2.0

1.5

1.0

0.5

0.0
-140
-120
-100
-80
-60
-40
-20
-0
20
40
60
80
100
120
140
160
180
200

BC year AD
Fig. 2. Tree-ring width series of three oaks from Zwolle-Stadshagen showing suppressed (heavily
shaded) and normal growth (lightly shaded) in the three trees (Z74, Z25, Z162) studied.
126 IAWA Journal, Vol. 26 (1), 2005

MATERIAL AND METHODS

Material selection and sampling

Stem discs of three sub-fossil oaks (Z25, Z74, Z162) with well-preserved heartwood
were selected from the excavation of an ancient woodland in Zwolle-Stadshagen, eastern
Netherlands. These oaks grew during the period 137 BC to AD 193 and show abrupt
and prolonged growth suppressions that consist almost entirely of earlywood and can
be cross-dated with growth suppressions in oaks from other sites in the Netherlands
and northwest Germany. From each tree, wood was analysed along one radial transect
spanning two successive suppression periods alternating with periods of normal growth
(Fig. 2). Each period of suppression and normal growth, covering between five and 20
tree rings, was subdivided further into two (radial) subsamples. The first, and last two
tree rings of each suppression and normal growth period were excluded to ensure that
there was (i) no contamination from possible lag effects in the use of stored photo-
synthates, and (ii) to avoid cross contamination during sampling. Samples were taken
using a razor blade cleaned in dichloromethane and methanol to prevent organic con-
tamination. Tests have shown that this treatment does not affect the carbon or oxygen
isotopic composition of the samples.

Determination of the molecular composition

The technique employed to determine the molecular composition followed that of
van Bergen and Poole (2002) and Poole and van Bergen (2002). From each period one
sample was selected to determine the molecular composition and ensure consistency
in molecular preservation. The material was powdered and extracted using methanol
and dichloromethane. After each extraction step the material was centrifuged and the
supernatant removed. The final solvent extracted residue was air-dried. All residues,
containing the insoluble organic matter representing the bulk of the wood, were stored
dry in the dark. Approximately 300 mg of each sample was subjected to off-line py-
rolysis (1 hour, 300 °C). The cellulose- and lignin-derived compound released were
derivatised using an excess of N,O-bis(trimethylsilyl)-trifluoroacetamide containing
1% trimethylchlorosilane (BSTFA + 1% TMCS) and pyridine, and analysed using a
Hewlett-Packard (HP, Wilmington, DE, USA) 6890 gas chromatograph (GC) (see Poole
& van Bergen 2002 for details). Compounds were identified using an HP 5890 series
II GC coupled to a Fisons Instruments VG platform II mass spectrometer (Manchester,
UK). Identification was based on mass spectral characteristics and data presented in
the literature (e.g. Poole & van Bergen 2002).

Cellulose extraction
The cellulose from each sample was extracted following the technique outlined in
Wiesberg (1974) following Sohn & Reiff (1942). An update of the procedure is given by
Loader et al. (1997). Briefly, between 10 and 30 mg of the ground sample was treated
in 5% NaOH solution for 2 hr at 60 °C. Afterwards, the solvent was renewed and the
sample again treated for 2 hr. Subsequently the sample was washed with hot distilled
water. This procedure transformed the sample into a more reactive form during which
the hemicelluloses, tannic acids and resins were dissolved. Finally the sample was added
Sass-Klaassen et al. — Carbon and oxygen isotope dendrochronology 127

to a 7% NaClO2 solution with 1% acetic acid for pH buffering. The solution gradu-
ally became pale yellow through the release of ClO2 which acted as the actual oxi-
dizing agent. The NaClO2 solution was renewed every 24 hr for five days. After two
to four days the pure cellulose was rinsed in hot distilled water and dried at 80 °C in
a vacuum oven.

δ 13C and δ 18O determination of bulk wood and cellulose

On-line bulk stable carbon isotope (δ13 C bulk wood) analysis was performed on the
extracted wood using a Fisons Instruments NA 1500 Elemental Analyser (EA). The CO 2
released was flushed through the ConFlo II interface and analysed using a Finningan
MAT Delta Plus isotope ratio mass spectrometer (IRMS) in the Faculty of Earth Sciences,
Utrecht University (The Netherlands). To ensure consistency between laboratories the
δ13 C bulk wood and δ13 C cellulose were also measured at the Research Centre in Jülich
(Germany) with an NA 1500 Elemental Analyser interfaced to an IRMS (Micromass
Optima). Similar equipment was used, applying the on-line pyrolysis method described
by Werner et al. (1996). Samples (300 μg) of bulk wood or cellulose were pyrolysed
to CO at 1080 °C. The δ18 O values are referred to VSMOW and the overall analytical
uncertainty is ± 0.25 ‰. Nine different commercial carbohydrates (celluloses, starches,
sugars) used as internal laboratory standards were calibrated against eight European
laboratories of the EU-project ISONET (EVK2-2001-00237). Sample replicates were
measured at intervals to test reproducibility.

RESULTS
Molecular composition
The resulting chromatograms of the off-line pyrolysates verified the presence of the
various chemical moieties (i.e. cellulose, lignin) (cf. van Bergen & Poole 2002; Poole
& van Bergen 2002). One representative chromatogram is shown (Fig. 3) revealing that
the wood samples are composed predominantly of cellulose, as shown by the presence
of levoglucosan and other polysaccharide pyrolysis products (cf. van Bergen & Poole
2002). Lignin, as revealed by guaiacols and syringols, is present but in much smaller
relative abundances.

Stable isotope content

As previously mentioned relative isotopic shifts rather than absolute signatures are
considered because chemical taphonomy has acted in a similar manner on all samples
studied. Consequently the measured δ13 C bulk wood and δ18 Obulk wood values can be in-
terpreted without further correction for differences in molecular composition. The rela-
tive difference between the isotopic values of the bulk wood and cellulose (Fig. 4a, b) is
because the bulk wood value represents the weighted mean average of the isotopically
heavy cellulose plus the isotopically light lignin (cf. Spiker & Hatcher 1987; Benner
et al. 1987; Poole & van Bergen 2002), whereas the isotopic value of cellulose simply
reflects the signature of the cellulose alone albeit possibly slightly altered chemically
(Schleser et al. 1999a).
128 IAWA Journal, Vol. 26 (1), 2005

:
Levoglucosan
Relative intensity

PS

20 30 40 50

: Levoglucosan

PS
Relative intensity

PS

PS

15 30 45
Retention time (min) 7
Fig. 3. Representative chromatogram of the off-line pyrolysate of tree Z74 (years 35–32 BC).
Each peak represents a different pyrolysis product and can be linked to the original source moie-
ties (i.e. cellulose, lignin; van Bergen & Poole 2002; Poole & van Bergen 2002). The most abun-
dant products, levoglucosan and polysaccharide pyrolysis product (PS), have been identified as
derivatives of cellulose.

Many authors observe that the isotopic composition of bulk wood and cellulose
correlate well and that the slope of the linear regression is not significantly different.
This has been reported for stable carbon isotope ratios (Livingston & Spittlehouse
1996; Borella et al. 1998; MacFarlane et al. 1999; Helle & Schleser 2004b), as well as
Sass-Klaassen et al. — Carbon and oxygen isotope dendrochronology 129

stable oxygen isotopes (Borella et al. 1999; Saurer et al. 2000). Interestingly, Loader
et al. (2003) found that δ13 C of bulk wood is a better climate proxy than δ13 C of α-
cellulose.

δ13C: Similarity of the δ13Cbulk wood values measured in Utrecht and those measured
in Jülich confirms the accuracy of measurements and the homogeneity of the samples.
Figure 4a shows the variation in δ13 Cbulk wood and δ13 Ccellulose between the alternating

cellulose bulk wood

-24 -24

-25 -25

-26 -26
δ13C (‰)

-27 -27

Z162
-28 -28
Z74
Z25
-29 -29
49 – 46
45 – 42
39 – 36
35 – 32
29 – 26
25 – 22
19 – 8
7– 4

49 – 46
45 – 42
39 – 36
35 – 32
29 – 26
25 – 22
19 – 8
a 7– 4

BC AD BC AD

cellulose bulk wood

30 30
29 29
28 28
27 27
26
δ18O (‰)

26
25 25
24 24
23 23
Z162
22 22
Z74
21 21
Z25
20 20
49 – 46
45 – 42
39 – 36
35 – 32
29 – 26
25 – 22
19 – 8
7– 4

49 – 46
45 – 42
39 – 36
35 – 32
29 – 26
25 – 22
19 – 8
7– 4

b
BC AD BC AD
Growth ring period

Fig. 4. δ 13 C (a) and δ 18 O (b) values for bulk wood and cellulose measured at the Research
Centre Jülich for normal (unshaded) and suppressed (shaded) growth periods for the three trees
(Z162, Z74, Z25) studied. Measurements done at Utrecht University are not shown as they are
virtually identical to those obtained from Jülich.
130 IAWA Journal, Vol. 26 (1), 2005

periods of normal (lightly shaded) and suppressed (heavily shaded) growth for the three
oak trees studied. In each sample the overall trend for both δ13 Cbulk wood and δ13 Ccellulose
are similar even though the two trends are offset by up to 2‰. The dominance of the
cellulose in the bulk samples is confirmed by the similarity of the δ13 Cbulk wood trend to
that of the δ13 Ccellulose. A significant difference in isotopic signature between suppressed
growth and normal growth is seen in tree Z74, in which the former has more negative
δ13 Cbulk wood and δ13 Ccellulose values relative to the periods of normal growth. The varia-
tion between normal and suppressed growth for the other samples is not striking but
the same general tendency can be seen in Z25. Z162 shows either no difference or a
slight bias towards a δ13C enrichment during the periods of suppressed growth.

δ18 O: The variation in δ18 O of suppressed and normal growth is illustrated in Figure
4b. As with the δ13 C values, there is an offset between the δ18 Obulk wood and δ18 Ocellulose
values accompanied by significant differences. The variability in δ18Obulk wood values is
greater than that of δ18Ocellulose values with the absolute levels of δ18Obulk wood values
varying between the individual tree ring sequences. Over the whole period δ18Obulk
18
wood of Z162 and Z74 is characterised by a declining trend, whereas δ Ocellulose of all
three individuals is relatively similar and does not show a long term trend although
a slight initial δ18 O depletion (i.e. in the first suppressed sample) can be detected in
periods of suppressed growth.

DISCUSSION

Although the wood of individual oaks from the excavation in Zwolle was not preserved
in exactly the same place, it had nevertheless been subject to a similar depositional (i.e.
bog) environment and this was reflected in the resulting similar molecular composition
(e.g. Fig. 3). Cellulose was still the dominant component whereas lignin was present
in smaller relative quantities. Thus, the overriding isotope signal is biased in favour
of cellulose but this bias is the same for each sample and further correction to ensure
comparison of like bulk values was not necessary.
For each of the three oaks, both δ13Cbulk wood and δ13Ccellulose values throughout the
tree-ring series indicated similar relative trends. In two cases, i.e. trees Z25 and Z162,
the molecular evidence confirmed that cellulose was the dominant moiety even though
additional 13 C-depleted moieties, including lignin, resulted in the bulk values being more
13 C depleted. Comparison between the δ13 C 13
bulk wood and δ Ccellulose values revealed
larger variations for Z74 suggesting slight differences in preservation, i.e. more/better
preserved lignin and/or molecular differences in the cellulose preserved.
Although the differences (< 2.4‰) in δ13C values between the suppressed and normal
growth phases are not greater than variations (1–3‰) documented for intra-ring vari-
ation (Schleser et al. 1999a; Leavitt 2002; Helle & Schleser 2004a, b), slight changes
nevertheless were observed. Trees Z25 and Z162 showed no apparent directional trend
from normal to suppressed growth in terms of either δ13Cbulk wood or δ13Ccellulose (Fig.
4a). This could be explained by extreme physiological stress culminating in a late
onset – or early cessation of radial growth as evidenced by narrow rings comprised
Sass-Klaassen et al. — Carbon and oxygen isotope dendrochronology 131

exclusively of earlywood vessels (Fig. 1). Earlywood formation starts before bud break
and is completed before the leaves have fully expanded, i.e. before they become net
exporters of assimilates. Consequently, earlywood incorporates carbon from previous
years and may not contain the same isotopic signature as that of the current year (Helle
& Schleser 2004b). In general, stable carbon isotope shifts can occur as a result of
carbon partitioning at metabolic branching points. The extent of such carbon isotope
shifts depends on pool sizes and various flux rates into the different metabolic pathways.
During periods of normal vegetative growth, carbon is partitioned between wood for-
mation and storage. During seasons with suppressed growth, the bog oaks formed no
latewood and therefore no isotopic signal was recorded because the assimilates were
stored (as starch) preferentially for next yearʼs earlywood formation. Unfortunately,
present knowledge permits only speculations regarding fractionations involved in the
seasonal interplay of major tree physiological processes such as accumulation and
remobilization of storage material, varying stem respiration and the formation of tree
rings. Therefore, it may well be that environmental signals evolving at the leaf level
are masked by counteracting post-photosynthetic physiological processes.
However, tree Z74 reacts differently from the others. This tree shows a tendency for
more negative δ13C values during suppressed growth years when compared with the
normal growth years. This is surprising as it is contrary to the findings of other workers
where periods of stress resulted in δ13C enrichment (e.g. Saurer et al. 1997a). Although
the growth pattern of Z74 is suppressed, this tree might not have experienced the degree
of physiological stress experienced by the other two oaks and thus the more depleted
signal could be explained by relatively enhanced humidity consistent with the findings
of Saurer et al. (1995). Additionally, Z74 shows a higher variability of δ13C than the
other two trees, whereas δ18 O-values are rather similar. The higher δ13C-values during
the drier phases of normal growth may indicate that this tree grew under drier condi-
tions relative to those of the other two trees. However, the similarity of δ18 O-values
suggests the same water source for all trees. Tree Z74 may have regulated stomatal
activity such that δ13C was affected by changing leaf internal CO2 partial pressure, but
transpiration rates remained constant leading to almost no changes in δ18 Ocellulose.
The differences in δ18 O observed between suppressed and normal growth years
(< 3.6‰) are again lower than the differences (4.5‰) documented for intra tree-ring
variation (Helle & Schleser 2003, 2004a). However, a similar scenario to that observed
with 13C values is noted for the δ18 O values (Fig. 4b) between normal and suppressed
growth years. Although there is no obvious overriding difference, a slight tendency
towards depleted δ18 O values in the samples of suppressed growth suggests a decrease
in evapotranspiration possibly due to relatively wet conditions associated with high
relative humidity. Alternatively, reduced stomatal conductance caused by inundation
might have reduced evapotransporation and thus the preferential loss of the H216 O,
leading to more 18 O depleted values.
The low variability of δ18 Ocellulose regardless of ring width possibly resulted from
a constant source value. In addition, the low variability of δ13 C observed for Z25 and
Z162 suggests that changes in air humidity were probably small and did not greatly
alter the transpiration rates of trees. However, it is also possible that environmentally
induced changes in 18 O enrichment of leaf water were superimposed onto changes in
132 IAWA Journal, Vol. 26 (1), 2005

uptake of differently labelled source water. This hypothesis is supported by the obser-
vations of Ewe and Sternberg (2002) who found no major variations between wet and
dry season in δ18 O in plant water for some woody plants from the Florida Everglades
even when they could show that source water taken up by plants had shifted from deep
groundwater during the dry season to shallow soil water during the wet season. Sea-
sonal isotopic changes in groundwater and soil water, respectively, were counteracted
by changes in water uptake by the trees.
The obvious differences between δ18 Ocellulose and δ18 Obulk wood cannot be explained
by variations in chemical composition of the ligno-cellulose which would have reflected
potential differences in chemical preservation. However, intra-molecular O-isotope
exchange during the initial stages of preservation might have affected bulk wood or cel-
lulose oxygen isotope signatures, while intra-molecular C-isotope signatures remained
unaltered.
Environmental conditions that drive the observed tree-ring pattern are not reflected
in obvious trends within the stable isotope signatures. Suppressed growth was probably
caused by dramatic hydrological changes and/or soon after, the onset of the growth
season resulting in a shortened growth season (hence the absence of latewood). These
results agree with those of Mayr et al. (2003), who also studied hydrogen and carbon
isotopes from tree rings of sub-fossil oaks originating from southern Germany. They
also found no uniform trend in δ13Ccellulose signatures among trees and advised caution
in trying to reconstruct palaeoclimate solely from stable isotopes.

CONCLUSION

This study applied stable isotope dendrochronology to well-dated 2000-year-old sub-fos-

sil bog oaks from The Netherlands. These trees have characteristic ring-width patterns
alternating between periods of normal and suppressed growth. Similar growth patterns
have been observed in other ancient wetland sites across northwest Europe suggesting
that the controlling factors for radial growth were regional rather than local (Baillie
1995; Pilcher et al. 1996; Leuschner et al. 2002). It was hypothesised that since the
trees were growing in a wet environment the overriding factor affecting the ring-width
pattern was the hydrological status which in turn could be explored using stable carbon
and oxygen isotopes of the cellulose and lignin. Determination of the chemical compo-
sition of the ligno-cellulose in the woods ensured that any differences in stable isotope
variation were not due to differential preservation of the inherent chemical moieties
(i.e. cellulose, lignin). It cannot be excluded that intra-molecular O-isotope exchange
during the initial stages of preservation has altered the δ18 Ocellulose and/or δ18 Obulk
wood signatures whilst intra-molecular C-isotope signatures have remained unaltered.
The absence of distinct trends in the stable isotope composition between suppressed
and normal growth periods is believed to be due to the absence of organic tissues, i.e.
latewood, which would be expected to contain this signal. This study implies that the
ring-width chronologies of bog oaks are a sensitive monitor of the external environment
but that environmental changes were not preserved in the stable carbon and oxygen
isotope ratios of wood and cellulose.
Sass-Klaassen et al. — Carbon and oxygen isotope dendrochronology 133

ACKNOWLEDGEMENTS

This research was made possible by funding from The Netherlands Organisation for Scientific Research
(NWO), number AWL/750.700.04 and ALW/809.32.004, the German Climate Research Program
(DEKLIM, grant no. 01LD0004) and EU 5th framework project ISONET (EVK2-2001-00237). The
authors thank Melanie Schrimpf, Gaby Reiss and Werner Laumer for their assistance during sampling,
sample preparation, cellulose extractions and isotope ratio measurements. The careful and critical
comments by Dr. W.T. Anderson and an anonymous reviewer greatly improved this manuscript.

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