UDK 903:612.39(497.4)"634f- Documenta Praehistorica XXVI Stable isotope evidence of the diet of the Neolithic population in Slovenia - a case study: Ajdovska jama Nives Ogrinc "J. Stefan" Institute. Department of Environmental Sciences. Ljubljana. Slovenia nives.ogrinc@ijs.si ABSTRACT - Tfw aim of this research nos to determine the nutrition habits of Neolithic people tiling in Stovenia between 4000 BC and 3400 BC. The specific isotopic composition of different types of food is reflected in the isotopic composition of the tissues of the consumer. Therefore, by measuring the isotopic composition in the tissues we can draw conclusions about the nutritional luibits of the consumer It e analysed the remains of human bones taken from Ajdovska jama atui determined the stable isotopic composition of carbon and nitrogen in the bone collagen. Our results indicate that the diet consisted primarily of herbivores, most probably domestic animals. IZVLEČEK - Namen našega delu je ugotoviti prehranjevalne navade neolitskega človeka na naših tleh iz petega in četrtega tisočletja BC. Izotopska sestava ogljika in dušika v hrani je različna in se odraža v izotopski sestavi tkiv uživalca, zato lahko na podlagi meritev izotopske sestave sklepamo o njegovem prehranjevanju Analizirali smo vzorce kostnih ostankov iz Ajdovske jame in določili izo-topsko sestavo ogljika in dušika v kostnem kolagenu. Rezultati meritev kažejo, da so pretežno prehrano naših /trednikov predstavljale rastlinojede, v glavnem domače živali. KEY WORDS - Ajdovska jama, Neolithic diet; stable isotopU• composition of carbon and nitrogen IiNTRODl CT10N Since its introduction in 1977, stable isotope analysis of bone collagen has become a very valuable tool for determining prehistoric human and animal diets (DeNiro and Epstein 1978. Tauber 1981, Scliwartcz and Schoeninger 1991; Lubell et al. 1994; Schtd-ting 1998). The inorganic and organic chemical constituents of bone provide a record of long-term dietary intake. Elements and amino acids liberated by the digestion of food are incorporated into bone minerals, collagen and non-collagenous bone proteins throughout a vertebrate's lifetime. Dietary information is thus recorded by carbon and nitrogen isotope ratios in bone collagen and by carbon isotope ratios in the carbonate component of the inorganic portion of bone minerals (bioapatite) and teeth mineral (Krueger and Sullivan 1984). The reconstruction of individuals diets using isotopic methods has been limited to the analysis of collagen preserved in bone, because bioapatite is more difficult to deal w ith due to problems of diagenesis (Schoeninger and DeNiro 1982). Carbon isotopes are fractionated by natural processes such as the photosynthetic assimilation of C0> and its adsorption in water. Carbon fractionation is affected by the type of metabolism used by a plant to fix CO; and differs in the marine and terrestrial foods chain and may therefore be used to elucidate questions on the origin of naturally occurring carbon compounds. Due to kinetic isotope effects, terrestrial plants that follow normal Calvin (C$) photosynthesis are depleted in the heavy carbon isotopes. as is shown in a change in 8'3C values from -6 to -8%o in atmospheric C0.> to -24 to -32%o in terrestrial plants. Terrestrial animals and human feeding on such plants show a similar HC content, although with a slight shift towards higher 5UC values. The absorption of C0> in water and the subsequent formation of bicarbonate are governed by kinetic isotope effects and by thermodynamic equilibrium processes, which lead to an enrichment in heavy carbon isotopes to 5' *C v alues closed to 0%o. When marine bicarbonate is assimilated during pho- 193 N.vos Ogrne «»synthesis in submerged plants, reaction kinetics again result in depletion in the heavy isotopes, in this case to Sl*C values about -10 to -18%o, and this fractionation is also reflected in marine animals and human whose food was based mostly on marine protein. The simple isotopic separation between terrestrial and marine plants and animals is partially obscured if the photosynthetic cycle has dominated in terrestrial plants. The isotopic composition of these plants ranged between -10 to -l6%o. The most important cultivated G, plants were maize, sugar cane and millet. However, in our study, which concerns temperate Europe, the influence of C, plants can be excluded. Little is known about the nitrogen isotopic composition of different types of food. It has been suggested that plants that can fix molecular nitrogen (due to the presence of symbiotic bacteria) have characteristically lower ISN/UN ratios than those which must assimilate other forms of inorganic nitrogen, such as ammonia or nitrate {Deln iche et til. 1979). The differences between the 6ISN values of the two ty pes of plant appear to vary depending on the location in which they grew and the tinte of year at w hich they were collected (DeNiro and Epstein 1981). The basis for the geographical and temporal variability of plant 5|SN values must be resolved before a dietary analy sis based on the isotopic ratios of animal nitrogen can be exploited to its full potential. It may also be possible to use the nitrogen isotopic method of dietary analysis to determine the relative amounts of terrestrial and aquatic food sources eaten by animals living in shore environments. Stable nitrogen isotopes, ISN, are also enriched in marine systems relative to terrestrial systems, but for such studies die degree of trophic level fractionation is more important Enrichment of HN through the trophic network is widely recognised among most animals, including invertebrates and vertebrates. leading to a value of 3.4 ±1.1 %o (Minagatca and Wada 1984. Wada et al. 1987). These facts suggest that the isotopic composition of organisms provides basic information not only about their food source, but also the trophic level. Most marine fish that are eaten by humans are carnivores, and marine food chains are relatively longer than terrestrial chains, therefore contents are relatively high. The is true of lake fish, so that humans consuming a substantial proportion of fish and/or mammals will have higher stable nitrogen value than is possible to attain in a purely terrestrial system (Schoeninger et al. 1983; Katzenberg 1989). Two important issues must also be considered. First, stable isotope results on human bones provide insights into the amount of protein an individual has consumed over approximately the last ten years of their lifetime (Chishohn et al. 1983. Schu arcz and Schoeninger 1991). And the second, the assumption that the bone collagen isotope ratio has not been modified by post-mortem processes. It was shown that the material isolated from prehistoric bones, with C/N ratios between 2.9 and 3-6 have not undergone diagenetic alteration (DeSiro 1985). It is possible they have, but previous studies suggest that such diagenetic shifts must be small enough for identification of the basic feeding behaviour of the individuals from their bone collagen isotope ratios to be possible. M Vyó y < ; : - • w x I r «• »? V- : J .. Fig. 1. Ajdovska Jama near Nemška vas is located in the south-eastern foot-hills of the Krško highlands. 194 Stable isotope evidence of the diet of the Neolithic population «1 Slovenia - a caw study; Ajdovska |ama Fig. 2. Human skeletons uere discovered in the le/t corridor and in the central liall of the cace. The aim of this paper is to determine the nutritional habits of Neolithic people living in Slovenia between around 4000 BC and 34(H) BC using isotopie methods. The isotopie composition of different plant and animal remains in association w ith human skeletons were also determined in order to be able to define more precisely the roles of plants and animal protein in the diet of the humans living at the time. MATERIALS AND METHODS lite human bone samples from Ajdovska jama near Nemška vas, also known as Kartušova jama located in the south-eastern foot-hills of the Krško highlands were collected in this study (Fig. 1). Human skeletons were discovered in the left corridor and in the central hall (Fig. 2). Anthropologists managed to identify 29 individuals, namely 13 adults (6 males and 7 females) and 16 infants. Different faunal species associated with the burials were also found. The most represented species were domesticated mammals, such as cattle, sheep, pigs, and plants - mostly wheat, which implies some domestic activity during both periods. Also, some remains of wild animals were found: brown bear. deer, field hare. The floral and faunal remains and the human bone samples were then transferred to labelled, polythene bags w hich were sealed until the start of pre-treat-ment. The AMS <*C analysis w as used to date the burial remains in Ajdovska jama. The results indicate that the samples are from two different periods ca 5300 and 6000 yrs BP. Sample preparation Since the accuracy of measurements mainly depends on the variability inherent in the collagen extraction and measurement techniques, we have compared two methods that have been developed for collagen extraction and purification. Four samples were selected for this test. The bone samples were cleaned in cold, distillate water in an ultrasonic bath to remove soil contaminants, and then oven-dried at 50°C to constant weight The samples were ground in a mill to -1 mm fine powder and subdivided into two portions. The first extraction method was that described by Longin (/97/). Approximately 1 g of bone powder was weighed into a 250 ml beaker and 150 ml of 1M hydrochloric acid (HCl) added to remove the acid soluble inorganic portions of the bone, any acid soluble protein and peptide fragments, and free amino acids. The acid also breaks down some of the hydrogen bonds of collagen, so that it becomes soluble in hot water. The pre-treatment time must be short (< 20 min). otherwise die proteinic chain is hydrolysed and the collagen becomes soluble in hot water and is then lost The acid solution is then discarded by filtration through a glass microfibre filter and well washed with distillate water. The remaining acid insoluble material, which includes unde-natured, and insoluble collagen, is extracted under reflux for 10 hours in a hot water (90°C) of pH = 3. The heating serves to denature and partially hydrolyse the intact collagen, making it soluble, while the acidic pH 3 avoids dissolv ing any non-acid-soluble contaminates. The solution is then filtered through an 8 (jm polyethylene filter to remove insoluble residues. and the collagen isolated by freeze-dtying the filtrate. Method 2 is the modification of the Longin method suggested by Richards and Mellars (1998). In this case, the inorganic portion is removed by extraction with 0.5 M HCl solution. The samples were kept at 4-5°C overnight. Powdered samples collected on glass microfibre filters were washed twice with dis tillate water. Then the residue was placed in a sealed 195 Nives Ogrinc tube (under reflux) in a pi 1 3 HCI solution, and gelatinised for 48 hours at 75°C. The solution is then decanted and filtered through an 8 (.un polyethylene filter and freeze-dried. This more gentle treatment is intended to reduce the collagen loss as compared to method 1. No measurable effects on the determining of and only a small effect on the 5,5N signal were obtained using these two methods. We conclude that the best collagen extraction technique is the second method proposed by Richards and Mellars (1998). Method 1 gives essentially the same results, but the collagen recovery may be lower. Other researchers have included a sodium hydroxide wash in Uieir preparation sequence (DeNiro and Epstein 1981:1'ate 1995). The sample was treated before the hot w ater extraction step w ith 0.5% sodium hydroxide (NaOH) for -20 h to remove base soluble contaminates such as humic acids. The results from other studies show that this pre-treatment has little effect on measured 8'iC and 81SS values, and because of the apparently reduced yields (Chrishol et at. 1983: Bonsall et al. 1997) we decided not to use this method. Isotopic analysis The isotopic composition of collagen was then determined using a Europe Scientific 20/20 continuous flow mass spectrometer with ANCA-SL solid-liquid preparation module. The technique involves the coupling of a preparation system employing the Dumas principle with a stable isotope mass spectrometer detector. This allows measurements of not only total nitrogen and carbon in a sample, but also their i*N and >3C levels. The "collagen" solid w as placed in tin cups and dropped sequentially into a combustion tube as a pulse of oxygen was injected. After various reduction reaction and chemical trapping, carried out in a helium carrier which transports released gases, the gas chromatography (GC) column separates N> and CO_> from trace impurities before analysis by the IRMS (Isotope Ratio Mass Spectrometer). The samples are analysed in batches that include known references (working standards). References calibrate isotopic abundance and elemental composition measurements and allow correction for drift. As a working standard, pure collagen was used which was calibrated vs. reference materials (IAE\-CH-7 polyethylene and NBS22 oil for carbon; L\EA-N-l and IAEA- NO-3 for nitrogen). Following standard procedure, the isotopic ratios are expressed in 8-notation in parts per mil (%o): Lcx/x) X)sampk' 'X Standard x 1000 For carbon, *X/X is1 *C/'-C and the standard is the V-PDB carbonate, while for nitrogen *X/X is HN/KN and the standard is atmospheric (air) nitrogen. The measurement uncertainties on the 8' *C value of -24.7 ± 1.5%o and 8'S.N value of +3-3 ± l%o. The difference in the isotopic composition in plants was also observed. Leguminous plants have 815N values of +1.9%o, w hile the non-legumes, in our case w heat, have 815N 196 Stake .sotope evidence ol tip diet ol the fveolitriic population in Slovenia - a case study Ajdovs*a |ama 25- 20 15 - 7- io- lo 5 -I ■ -r -30 ■ -25 T -20 T---1 -15 -10 6I}C [%o] Fig. J. SUC and cPf.V values for foods available in Seolithic and terrestrial (T) herbivores established by Schuarcz (1991). Sample Age 8>3C 8»N C/N (yr BP) [%.] 1%«) ANIMALS domestic catde 5300 -21.1 +5.8 34 domestic sheep 5300 -19.6 +7.0 32 deer * 5300 -24.3 +6.3 6.7 * domestic cattle > -20.4 +6.7 34 brown bear > -22.1 +1.4 3.2 deer * ? -24.2 +5.7 6.5 * domestic cattle 6000 -21.7 +5.8 34 domesUc cattle 6000 -20.4 +5.8 3.2 domestic cattle 6000 -20.0 +6.0 3.2 domestic pig 6000 -20.0 +5.7 3.3 field hare 6000 -21.6 +3.8 33 brown bear 6000 -22.0 +0.0 3.7 deer 6000 -21.5 +5.9 3.7 PLANTS wheat (mono-grain) -24.6 +4.2 wheat (two-grain) -25.3 +2.8 barley -25.9 +4.3 mixture peas -23.2 +1.9 Tab. 1. The isotopic composition of carbon and nitrogen of plants and animal remains associated u ith human skeletons. values of +3.8 ± 0. •*%<>. Also, the results obtained from animal bones show that their diets favoured Cj plants. The clear distinction between domestic and wild animals was observed in the 6'^N values. Samples from terrestrial wild herbivores have 8>*N values in the range of 0 - 6%o (average - 3-7%o), while seven samples from domestic animals analysed have 8'\N values between 5 -7%o (average - 6.1%o). These results are in good agreement with the two studies concerning analyses of wild animals performed by Schwartcz (1991), and domesticated animals av ailable today in temperate Europe (Botisall el al. 1997). It is still not clear why the (1SN values of wild and domesticated herbivores differ so markedly. Most probably the difference is connected with feeding patterns. Domesticated animals could be fed foodstuffs that were not available to wild herbivores. It is conceivable that animals kept by farmers ingested some cultivated vegetables (the 8ISN v alues are -3%« higher than that observed in plants - wheat, legumes) and also a certain amount of human food refuse. Our results for prehistoric domesticated animals correspond well with those obtained from modern livestock, suggesting that animals had similar foodstuffs and should be regarded as having more omnivorous diets than their wild counterparts. Palaeodietary reconstruction The and 6'^N of collagen extracted from human bones from two different periods 5300 and 6000 BP are collected in table 2 and graphically presented in figure 4. No significant difference in the dietary habits of the population between the two periods was found. The isotopic composition of carbon in the human samples range between -22.5 and -19.6%«, while 51<>N values range from +4.9 to +11.5%o. The 8'^N and 8'X] values plotted in a single, well-defined cluster suggest a diet that was relatively homogeneous and predominantly based on a purely terrestrial system. In other words, essentially all of the protein in the diet over at lexst 10 or so years of their lives came from the terrestrial system. The very uniform 8'^N values, around 9%» through this period, are indicative of a population obtaining most of its protein from herbivores, domestic and wild animals, and relatively little from plant foods. Similar values are observed in Neolithic human remains from southern Portugal (Straus el al. 1992; Lubell et al. 1994). Whatever the case, there is no good linear correlation between 8'*N and 81SC values. A simple explanation of the linear trend of these two values is that the individuals whose collagen is plotted along the line were eating varying proportions from only two isotopically distinct, homogeneous food sources. So, the data suggests that diets were not homogeneous and there was a de- 197 Nives Ogrinc 25 2015- l • j2 105 - M o t -p' T -30 -25 -20 6,3C [%>] ■ 5.300 BP o 6.000 BP 1 -10 -15 Sample Age (yr BP) 5'JC [%.] 8|5N |%o] C/N HUMAN Male 5300 -22.5 +5.6 2.9 Male 5300 -21.2 +5.6 34 Male $300 -20.5 +9.0 35 Female 5300 -20.9 +8.1 3.4 Female 5300 -20.7 +93 3.4 Female 5300 -20.5 +8.7 3.3 Child (boy 10-12 yrs) 5300 -20.3 +9.4 32 Child (boy 10 yTs) 5300 -20.8 +7.5 2.9 Child (7-8 yrs) 5300 -20.8 +8.2 2.8 Child (boy 6 yrs) $300 -22.4 +5.8 3.5 Child (6 yrs) >300 -20.6 +9.1 30 Child (4 yrs) >300 -21.0 +6.6 3.2 Child (2 yrs) 5300 -20.7 + 10.0 33 Male 6000 -22.2 +4.9 3.2 Male 6000 -20.9 +7.7 3.2 Male 6000 -20.6 +7.9 3.3 Female 6000 -20.9 +8.5 32 Female 6000 -20.5 +7.9 3.2 Female 6000 -20.2 +7.3 3.3 Child (6-7 yrs) 6000 -21.6 +8.3 3.6 Child (5 yrs) 6000 -20.3 +10.7 3.5 Child (boy 4 yrs) 60(H) -21.3 +6.6 3-6 Child (1-2 yrs) 6000 -20.7 +10.2 2.9 Child (1-2 yrs) 6000 -20.4 ♦11.5 35 Child (1 yr) 6000 -20.7 +10.8 35 Fig. 4. Scalier diagram of SMC and values in human samples from AJdovska Jama. The ranges of aquatic carnivores and omnivores (A), marine carnivores and omnivores (M) and terrestrial foods (7) established by Schuartcz ^1991,) for 5UC and +?"«»for S».N Is added to arrive at consumer values). crease in the diversity of food choices during both periods in the Neolithic. The results indicate different nutritional habits among the Neolithic population in the central part of Kurope in comparison with the study of Bonsall et al. (1997). ITie results of this study, which includes the Mesolithic and also the earliest Neolithic inhabitants, suggest that in the Mesolithic period people had high protein diets derived mostly from riverine food sources. A shift in dietary patterns occurred between 7600 and 7300 BP. reflecting the intake of higher proportions of terrestrial food. It seems that changes coincide with die introduction of cultivation in the Iron Gates, which was not so dramatic as that seen in some other areas of Europe, such as Portugal (Straus et al. 1992; Lu-bell et al. 1994). TradiUonal food sources were not abandoned in favour of agricultural produce. Comparisons of the stable isotopic ratios of collagen from males, females and children are presented graphically in figure 5. It is seen from the results that in a given population w ith a range of possible foods, the individuals in the population have individual, personal, preferences in diet which in turn determine their individual 6-values. The different nutritional habits of individuals can be seen more clearly from the 5ISN values ranged from +4.9 to +1 1.5%o. The lowest v alues of 815N found in human collagen and average 51SN value of 6.1 ± 0.9%» for domestic Tab. 2. The collagen stable isotope values in hunum skeletons. Ihe table includes the amount of carbon vs. nitrogen in the extracted collagen samples are collected. animal collagen, fit reasonably well with the expected values of 6%o for vegetarian humans and herbivores. respectively. It is interesting that the lowest values are found in males' collagen, and there is a tendency for females to be associated with higher 51H: and S"S ivlues of \eoliihlc' males, females and children from Ajdorska jama. difference in nutritional habits may he related to different food supply activities. It is very difficult to see how the nutritional demands of pregnane) or the division of labour could account for the differences seen in men and women. Comparing the results of adults and children also indicates significant differences. The highest isotopic composition of nitrogen was found in the bones of one- and two-year-old children. The values are approximately 3°oo higher than those found in the female bones, indicating the weaning effect". These results could not be compared with other studies, because there is no data available on the bone samples of children at the same age until now . What is more remarkable is that the infants between 4 and 10 years old have lower 5ISN values in comparison with children one to two years old. These results indicate a new dietary regime that has been identified in die isotopic composition of collagen in a short period over two and ten years. The most probable reason is that in children bone collagen deposition has a very high turnover rate in comparison to adults. Therefore, bone chemistry changes quickly and can reflect new nutrition habits. A difference between older children is also observed. From the S'*C and 8,SN values (approx. -22%« and +6%o. respectively) it is seen that the protein consumed by some children is based mostly on plants, most probably on cereals, while the diet of other children favours meat from domesticated animals. The results correlate extremely well w ith the hypothesised model of human diets calculated hv R. Schulting (1998 206. Tah. I). In conclusion it is worth noting that the stable isotope evidence suggests that the Neolithic population living in Slovenia had individual, personal preferences in diet in which the bulk of the protein was derived from terrestrial food sources. This diet was based mostly on herbivores, domestic and wild animals and relatively little on plant foods. The most interesting and original results are obtained in infants and young children. The significant higher 6HN values certainly relate to "weaning effects", while older children had new dietary habits which markedly changed the isotopic signature of bone collagen in a relatively short period. ACKNOWLEDGEMENT I would like to thank Prof. Mihael Budja for his indispensable collaboration and the opportunity to partidpate in an enjoyable seminar. I am also grateful to Mrs. Milena Horvat. who provided the bone samples from Ajdovska jama. 199 Nives Ogrinc REFERENCES BONSALL C., LE.NNON R.. McSWEENEY K., STEWART C.. DARKNESS D.. BORONEANT V.. BARTOSIEW1CZ L„ PAYTON R.. and CHAPMAN J. 1997. Mesolithic and Early Neolithic in Iron Gates: A Palaeodietary Perspec-ti ve. Journal of European Archaeology 5: 50-92. CIIRISIIOLM B. E.. NELSON D. E. SCHWARZ H. P. 1983- Marine and Terresterial Protein in Prehistoric Diets on the British Columbia Coast Current Anthropology. 24:396-398. LUBELL D , JACKES M., SCHWARCZ H., KNYF M. and MEIKLEJOHN C. 1994 The Mesolithie-Neolithic transition in Portugal: Isotopic and dental evidence of diet Journal ofArchaelogical Science 21:201-216. DeNIRO M.J. 1985. Post-mortem preservation and alteration of in t iro bone collagen isotope ratios in relation to palcodietarv reconstruction. Aalure 317: 806-809. DeNIRO M.J. and EPSTEIN S. 1978. Influence of diet on the distribution of Carbon Isotopes in Animals. Geochim. Cosmochim. Acta. 42: 495-506. 1981. Influence of diet on the distribution of Nitrogen Isotopes in Animals. Geochim. Cosmochim. Acta. 45:341-351. KATZENBERG M. A. 1989. Stable isotope analysis of archaeological faunal remains from Southern Onta-no. Journal ofArchaelogical Science 16. 319-329. KRUEGER H. W. and SULLIVAN C. H. 1984. Models for carbon isotope fractionation between diet and bone. In J. R. Turnlund and P. E.Johnson (eds.), Stable Isotopes in Nutrition. American Societ)Symfx>-sium Series: 205-220. MINAGAWA M. and WADA E. 1984. Stepwise enrichment of i*N along food chains: further evidence and the relation between 5 ''N in animal alge. Geochim. Cosmochim. Acta. 48: 1135-1140. OWENS N. 1987. Natural variations in "N in the marine environment. Advances in Marine Biology 24: 389-451. PATE F. D. 1995. Stable carhon isotope assessment of hunter-gatherer mobility in prehistoric South Australia./o/inw/ of Archaeological Sciences 22. 81- 87. RICHARDS M. P. and MELLARS P. A. 1998. Stable isotopes and the seasonality of the Oronsay middens. Antiquity 72: 178-184.' SCHWARTCZ H. P. 1991. Some theoretical aspects of isotope paleodiet studies. Journal of Archaeological Sciences 18: 261-275. SCHWARTCZ H. P. and SHOENINGER M. J. 1991. Stable isotope analyses in human nutritional ecology. Year book of Physical Anthropology 34: 283-321. SCHOENINGER M. J. 1985. Trophic level effects in <5N/,4N and UC/'-C ratios in bone collagen and strontium levels in bone mineral. Journal of Human Evolution 14: 515-525. SCHOENINGER M.J. and DeNIRO M.J. 1982. Carbon isotope ratios of apatite from fossil bone cannot be used to reconstruct diets of animals. Nature 297: 577-578. SCHOENINGER M. J. DeNiro M. J. and TAllBER 11. 1983. Stable nitrogen isotope ratios of bone collagen reflect marine and terrestrial components of prehistoric human diet. Science 220. 1381-1383 SCHULT1NG R.J. 1998. Slighting the sea: Stable isotope evidence for the transition to farming in northwestern Europe. In Budja M. (ed.). Doaimenta !*rae-historica XXV, Neolithic studies. 203-219. STRAUS L. G.. ALTL1NA J.. FORD D.. MARAMBAT L.. RHINE J. S„ SCHWARCZ H. P. and VERNETJ.-L 1992. Early farming in the Algarve (Southern Portugal): A preliminary view from two cave excavations near Faro. Trahallios de Antropologia e Etnologia (Porto) 32: 141-162. TAUBF.R II. 1981. I3C evidence for dietary habits of prehistoric man in Denmark. Nature 292. 332-333- VAN DER MERWE N. J. and VOGELJ. C. 1978. l*C content of human collagen as a measure of prehistoric diet in Woodland North America. Nature 276. 815-816. WADA E„ MINAGAWA M., M1ZUTAN1 H„ TSUJ1 T., IMAIZUM1 R. and KARASAWA K. 1987. Biogeochemi-cal studies on the transport of organic matter along the Otsuchi river watershed, lapan. Estuarine Coastal Shelf Sci. 25:321-336. 200