Dating the Paleolithic: Trapped charge methods and amino acid geochronology.

Despite the vast array of different geochronological tools available, dating the Paleolithic remains one of the discipline's greatest challenges. This review focuses on two different dating approaches: trapped charge and amino acid geochronology. While differing in their fundamental principles, both exploit time-dependent changes in signals found within crystals to generate a chronology for the material dated and hence, the associated deposits. Within each method, there is a diverse range of signals that can be analyzed, each covering different time ranges, applicable to different materials and suitable for different paleoenvironmental and archaeological contexts. This multiplicity of signals can at first sight appear confusing, but it is a fundamental strength of the techniques, allowing internal checks for consistency and providing more information than simply a chronology. For each technique, we present an overview of the basis for the time-dependent signals and the types of material that can be analyzed, with examples of their archaeological application, as well as their future potential.

Estimating the dwarfing rate of an extinct Sicilian elephant.

Evolution on islands, together with the often extreme phenotypic changes associated with it, has attracted much interest from evolutionary biologists. However, measuring the rate of change of phenotypic traits of extinct animals can be challenging, in part due to the incompleteness of the fossil record. Here, we use combined molecular and fossil evidence to define the minimum and maximum rate of dwarfing in an extinct Mediterranean dwarf elephant from Puntali Cave (Sicily).1 Despite the challenges associated with recovering ancient DNA from warm climates,2 we successfully retrieved a mitogenome from a sample with an estimated age between 175,500 and 50,000 years. Our results suggest that this specific Sicilian elephant lineage evolved from one of the largest terrestrial mammals that ever lived3 to an island species weighing less than 20% of its original mass with an estimated mass reduction between 0.74 and 200.95 kg and height reduction between 0.15 and 41.49 mm per generation. We show that combining ancient DNA with paleontological and geochronological evidence can constrain the timing of phenotypic changes with greater accuracy than could be achieved using any source of evidence in isolation.

Ionisation bias undermines the use of matrix-assisted laser desorption/ionisation for estimating peptide deamidation: Synthetic peptide studies demonstrate electrospray ionisation gives more reliable response ratios.

RATIONALE: Although mass spectrometry (MS) is routinely used to determine deamination in peptide mixtures, the effects of the choice of ionisation source have not yet been investigated. In particular, matrix-assisted laser desorption/ionisation (MALDI) has become a popular tool with which to measure levels of glutamine deamidation in ancient proteins. Here we use model synthetic peptides to rigorously compare MALDI and electrospray ionisation (ESI). METHODS: We used two synthetic peptides, with glutamine (Q) in one substituted for glutamic acid (E) in the other, to investigate the suitability of MALDI and ESI sources for the assessment of deamidation in peptides using MS. We also compared measurements of the same Q- and E-containing peptide mixtures using two different mass analysers (time-of-flight (TOF) and Fourier transform ion cyclotron resonance (FT-ICR)). RESULTS: When standard mixtures of the Q- and E-containing peptides were analysed using MALDI, under-representation of the E-containing peptide was observed. This observation was consistent between analyses carried out using either TOF or FT-ICR-MS. When the same mixtures were analysed using ESI FT-ICR-MS, no ionisation bias was observed. CONCLUSIONS: MALDI may not be a suitable ionisation method for the determination of deamidation in peptide mixtures. However, ESI was successfully used to determine the ratio in known mixtures of Q- and E-containing peptides. These preliminary observations warrant further investigation into ionisation bias when measuring deamidation in other peptide sequences.

Collagen proteins exchange oxygen with demineralisation and gelatinisation reagents and also with atmospheric moisture.

RATIONALE: The oxygen (O) isotope composition of collagen proteins is a potential indicator of adult residential location, useful for provenancing in ecology, archaeology and forensics. In acidic solution, proteins can exchange O from carboxylic acid moieties with reagent O. This study investigated whether this exchange occurs during demineralisation and gelatinisation preparation of bone/ivory collagen. METHODS: EDTA and HCl demineralisation or gelatinisation reagents were made up in waters with different δ18 O values, and were used to extract collagen from four skeletal tissue samples. Aliquots of extracted collagen were exposed to two different atmospheric waters, at 120°C and ambient temperature, and subsequently dried in a vacuum oven at 40°C or by freeze drying. Sample δ18 O values were measured by HT-EA pyrolysis/IRMS using a zero-blank autosampler. RESULTS: Collagen samples exchanged O with both reagent waters and atmospheric water, which altered sample δ18 O values. Exchange with reagent waters occurred in all extraction methods, but was greater at lower pH. Damage to the collagen samples during extraction increased O exchange. The nature of exchange of O with atmospheric water depended on the temperature of exposure: kinetic fractionation of O was identified at 120°C but not at ambient temperature. Exchange was difficult to quantify due to the high variability of δ18 O values between experimental replicates. CONCLUSIONS: Studies of δ18 O values in collagen proteins should avoid extraction methods using acidic solutions.

Palaeogenomes of Eurasian straight-tusked elephants challenge the current view of elephant evolution.

The straight-tusked elephants Palaeoloxodon spp. were widespread across Eurasia during the Pleistocene. Phylogenetic reconstructions using morphological traits have grouped them with Asian elephants (Elephas maximus), and many paleontologists place Palaeoloxodon within Elephas. Here, we report the recovery of full mitochondrial genomes from four and partial nuclear genomes from two P. antiquus fossils. These fossils were collected at two sites in Germany, Neumark-Nord and Weimar-Ehringsdorf, and likely date to interglacial periods ~120 and ~244 thousand years ago, respectively. Unexpectedly, nuclear and mitochondrial DNA analyses suggest that P. antiquus was a close relative of extant African forest elephants (Loxodonta cyclotis). Species previously referred to Palaeoloxodon are thus most parsimoniously explained as having diverged from the lineage of Loxodonta, indicating that Loxodonta has not been constrained to Africa. Our results demonstrate that the current picture of elephant evolution is in need of substantial revision.

Dietary modulation of cortical excitation and inhibition.

The balance of excitatory and inhibitory neurotransmitters in the brain affects both neural responses and behaviour in humans and animals. Here we investigated whether dietary intervention aimed at increasing levels of the inhibitory neurotransmitter gamma-aminobutyric acid (GABA) can influence neural responses to basic sensory stimuli. Using a steady-state electroencephalography (EEG) paradigm, we found that the neural response to visual patterns was reduced in individuals who consumed a yeast extract product rich in substances associated with the production of GABA (glutamate and B vitamins), but not in a control group who consumed a placebo substance ( n = 14 per group). This demonstrates that the balance of excitation and inhibition in the brain can be influenced by dietary interventions, suggesting possible clinical benefits in conditions (e.g. epilepsy) where inhibition is abnormal.

Lessons from Star Carr on the vulnerability of organic archaeological remains to environmental change.

Examples of wetland deposits can be found across the globe and are known for preserving organic archaeological and environmental remains that are vitally important to our understanding of past human-environment interactions. The Mesolithic site of Star Carr (Yorkshire, United Kingdom) represents one of the most influential archives of human response to the changing climate at the end of the last glacial in Northern Europe. A hallmark of the site since its discovery in 1948 has been the exceptional preservation of its organic remains. Disturbingly, recent excavations have suggested that the geochemistry of the site is no longer conducive to such remarkable survival of organic archaeological and environmental materials. Microcosm (laboratory-based) burial experiments have been undertaken, alongside analysis of artifacts excavated from the site, to assess the effect of these geochemical changes on the remaining archaeological material. By applying a suite of macroscopic and molecular analyses, we demonstrate that the geochemical changes at Star Carr are contributing to the inexorable and rapid loss of valuable archaeological and paleoenvironmental information. Our findings have global implications for other wetland sites, particularly archaeological sites preserved in situ.

Provenancing Archaeological Wool Textiles from Medieval Northern Europe by Light Stable Isotope Analysis (δ13C, δ15N, δ2H).

We investigate the origin of archaeological wool textiles preserved by anoxic waterlogging from seven medieval archaeological deposits in north-western Europe (c. 700-1600 AD), using geospatial patterning in carbon (δ13C), nitrogen (δ15N) and non-exchangeable hydrogen (δ2H) composition of modern and ancient sheep proteins. δ13C, δ15N and δ2H values from archaeological wool keratin (n = 83) and bone collagen (n = 59) from four sites were interpreted with reference to the composition of modern sheep wool from the same regions. The isotopic composition of wool and bone collagen samples clustered strongly by settlement; inter-regional relationships were largely parallel in modern and ancient samples, though landscape change was also significant. Degradation in archaeological wool samples, examined by elemental and amino acid composition, was greater in samples from Iceland (Reykholt) than in samples from north-east England (York, Newcastle) or northern Germany (Hessens). A nominal assignment approach was used to classify textiles into local/non-local at each site, based on maximal estimates of isotopic variability in modern sheep wool. Light element stable isotope analysis provided new insights into the origins of wool textiles, and demonstrates that isotopic provenancing of keratin preserved in anoxic waterlogged contexts is feasible. We also demonstrate the utility of δ2H analysis to understand the location of origin of archaeological protein samples.

Biomineralisation by earthworms - an investigation into the stability and distribution of amorphous calcium carbonate.

BACKGROUND: Many biominerals form from amorphous calcium carbonate (ACC), but this phase is highly unstable when synthesised in its pure form inorganically. Several species of earthworm secrete calcium carbonate granules which contain highly stable ACC. We analysed the milky fluid from which granules form and solid granules for amino acid (by liquid chromatography) and functional group (by Fourier transform infrared (FTIR) spectroscopy) compositions. Granule elemental composition was determined using inductively coupled plasma-optical emission spectroscopy (ICP-OES) and electron microprobe analysis (EMPA). Mass of ACC present in solid granules was quantified using FTIR and compared to granule elemental and amino acid compositions. Bulk analysis of granules was of powdered bulk material. Spatially resolved analysis was of thin sections of granules using synchrotron-based µ-FTIR and EMPA electron microprobe analysis. RESULTS: The milky fluid from which granules form is amino acid-rich (≤ 136 ± 3 nmol mg-1 (n = 3; ± std dev) per individual amino acid); the CaCO3 phase present is ACC. Even four years after production, granules contain ACC. No correlation exists between mass of ACC present and granule elemental composition. Granule amino acid concentrations correlate well with ACC content (r ≥ 0.7, p ≤ 0.05) consistent with a role for amino acids (or the proteins they make up) in ACC stabilisation. Intra-granule variation in ACC (RSD = 16%) and amino acid concentration (RSD = 22-35%) was high for granules produced by the same earthworm. Maps of ACC distribution produced using synchrotron-based µ-FTIR mapping of granule thin sections and the relative intensity of the ν2: ν4 peak ratio, cluster analysis and component regression using ACC and calcite standards showed similar spatial distributions of likely ACC-rich and calcite-rich areas. We could not identify organic peaks in the µ-FTIR spectra and thus could not determine whether ACC-rich domains also had relatively high amino acid concentrations. No correlation exists between ACC distribution and elemental concentrations determined by EMPA. CONCLUSIONS: ACC present in earthworm CaCO3 granules is highly stable. Our results suggest a role for amino acids (or proteins) in this stability. We see no evidence for stabilisation of ACC by incorporation of inorganic components. Graphical abstractSynchrotron-based µ-FTIR mapping was used to determine the spatial distribution of amorphous calcium carbonate in earthworm-produced CaCO3 granules.

An aminostratigraphy for the British Quaternary based on Bithynia opercula.

Aminostratigraphies of Quaternary non-marine deposits in Europe have been previously based on the racemization of a single amino acid in aragonitic shells from land and freshwater molluscs. The value of analysing multiple amino acids from the opercula of the freshwater gastropod Bithynia, which are composed of calcite, has been demonstrated. The protocol used for the isolation of intra-crystalline proteins from shells has been applied to these calcitic opercula, which have been shown to more closely approximate a closed system for indigenous protein residues. Original amino acids are even preserved in bithyniid opercula from the Eocene, showing persistence of indigenous organics for over 30 million years. Geochronological data from opercula are superior to those from shells in two respects: first, in showing less natural variability, and second, in the far better preservation of the intra-crystalline proteins, possibly resulting from the greater stability of calcite. These features allow greater temporal resolution and an extension of the dating range beyond the early Middle Pleistocene. Here we provide full details of the analyses for 480 samples from 100 horizons (75 sites), ranging from Late Pliocene to modern. These show that the dating technique is applicable to the entire Quaternary. Data are provided from all the stratotypes from British stages to have yielded opercula, which are shown to be clearly separable using this revised method. Further checks on the data are provided by reference to other type-sites for different stages (including some not formally defined). Additional tests are provided by sites with independent geochronology, or which can be associated with a terrace stratigraphy or biostratigraphy. This new aminostratigraphy for the non-marine Quaternary deposits of southern Britain provides a framework for understanding the regional geological and archaeological record. Comparison with reference to sites yielding independent geochronology, in combination with other lines of evidence, allows tentative correlation with the marine oxygen isotope record.

Assessing amino acid racemization variability in coral intra-crystalline protein for geochronological applications.

Over 500 Free Amino Acid (FAA) and corresponding Total Hydrolysed Amino Acid (THAA) analyses were completed from eight independently-dated, multi-century coral cores of massive Porites sp. colonies. This dataset allows us to re-evaluate the application of amino acid racemization (AAR) for dating late Holocene coral material, 20 years after Goodfriend et al. (GCA56 (1992), 3847) first showed AAR had promise for developing chronologies in coral cores. This re-assessment incorporates recent method improvements, including measurement by RP-HPLC, new quality control approaches (e.g. sampling and sub-sampling protocols, statistically-based data screening criteria), and cleaning steps to isolate the intra-crystalline skeletal protein. We show that the removal of the extra-crystalline contaminants and matrix protein is the most critical step for reproducible results and recommend a protocol of bleaching samples in NaOCl for 48 h to maximise removal of open system proteins while minimising the induced racemization. We demonstrate that AAR follows closed system behaviour in the intra-crystalline fraction of the coral skeletal proteins. Our study is the first to assess the natural variability in intra-crystalline AAR between colonies, and we use coral cores taken from the Great Barrier Reef, Australia, and Jarvis Island in the equatorial Pacific to explore variability associated with different environmental conditions and thermal histories. Chronologies were developed from THAA Asx D/L, Ala D/L, Glx D/L and FAA Asx D/L for each core and least squares Monte Carlo modelling applied in order to quantify uncertainty of AAR age determinations and assess the level of dating resolution possible over the last 5 centuries. AAR within colonies follow consistent stratigraphic aging. However, there are systematic differences in rates between the colonies, which would preclude direct comparison from one colony to another for accurate age estimation. When AAR age models are developed from a combined dataset to include this natural inter-colony variability THAA Asx D/L, Glx D/L and Ala D/L give a 2σ age uncertainty of ±19, ±38 and ±29 year, for the 20th C respectively; in comparison 2σ age uncertainties from a single colony are ±12, ±12 and ±14 year. This is the first demonstration of FAA D/L for dating coral and following strict protocols 2σ precisions of ±24 years can be achieved across different colonies in samples from the last 150 years, and can be ±10 years within a core from a single colony. Despite these relatively large error estimates, AAR would be a valuable tool in situations where a large number of samples need to be screened rapidly and cheaply (e.g. identifying material from mixed populations in beach or uplift deposits), prior to and complementing the more time-consuming geochronological tools of U/Th or seasonal isotopic timeseries.

A chronological framework for the British Quaternary based on Bithynia opercula.

Marine and ice-core records show that the Earth has experienced a succession of glacials and interglacials during the Quaternary (last ∼2.6 million years), although it is often difficult to correlate fragmentary terrestrial records with specific cycles. Aminostratigraphy is a method potentially able to link terrestrial sequences to the marine isotope stages (MIS) of the deep-sea record. We have used new methods of extraction and analysis of amino acids, preserved within the calcitic opercula of the freshwater gastropod Bithynia, to provide the most comprehensive data set for the British Pleistocene based on a single dating technique. A total of 470 opercula from 74 sites spanning the entire Quaternary are ranked in order of relative age based on the extent of protein degradation, using aspartic acid/asparagine (Asx), glutamic acid/glutamine (Glx), serine (Ser), alanine (Ala) and valine (Val). This new aminostratigraphy is consistent with the stratigraphical relationships of stratotypes, sites with independent geochronology, biostratigraphy and terrace stratigraphy. The method corroborates the existence of four interglacial stages between the Anglian (MIS 12) and the Holocene in the terrestrial succession. It establishes human occupation of Britain in most interglacial stages after MIS 15, but supports the notion of human absence during the Last Interglacial (MIS 5e). Suspicions that the treeless 'optimum of the Upton Warren interstadial' at Isleworth pre-dates MIS 3 are confirmed. This new aminostratigraphy provides a robust framework against which climatic, biostratigraphical and archaeological models can be tested.

Direct terrestrial-marine correlation demonstrates surprisingly late onset of the last interglacial in central Europe.

An interdisciplinary study of a small sedimentary basin at Neumark Nord 2 (NN2), Germany, has yielded a high-resolution record of the palaeomagnetic Blake Event, which we are able to place at the early part of the last interglacial pollen sequence documented from the same section. We use this data to calculate the duration of this stratigraphically important event at 3400 ± 350 yr. More importantly, the Neumark Nord 2 data enables precise terrestrial-marine correlation for the Eemian stage in central Europe. This shows a remarkably large time lag of ca. 5000 yr between the MIS 5e 'peak' in the marine record and the start of the last interglacial in this region.

Is amino acid racemization a useful tool for screening for ancient DNA in bone?

Many rare and valuable ancient specimens now carry the scars of ancient DNA research, as questions of population genetics and phylogeography require larger sample sets. This fuels the demand for reliable techniques to screen for DNA preservation prior to destructive sampling. Only one such technique has been widely adopted: the extent of aspartic acid racemization (AAR). The kinetics of AAR are believed to be similar to the rate of DNA depurination and therefore a good measure of the likelihood of DNA survival. Moreover, AAR analysis is only minimally destructive. We report the first comprehensive test of AAR using 91 bone and teeth samples from temperate and high-latitude sites that were analysed for DNA. While the AAR range of all specimens was low (0.02-0.17), no correlation was found between the extent of AAR and DNA amplification success. Additional heating experiments and surveys of the literature indicated that d/l Asx is low in bones until almost all the collagen is lost. This is because aspartic acid is retained in the bone within the constrained environment of the collagen triple helix, where it cannot racemize for steric reasons. Only if the helix denatures to soluble gelatin can Asx racemize readily, but this soluble gelatine is readily lost in most burial environments. We conclude that Asx d/l is not a useful screening technique for ancient DNA from bone.

Preservation of ancient DNA in thermally damaged archaeological bone.

Evolutionary biologists are increasingly relying on ancient DNA from archaeological animal bones to study processes such as domestication and population dispersals. As many animal bones found on archaeological sites are likely to have been cooked, the potential for DNA preservation must be carefully considered to maximise the chance of amplification success. Here, we assess the preservation of mitochondrial DNA in a medieval cattle bone assemblage from Coppergate, York, UK. These bones have variable degrees of thermal alterations to bone collagen fibrils, indicative of cooking. Our results show that DNA preservation is not reliant on the presence of intact collagen fibrils. In fact, a greater number of template molecules could be extracted from bones with damaged collagen. We conclude that moderate heating of bone may enhance the retention of DNA fragments. Our results also indicate that ancient DNA preservation is highly variable, even within a relatively recent assemblage from contexts conducive to organic preservation, and that diagenetic parameters based on protein diagenesis are not always useful for predicting ancient DNA survival.

Differentiation of MIS 9 and MIS 11 in the continental record: vegetational, faunal, aminostratigraphic and sea-level evidence from coastal sites in Essex, UK.

Multidisciplinary investigations of the vegetational, faunal and sea-level history inferred from the infills of buried channels on the coast of eastern Essex have a direct bearing on the differentiation of MIS 11 and MIS 9 in continental records. New data are presented from Cudmore Grove, an important site on Mersea Island that can be linked to the terrace sequence of the River Thames. The vegetational history has been reconstructed from a pollen sequence covering much of the interglacial represented. The temperate nature of the climate is apparent from a range of fossil groups, including plant remains, vertebrates (especially the rich herpetofauna), molluscs and beetles, which all have strong thermophilous components. The beetle data have been used to derive a Mutual Climatic Range reconstruction, suggesting that mean July temperatures were about 2 degrees C warmer than modern values for southeast England, whereas mean January temperatures may have been slightly colder. The sea-level history has been reconstructed from the molluscs, ostracods and especially the diatoms, which indicate that the marine transgression occurred considerably earlier in the interglacial cycle than at the neighbouring Hoxnian site at Clacton. There are a number of palynological similarities between the sequence at Cudmore Grove and Clacton, especially the presence of Abies and the occurrence of Azolla filiculoides megaspores. Moreover, both sites have yielded Palaeolithic archaeology, indeed the latter is the type site of the Clactonian (flake-and-core) industry. However, the sites can be differentiated on the basis of mammalian biostratigraphy, new aminostratigraphic data, as well as the differences in the sea-level history. The combined evidence suggests that the infill of the channel at Cudmore Grove accumulated during MIS 9, whereas the deposits at Clacton formed during MIS 11. The infill of a much later channel, yielding non-marine molluscs and vertebrates including Hippopotamus, appears to have formed during the Ipswichian (MIS 5e). This evidence is compared with other important sites of late Middle Pleistocene age in Britain and elsewhere on the continent and the importance of a multidisciplinary approach is stressed.

A new approach to amino acid racemization in enamel: testing of a less destructive sampling methodology.

Aspartic acid racemization has been found to be an accurate measure of age at death for recent forensic material. This paper examines the practicality of using acid etching of the tooth surface to extract amino acids from the enamel for racemization analysis. By serial etching of the tooth and contamination of the teeth with bovine serum albumin prior to etching, the ability of etching to remove contamination was assessed. The destructiveness of the method was visualized and quantified using micro-computed tomography (micro-CT). By bleaching the teeth and by deeper etching it was possible to obtain more consistent values. While etching had little effect on the enamel at the macroscale, it did have an impact at the microscale. The quantities of enamel removed varied depending upon the tooth morphology, but were not large. Acid etching of enamel thus appears to be a promising new method for extracting proteins for amino acid racemization age estimation noninvasively.

New evidence for complex climate change in MIS 11 from Hoxne, Suffolk, UK.

The climatic signal of Marine Isotope Stage (MIS) 11 is well-documented in marine and ice-sheet isotopic records and is known to comprise at least two major warm episodes with an intervening cool phase. Terrestrial records of MIS 11, though of high resolution, are often fragmentary and their chronology is poorly constrained. However, some notable exceptions include sequences from the maar lakes in France and Tenaghi Philippon in Greece. In the UK, the Hoxnian Interglacial has been considered to correlate with MIS 11. New investigations at Hoxne (Suffolk) provide an opportunity to re-evaluate the terrestrial record of MIS 11. At Hoxne, the type Hoxnian Interglacial sediments are overlain by a post-Hoxnian cold-temperate sequence. The interglacial sediments and the later temperate phase are separated by the so-called 'Arctic Bed' from which cold-climate macroscopic plant and beetle remains have been recovered. The later temperate phase was deposited during an episode of boreal woodland and is associated with the artefacts, a diverse vertebrate fauna and molluscs. New amino acid geochronological data and biostratigraphical considerations suggest that the post-Hoxnian sequence correlates with late substages of MIS 11. The paper further investigates the correlation of the sequence at Hoxne with the palynological sequences found elsewhere in Europe and adjacent offshore areas.

Ancient biomolecules from deep ice cores reveal a forested southern Greenland.

It is difficult to obtain fossil data from the 10% of Earth's terrestrial surface that is covered by thick glaciers and ice sheets, and hence, knowledge of the paleoenvironments of these regions has remained limited. We show that DNA and amino acids from buried organisms can be recovered from the basal sections of deep ice cores, enabling reconstructions of past flora and fauna. We show that high-altitude southern Greenland, currently lying below more than 2 kilometers of ice, was inhabited by a diverse array of conifer trees and insects within the past million years. The results provide direct evidence in support of a forested southern Greenland and suggest that many deep ice cores may contain genetic records of paleoenvironments in their basal sections.

The earliest record of human activity in northern Europe.

The colonization of Eurasia by early humans is a key event after their spread out of Africa, but the nature, timing and ecological context of the earliest human occupation of northwest Europe is uncertain and has been the subject of intense debate. The southern Caucasus was occupied about 1.8 million years (Myr) ago, whereas human remains from Atapuerca-TD6, Spain (more than 780 kyr ago) and Ceprano, Italy (about 800 kyr ago) show that early Homo had dispersed to the Mediterranean hinterland before the Brunhes-Matuyama magnetic polarity reversal (780 kyr ago). Until now, the earliest uncontested artefacts from northern Europe were much younger, suggesting that humans were unable to colonize northern latitudes until about 500 kyr ago. Here we report flint artefacts from the Cromer Forest-bed Formation at Pakefield (52 degrees N), Suffolk, UK, from an interglacial sequence yielding a diverse range of plant and animal fossils. Event and lithostratigraphy, palaeomagnetism, amino acid geochronology and biostratigraphy indicate that the artefacts date to the early part of the Brunhes Chron (about 700 kyr ago) and thus represent the earliest unequivocal evidence for human presence north of the Alps.