Metal exposure and biological responses in resident and transplanted blue mussels (Mytilus edulis) from the Scheldt estuary

The Western Scheldt river and estuary is known to be highly polluted as it receives industrial, agricultural and domestic effluents from one of the most populated and industrialised zones in Europe. Aquatic organisms are exposed to pollutants, specifically metals that are present in different environmental phases, e.g. dissolved, suspended material or sediment phases. The objective of this study was to study the relationship that exists between environmental metal levels, the degree of metal uptake by aquatic organisms with the concomitant biological responses. For this purpose the bivalve mollusk, Mytilus edulis, was selected as bioaccumulation indicator organism. Environmental (water and sediment) and mussel samples were collected during the late winter (March 2000) from sampling sites in the Scheldt estuary. Sites were selected to represent a salinity gradient from the mouth of the estuary to the furthest distribution area of mussels upstream in the system. Together with standard water quality parameters (e.g. salinity, dissolved oxygen, dissolved organic carbon, etc.) concentrations of twelve metals were analysed in the water (dissolved and suspended matter) and sediments. Levels of these metals were also measured in the soft tissue of M. edulis, together with concomitant biomarker responses in resident mussel populations at each site. The biomarkers that were included in this study were condition index, scope for growth, survival in air, cell membrane stability, DNA damage, and metallothioneins. Data were subjected to multivariate statistical analysis. The physico-chemical parameters and metals in the environmental samples clustered the sites to reflect the distribution based on the salinity gradient. Bioaccumulation results revealed increased metal uptake along a pollution gradient with the highest metal bioaccumulation occurring at the upstream most sites and therefore closest to the in the industrial activities. However, the biomarker responses clustered the sites in a manner that reflected the influence of combination of internal exposure (bioaccumulation) and external exposure (physico-chemical conditions). These differences in biomarker responses clearly demonstrated were attributed to abiotic factors other than metal pollution alone e.g. localized short-term increases in increased suspended sediment concentrations and decreased dissolved oxygen concentrations.     

Multivariate discriminant analysis distinguishes metal- from non metal-related biomarker responses in the clam Chamaelea gallina

Molecular biomarkers are among the most sensitive and earliest responses to pollutants. However, lack of detailed knowledge on variability of responses and their possible seasonal variation limit their use. In addition, the seasonality of biological processes modulates the response of organisms to pollutant stressors. Using multivariate statistics, we have studied the influence of environmental and biological factors on the response of a battery of molecular biomarkers in the clam Chamaelea gallina collected along the South-Spanish littoral. Multivariate discriminant analysis clearly distinguished biomarker response between clean and polluted areas, using heavy metals as indicator of pollution. Such differences disappeared when the dataset was normalised for metal content, thus indicating that pollution was the main significant cause of the changes observed between clean and polluted sites. In conclusion, this work shows that, when applying a complete biomarker panel, multivariate statistical tools can be used to discern pollutant- from non pollutant-related responses.     

Seasonal variations of biomarker responses in the marine blue mussel (Mytilus spp.)

Biomarkers, which can detect changes at the biochemical level, have been used for many years as early warning tools in environmental monitoring. In order to distinguish between natural variability and the potential effects of anthropogenic pollution, it is essential to identify background levels and establish the potential influence of abiotic (season, temperature and salinity) and biotic (gametogenesis) factors. In this study, we examined various biomarkers of stress (glutathione S-transferase (GST)), reproduction (vitellin-like proteins) and damage (lipid peroxidation (LPO) and DNA damage (DNA)) in marine mussels (Mytilus spp.) from a known pristine hybrid zone. Levels of all biomarker expression varied between seasons and appeared to be linked to the reproductive cycle. Oxidative stress in winter, with low GST expression and a higher expression of LPO and DNA damage displayed could be explained by low food availability. These data provide vital baseline information for future ecotoxicological and environmental monitoring studies.     

Heavy metal concentrations and biomarkers of oxidative stress in native mussels (Mytilus edulis chilensis) from Beagle Channel coast (Tierra del Fuego, Argentina)

The aim of this study was to evaluate the usefulness of oxidative stress biomarkers of pollution in native mussels Mytilus edulis chilensis from the Beagle Channel. Spatial and seasonal variations of catalase, glutathione-S-transferase and lipid peroxidation in gills and digestive gland were analyzed in relation to environmental parameters, heavy metals in sediment and in tissue. Four sites with anthropogenic impact and a control site were selected and monitored during the four seasons of 2007. We found significant differences among sites in concentrations of dissolved nutrients and heavy metals in sediments, with the highest values recorded at sites with anthropogenic pressure. Different patterns were observed between concentrations of metals in tissues and in sediments suggesting differences in bioavailability. There were also significant differences in biomarker responses among sites, despite the strong seasonal variability. Our results showed relatively moderate levels of pollution in the study area as a result of urban influences.     

Seasonal variations of physiological and cellular biomarkers and their use in the biomonitoring of north Adriatic coastal waters (Croatia)

Lysosomal membrane stability, the content of neutral lipids and lipofuscin as cellular biomarkers were measured in the digestive gland of mussels, Mytilus galloprovincialis Lam., during a one year survey at selected sites along the coast of the northern Adriatic. The ability of mussels to survive in air as a general physiological response was tested at the same sites. Annual changes of sea temperature, salinity, O(2) saturation, pH and chlorophyll a concentration were measured and the effects of these environmental factors on the biomarkers selected for this study were determined. Survival of mussels after aerial exposure was significantly (p < 0.05) reduced in mussels collected from the contaminated harbors of Pula and Rijeka compared to clean control sites. Lysosomal membrane stability of mussels from urban and industrial areas was also lower indicating exposure to different environmental contaminants. At the same sites, the content of neutral lipids and lipofuscin in digestive cells was significantly higher compared to the reference sites, suggesting alterations in lipid metabolism and contaminant-induced peroxidation of cell membranes. The greatest influence of natural environmental factors was observed for the aerial survival of mussels and the neutral lipids and lipofuscin content in the digestive gland, indicating that the annual cycles of biomarker responses should be well understood for a correct interpretation of biomonitoring results.     

Metal exposure and biological responses in resident and transplanted blue mussels (Mytilus edulis) from the Scheldt estuary

The Western Scheldt river and estuary is known to be highly polluted as it receives industrial, agricultural and domestic effluents from one of the most populated and industrialised zones in Europe. Aquatic organisms are exposed to pollutants, specifically metals that are present in different environmental phases, e.g. dissolved, suspended material or sediment phases. The objective of this study was to study the relationship that exists between environmental metal levels, the degree of metal uptake by aquatic organisms with the concomitant biological responses. For this purpose the bivalve mollusk, Mytilus edulis, was selected as bioaccumulation indicator organism. Environmental (water and sediment) and mussel samples were collected during the late winter (March 2000) from sampling sites in the Scheldt estuary. Sites were selected to represent a salinity gradient from the mouth of the estuary to the furthest distribution area of mussels upstream in the system. Together with standard water quality parameters (e.g. salinity, dissolved oxygen, dissolved organic carbon, etc.) concentrations of twelve metals were analysed in the water (dissolved and suspended matter) and sediments. Levels of these metals were also measured in the soft tissue of M. edulis, together with concomitant biomarker responses in resident mussel populations at each site. The biomarkers that were included in this study were condition index, scope for growth, survival in air, cell membrane stability, DNA damage, and metallothioneins. Data were subjected to multivariate statistical analysis. The physico-chemical parameters and metals in the environmental samples clustered the sites to reflect the distribution based on the salinity gradient. Bioaccumulation results revealed increased metal uptake along a pollution gradient with the highest metal bioaccumulation occurring at the upstream most sites and therefore closest to the in the industrial activities. However, the biomarker responses clustered the sites in a manner that reflected the influence of combination of internal exposure (bioaccumulation) and external exposure (physico-chemical conditions). These differences in biomarker responses clearly demonstrated were attributed to abiotic factors other than metal pollution alone e.g. localized short-term increases in increased suspended sediment concentrations and decreased dissolved oxygen concentrations.     

Monitoring biomarkers in fish (Lepidorhombus boscii and Callionymus lyra) from the northern Iberian shelf after the Prestige oil spill

Hepatic biomarker responses were measured in two demersal fish species (Lepidorhombus boscii and Callionymus lyra) from the northern Iberian shelf associated with the massive Prestige oil spill (POS), five months after the accident. The biomarkers selected were 7-ethoxyresorufin-O-deethylase (EROD), glutathione-S-transferase (GST), glutathione reductase (GR), catalase (CAT), and DNA integrity. Interspecies differences and spatial variations in biomarker responses were observed along the shelf. GST, GR and CAT activities were significantly elevated in L. boscii in the most oil impacted area (Finisterre) and positively correlated (p<0.05) with POS tar aggregate densities. The lack of previous data from the area together with the existence of chronic background pollution of the shelf implies that the observed biomarker responses cannot be solely attributed to the petroleum hydrocarbon components of the spilled oil. This first biological effect assessment showed that L. boscii is a potentially suitable target species to be used in future biomonitoring programmes along the northern Iberian shelf.     

Metal exposure and biological responses in resident and transplanted blue mussels (Mytilus edulis) from the Scheldt estuary

The Western Scheldt river and estuary is known to be highly polluted as it receives industrial, agricultural and domestic effluents from one of the most populated and industrialised zones in Europe. Aquatic organisms are exposed to pollutants, specifically metals that are present in different environmental phases, e.g. dissolved, suspended material or sediment phases. The objective of this study was to study the relationship that exists between environmental metal levels, the degree of metal uptake by aquatic organisms with the concomitant biological responses. For this purpose the bivalve mollusk, Mytilus edulis, was selected as bioaccumulation indicator organism. Environmental (water and sediment) and mussel samples were collected during the late winter (March 2000) from sampling sites in the Scheldt estuary. Sites were selected to represent a salinity gradient from the mouth of the estuary to the furthest distribution area of mussels upstream in the system. Together with standard water quality parameters (e.g. salinity, dissolved oxygen, dissolved organic carbon, etc.) concentrations of twelve metals were analysed in the water (dissolved and suspended matter) and sediments. Levels of these metals were also measured in the soft tissue of M. edulis, together with concomitant biomarker responses in resident mussel populations at each site. The biomarkers that were included in this study were condition index, scope for growth, survival in air, cell membrane stability, DNA damage, and metallothioneins. Data were subjected to multivariate statistical analysis. The physico-chemical parameters and metals in the environmental samples clustered the sites to reflect the distribution based on the salinity gradient. Bioaccumulation results revealed increased metal uptake along a pollution gradient with the highest metal bioaccumulation occurring at the upstream most sites and therefore closest to the in the industrial activities. However, the biomarker responses clustered the sites in a manner that reflected the influence of combination of internal exposure (bioaccumulation) and external exposure (physico-chemical conditions). These differences in biomarker responses clearly demonstrated were attributed to abiotic factors other than metal pollution alone e.g. localized short-term increases in increased suspended sediment concentrations and decreased dissolved oxygen concentrations.     

Factors shaping submerged bryophyte communities: A conceptual model for small mountain streams in Germany

Several models explaining species composition of aquatic bryophytes are available for specific regions. However, a more general, conceptual model applicable to a broader range of regions is lacking.We present a conceptual model ranking environmental factors determining submerged bryophyte communities in small mountain streams. It was tested on a dataset of 54 stream sections after removing the effect of stream size and altitude. Species responses were modeled with pH as predictor variable based on 97 stream sites covering six mountain regions all over Germany. Multiple regressions revealed the importance of primary growth factors (light, Ep(CO₂)) and substrate for the total submerged bryophyte coverage.The known distinction of hard- and softwater bryoflora was clearly supported. The floristic composition of headwaters was predominantly determined by the bicarbonate/ionic strength complex. Species response to pH values supported this result and thus our conceptual model. The primary growth resources light, Ep(CO₂) and availability of coarse streambed material explained one third (R_adjusted² = 0.34) of total submerged bryophyte cover. Disturbances, predominantly spates, reduce biomass but do not affect the basic floristic structure.In conclusion, conceptual models and monitoring methods focusing on aquatic bryophytes need to clearly distinguish “aquatic” from “submersed by chance”. All “aquatic bryophytes” found in Germany can also occur at least temporarily at non-submerged sites. Therefore, a distinction between primary growth factors and additional resources is recommended to disentangle factors determining aquatic bryophyte communities.     

Comparison of coral growth and survival under enclosed,semi-natural conditions and in the field

Two closely related scleractinian coral species, Porites cylindrica and Porites rus, were transplanted to two different locations: the natural environment on the reef flat, and culture tanks on land. The use of tanks enabled the regulation of certain environmental factors, and, hence, the evaluation of specific responses of the corals to these factors. For both species, growth and survival were much better in the field than in the land-based tanks most probably due to unrestricted water circulation. Since the two species were subjected to identical experimental treatments, it was possible to distinguish inherent differences between them in terms of responses to external variables. Porites cylindrica was more susceptible than P. rus to predation by corallivores. Predators, as well as grazers, occurred in significant numbers in the field, but not in the land-based tanks. Porites rus, on the other hand, succumbed more readily to overgrowth by macroalgae which thrived in the culture tanks presumably because of significantly higher nutrient levels and the conspicuous absence of grazers. These results have broader ecological implications because of accelerated environmental changes taking place in present-day reefs due to human impact. Major examples are eutrophication and alterations in water circulation which frequently result in sub-optimal conditions for coral survival and growth.     

Relationship between the Earth tidal factor and phase lag of groundwater levels in confined aquifers and the Wenchuan M s8.0 earthquake of 2008

This work focuses on variations of the Earth tidal factor and phase lag derived from groundwater observations before and after major earthquakes. It is based on an analysis of the data from four observational wells at boundaries between distinct active blocks of China mainland. These wells are also situated on several active fault zones and have exhibited considerable responses to the Wenchuan M _s8.0 earthquake of 2008 in China. We collected hourly records of water levels of these wells from 2007 to 2009 and processed these data for analysis. The tidal factors, phase lags, and phase-difference changes of tidal residuals of each well were calculated. We found that when the Wenchuan quake happened, the tidal factors of the 4 wells were changing rapidly, while their phase lags and phase differences of tidal residuals declined swiftly, which may reflect the stress and strain changes of the well-aquifer system during the seismic generation.     

Polar motion influences in the gravity data recorded by superconducting gravimeters

Gravity data stored in the GGP database (GGP-ISDC) are used to study the small gravity variations caused by polar motion. In a first step the dominant tidal signal and the instrumental drift have to be eliminated from the gravity data. In most cases it is sufficient to model the instrumental drift by polynomials of low degree. The resulting non-tidal gravity variations are split up into their main constituents by fitting two sinusoidal waves with periods of 365.25 days (annual wobble) and 432 days (Chandler wobble). In a similar way the gravity effect of the observed polar motion (IERS-Data) is processed. The ratio between the correspondent amplitudes gives the amplitude factors δ of both wobbles.In a more sophisticated model an additional annual wave was included, destined to absorb disturbing influences with annual period (e.g. environmental influences of different origin). The amount of these influences and the success of their elimination are very different at the individual stations.Besides the comparison of the amplitude factors it also was tried to compare the gravity residuals itself. For that purpose the data series recorded at the different stations were transferred to a common reference point (0°E, 45°N). The graph of the stacked data series gives a first impression of the accordance of the data series recorded at the different stations. Since randomly distributed disturbing influences are reduced by the averaging the amplitude factors derived from the mean of the stacked data series are more reliable than the values derived from the data at the individual stations.In the end 12 data series were included in a common processing. Amplitude factors of 1.183 for the annual and 1.168 for the Chandler wobble result with mean errors less than ±0.010 (roughly estimated). Although corrections for environmental influences were not included directly, the additionally fitted annual wave reduced the scatter of the amplitude factors in the annual range considerably. In contrast to that the amplitude factor of the Chandler wobble remains nearly unaffected, confirming the assumption that the disturbing environmental influences do not extend into the period range of the Chandler wobble.     

Daily variation characteristics of CO_2 emission fluxes and contributions of environmental factors in semiarid grassland of Inner Mongolia, China

Soil respiration refers to the process of soil gener-ating and emitting CO2to the atmosphere under the synthetic effect of different environmental factors,which includes mineralization of soil organic matter involved by microorganism and respiration of plant root system and soil animals.The emission of CO2to the atmosphere through soil respiration is the most important link of carbon cycle process of grassland ecosystem,and also the key ecological process of grassland ecosystem exerting effec…     

Phenological Responses to ENSO in the Global Oceans

Phenology relates to the study of timing of periodic events in the life cycle of plants or animals as influenced by environmental conditions and climatic forcing. Phenological metrics provide information essential to quantify variations in the life cycle of these organisms. The metrics also allow us to estimate the speed at which living organisms respond to environmental changes. At the surface of the oceans, microscopic plant cells, so-called phytoplankton, grow and sometimes form blooms, with concentrations reaching up to 100 million cells per litre and extending over many square kilometres. These blooms can have a huge collective impact on ocean colour, because they contain chlorophyll and other auxiliary pigments, making them visible from space. Phytoplankton populations have a high turnover rate and can respond within hours to days to environmental perturbations. This makes them ideal indicators to study the first-level biological response to environmental changes. In the Earth’s climate system, the El Niño–Southern Oscillation (ENSO) dominates large-scale inter-annual variations in environmental conditions. It serves as a natural experiment to study and understand how phytoplankton in the ocean (and hence the organisms at higher trophic levels) respond to climate variability. Here, the ENSO influence on phytoplankton is estimated through variations in chlorophyll concentration, primary production and timings of initiation, peak, termination and duration of the growing period. The phenological variabilities are used to characterise phytoplankton responses to changes in some physical variables: sea surface temperature, sea surface height and wind. It is reported that in oceanic regions experiencing high annual variations in the solar cycle, such as in high latitudes, the influence of ENSO may be readily measured using annual mean anomalies of physical variables. In contrast, in oceanic regions where ENSO modulates a climate system characterised by a seasonal reversal of the wind forcing, such as the monsoon system in the Indian Ocean, phenology-based mean anomalies of physical variables help refine evaluation of the mechanisms driving the biological responses and provide a more comprehensive understanding of the integrated processes.     

Variations in the quasi-static electric field in the near-Earth’s atmosphere during geomagnetic storms in November 2004

The effects of the geomagnetic storms of November 8 and 10, 2004, in variations in the strength and power spectra of the electric field in the near-Earth’s atmosphere in Kamchatka were studied, together with the meteorological and geophysical phenomena observed simultaneously. A sequence of strong solar flares was shown to cause an anomalous increase in air temperature and humidity. This resulted in the excitation of anomalously strong thunderstorm processes in the atmosphere during the storm of November 8 and made it impossible to distinguish the effects associated with cosmic rays on this background. During the storm of November 10, on the background of weak variations in meteorological parameters, an increase in the strength and intensity of power spectra of the electric field on the day before the storm of November 10 was detected; it was followed by an attenuation of these parameters on the date of the storm. These effects were supposed to be associated with the action of cosmic rays on currents of the global electric circuit. It was shown that the influence of the Forbush effect of galactic cosmic rays in the power spectrum of the electric field first of all shows as the amplification of the component with the period T ～ 48 h; in variations in humidity, the effect shows as the amplification of the component with T ～ 24 h. Cause-and-effect relationships between variations in the electric field strength and the horizontal component of the geomagnetic field were shown to be absent both under the conditions of “fair weather” and during the storm of November 10. A diurnal negative-difference atmospheric pressure was detected on the second day after the geomagnetic storms of November 8 and 10.     

A simple osmium post-fixation paraffin-embedment technique to identify lipid accumulation in fish liver using medaka (Oryzias latipes) eggs and eleutheroembryos as lipid rich models

Hepatic lipidosis is a non-specific biomarker of effect from pollution exposure in fish. Fatty liver is often misdiagnosed or overlooked in histological assessments due to the decreasing application of specific fat procedures and stains. For example, ethanol dehydration in standard paraffin processing removes lipids, leaving vacuoles of which the precise nature is unknown. Lipids can be identified using osmium post-fixation in semi-thin resin sections or transmission electron microscopy. However, both are expensive and technically demanding procedures, often not available for routine environmental risk assessment and monitoring programs. The current emphasis to reduce and refine animal toxicity testing, requires refinement of the suite of histopathological techniques currently available to maximize information gained from using fish for toxicity testing and as bio-indicators of environmental quality. This investigation has successfully modified an osmium post-fixation technique to conserve lipids in paraffin-embedded tissues using medaka (Oryzias latipes) eleutheroembryos and eggs (embryos) as lipid rich models.     

Algal bioassays: their significance in monitoring water quality with respect to nutrients and toxicants

The review focuses on the use of primary producers as a biological tool for evaluating the impact of damage by human activity (eutrophication, toxicity) on the aquatic environment. Studies are discussed following a reductionist approach by using algal bioassays (Selenastrum capricornutum). Variations of algal growth potential (AGP) within watersheds show the impact of human activities such as agriculture and urbanization, on water quality. The study of variation of the AGP in time allowed the investigation of the effect of abiotic (temperature, flow rate) and biotic factors (indigenous primary production) on the concentration of nutrients potentially available to phytoplankton. Seasonal changes of the AGP have further shown the impact of non-point (runoff) or point sources (sewage effluents) on the aquatic system. A staggered relationship was observed between AGP values and chlorophyll a content of indigenous phytoplankton. Values obtained in the laboratory by means of this type of approach would therefore appear to be transferably to natural systems. S. capricornutum was also used to identify toxic characteristics of substances (in pure form or used in formulations) and effluents released into the environment. It was shown that the user of bioassays should use care when evaluating results from tests requiring pretreatment such as storage, autoclaving and filtration. Cautious interpretation is also recommended in order to distinguish between effects of growth-limiting essential elements and the presence of toxic substances. In general it appears from this review that the AGP provides helpful information for a sound management of the aquatic environment.     

Integrated use of biomarkers in the mussel Mytilus galloprovincialis for assessing off-shore gas platforms in the Adriatic Sea: results of a two-year biomonitoring program

Despite a large number of gas platforms existing in the Adriatic Sea, which is a semi-enclosed basin characterized by a slow turnover rate and increasing industrial as well as other anthropogenic activities, the effects of these structures on the aquatic ecosystem require further investigation. Since 1998, multidisciplinary studies have been performed by CNR-ISMAR to comply with legislation and to support the development of protocols for the monitoring of offshore activities in the Adriatic Sea. The present study was developed to implement a biomonitoring plan to assess the ecotoxicological effects of the extraction activities of an off-shore gas platform. Biomarkers were evaluated in mussels collected from the platform in relation to physiological stress, DNA damage, cellular damage, oxidative stress and exposure effects. Organic contaminants and trace element bioaccumulation were also assessed in the soft body of the mussels to correlate bioaccumulation of pollutants with biomarker responses. The results indicate an absence of platform-related environmental stress.     

First evidence of altered vitellogenin-like protein levels in clam Tapes philippinarum and in cockle Cerastoderma glaucum from the Lagoon of Venice

Possible xenoestrogenic effects were investigated, for the first time, in two bivalve species from the Lagoon of Venice (Italy): the clam Tapes philippinarum and the cockle Cerastoderma glaucum. Bivalves were collected far from their reproductive phase at the very early stage of gametogenesis (January), and in the pre-spawning period (June) in six sites. Vitellogenin (Vg)-like proteins (a biomarker of exposure to estrogenic compounds) were measured by the alkali-labile phosphate method (ALP), in both haemolymph and digestive gland from males and females (when it was possible to distinguish sex by microscopic observation of gonadal tissue). Haemolymph calcium concentrations (a parameter considered closely related to the presence of Vg-like proteins) and the bivalve condition index (CI) were also measured. In both seasons, bivalves collected at Campalto (near a sewage treatment plant) and Marghera (a highly contaminated area) had higher Vg-like protein levels, particularly in haemolymph, than animals from the other sampling sites. Interestingly, CI had high values in these polluted sites. In June only, Vg-like proteins and Ca²⁺ levels in haemolymph exhibited similar trends in both bivalve species at most sampling sites. The responsiveness of bivalves to environmental xenoestrogens was higher in June, allowing better discrimination among sites. The present study demonstrates that animals from highly polluted areas have increased Vg-like protein levels. As endocrine disruption due to exposure to estrogenic compounds may cause fertility reduction, alterations in the sex ratio, and a decrease in reproductive rate, a condition of potential risk for bivalve populations in estuarine areas is highlighted.     

A statistical assessment on the stochastic relationship between biomarker concentrations and environmental exposures

Biological monitoring has gradually developed into a powerful tool in the identification and quantification of exposures to occupational and/or environmental hazards in environmental and occupational health studies. Aggregate individual exposure to pollutants and evidence for exploring dose-response relationship in the human bodies can be assessed through biomarker measurements. The existence of inter-individual differences among a study population, however, often hampers the relationship assessment between exposure and the biomarker. In this paper, a statistical random effects model identified from a simplified one-compartment pharmacokinetic model is applied to establish the dynamic relationship between a biomarker and its corresponding external exposure. This model avoids the complex parameter estimation problem encountered using a physiologically based pharmacokinetic (PBPK) model, and incorporates inter-individual variations often ignored in the usual regression approach. In addition, the relevant parameters for the generic kinetic process can be estimated directly. As a guideline for preliminary sampling strategy, tables of required sample sizes and the number of repeated measurements to achieve the desired statistical power and test efficiency are given. The currently established biological exposure indices (BEIs) for benzene and methyl chloroform are employed to illustrate the impact of inter- individual variations on the percentages of protection for workers exposed to the threshold limit value (TLV) of the corresponding chemical.This research was partially supported by the Environmental Protection Agency of Taiwan (Contract No. EPA-88-E102–03–003).