Evaluating manufacturing process profile comparability with multivariate equivalence testing: Case study of cell‐culture small scale model transfer

This article studies the Generalized Mahalanobis Distance (GMD) approach proposed by Hoffelder which measures the dissimilarity of two multivariate Gaussian distributions with arbitrary covariance matrices and unequal sample sizes. This investigation demonstrated that, with appropriate adjustment, the GMD approach can achieve the targeted nominal Type I error and provide sufficient power for testing equivalence between two profile populations. The adjusted GMD approach was applied to examine the equivalence of harvest profiles between a 12L small scale model and 2000L manufacturing scale in a transfer study performed at Sanofi Specialty Care Framingham Biologics. The harvest profiles were evaluated based on three critical parameters (Productivity, Lactate Production, and Total Cell Density) and deemed practically equivalent using a pre‐defined equivalence margin per the adjusted GMD approach. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 34:187–195, 2018     

Gastroretentive drug delivery system of ranitidine hydrochloride: Formulation and in vitro evaluation

The purpose of this research was to prepare a gastroretentive drug delivery system of ranitidine hydrochloride. Guar gum, xanthan gum, and hydroxypropyl methylcellulose were evaluated for gel-forming properties. Sodium bicarbonate was incorporated as a gas-generating agent. The effects of citric acid and stearic acid on drug release profile and floating properties were investigated. The addition of stearic acid reduces the drug dissolution due to its hydrophobic nature. A 3² full factorial design was applied to systemically optimize the drug release profile. The amounts of citric acid anhydrous (X₁) and stearic acid (X₂) were selected as independent variables. The times required for 50% (t₅₀) and 80% drug dissolution (t₈₀), and the similarity factor f₂ were selected as dependent variables. The results of the full factorial design indicated that a low amount of citric acid and a high amount of stearic acid favors sustained release of ranitidine hydrochloride from a gastroretentive formulation. A theoretical dissolution profile was generated using pharmacokinetic parameters of ranitidine hydrochloride. The similarity factor f₂ was applied between the factorial design batches and the theoretical dissolution profile. No significant difference was observed between the desired release profile and batches F2, F3, F6, and F9. Batch F9 showed the highest f2 (f2=75) among all the batches, and this similarity is also reflected in t₅₀ (∼214 minutes) and t₈₀ (∼537 minutes) values. These studies indicate that the proper balance between a release rate enhancer and a release rate retardant can produce a drug dissolution profile similar to a theoretical dissolution profile.     

Seasonal soil‐surface carbon fluxes from the root systems of young Pinus radiata trees growing at ambient and elevated CO2 concentration

Elevated atmospheric CO₂ concentration may result in increased below‐ground carbon allocation by trees, thereby altering soil carbon cycling. Seasonal estimates of soil surface carbon flux were made to determine whether carbon losses from Pinus radiata trees growing at elevated CO₂ concentration were higher than those at ambient CO₂ concentration, and whether this was related to increased fine root growth. Monthly soil surface carbon flux density (f) measurements were made on plots with trees growing at ambient (350) and elevated (650 μmol mol^?1) CO₂ concentration in large open‐top chambers. Prior to planting the soil carbon concentration (0.1%) and f (0.28 μmol m^?2 s^?1 at 15 °C) were low. A function describing the radial pattern of f with distance from tree stems was used to estimate the annual carbon flux from tree plots. Seasonal estimates of fine root production were made from minirhizotrons and the radial distribution of roots compared with radial measurements of f. A one‐dimensional gas diffusion model was used to estimate f from soil CO₂ concentrations at four depths. For the second year of growth, the annual carbon flux from the plots was 1671 g y^?1 and 1895 g y^?1 at ambient and elevated CO₂ concentrations, respectively, although this was not a significant difference. Higher f at elevated CO₂ concentration was largely explained by increased fine root biomass. Fine root biomass and stem production were both positively related to f. Both root length density and f declined exponentially with distance from the stem, and had similar length scales. Diurnal changes in f were largely explained by changes in soil temperature at a depth of 0.05 m. Ignoring the change of f with increasing distance from tree stems when scaling to a unit ground area basis from measurements with individual trees could result in under‐ or overestimates of soil‐surface carbon fluxes, especially in young stands when fine roots are unevenly distributed.     

Artificial neural networks in the modeling and optimization of aspirin extended release tablets with eudragit L 100 as matrix substance

The purpose of the present study was to model the effects of the concentration of Eudragit L 100 and compression pressure as the most important process and formulation variables on the in vitro release profile of aspirin from matrix tables formulated with Eudragit L 100 as matrix substance and to optimize the formulation by artificial neural network. As model formulations, 10 kinds of aspirin matrix tablets were prepared. The amount of Eudragit L 100 and the compression pressure were selected as causal factors. In vitro dissolution time profiles at 4 different sampling times were chosen as responses. A set of release parameters and causal factors were used as tutorial data for the generalized regression neural, network (GRNN) and analyzed using a computer. Observed results of drug release studies indicate that drug release rates vary widely between investigated formulations, with a range of 5 hours to more than 10 hours to complete dissolution. The GRNN model was optimized. The root mean square value for the trained network was 1.12%, which indicated that the optimal GRNN model was reached. Applying the generalized distance function method, the optimal tablet formulation predicted by GRNN was with 5% of Eudragit L 100 and tablet hardness 60N. Calculated difference (f ₁ 2.465) and similarity (f ₂ 85.61) factors indicate that there is no difference between predicted and experimentally observed drug release profiles for the optimal formulation. This work illustrates the potential for an artificial neural network, GRNN, to assist in development of extended release dosage forms.     

Development and Validation of a Discriminating <Emphasis Type="Italic">In Vitro</Emphasis> Dissolution Method for a Poorly Soluble Drug,Olmesartan Medoxomil: Comparison Between Commercial Tablets

A dissolution test for tablets containing 40 mg of olmesartan medoxomil (OLM) was developed and validated using both LC-UV and UV methods. After evaluation of the sink condition, dissolution medium, and stability of the drug, the method was validated using USP apparatus 2, 50 rpm rotation speed, and 900 ml of deaerated H₂O + 0.5% sodium lauryl sulfate (w/v) at pH 6.8 (adjusted with 18% phosphoric acid) as the dissolution medium. The model-independent method using difference factor (f ₁) and similarity factor (f ₂), model-dependent method, and dissolution efficiency were employed to compare dissolution profiles. The kinetic parameters of drug release were also investigated. The obtained results provided adequate dissolution profiles. The developed dissolution test was validated according to international guidelines. Since there is no monograph for this drug in tablets, the dissolution method presented here can be used as a quality control test for OLM in this dosage form, especially in a batch to batch evaluation.     

Evolution of eusociality in diploid species

Haplodiploid species are naturally biased by their genetic structure toward the evolution of sterile worker castes, as shown by W. D. Hamilton (1964. J. Theoret. Biol., 7, 1–16, 17–52). Diploid species do not have this intrinsic genetic bias toward eusociality. Nonetheless, true sociality has evolved in the diploid ancestors of the modern termites, and varying degrees of quasisociality are not uncommon in diploid species, including mammals. A genetic bias toward investment in relatives rather than offspring can arise in a diploid species as a result of inbreeding. The consequences of several regular incestuous breeding systems are analyzed in detail. It is shown that, under certain conditions, there is a natural bias toward an alternation of inbred and outbred generations. As this alternation proceeds, the genetic bias toward eusociality rapidly approaches an asymptotic value of 4(1 + 2f₀)/3(1 + 3f₀), where f₀ is the average coefficient of relationship for the outbreeding pairs. For f₀ close to zero, the genetic bias toward eusociality is close to 1.33, which is even larger than the genetic bias of 1.25 in haplodiploid species. Under other conditions there may be repeated incestuous matings between successive outbreeding generations. In this case the bias toward eusociality can be as large as 2.     

Use of the tetrachoric correlation model in assessing clinical tests

Categorical data on n patients are classified according to the results on an initial clinical test T, and also with respect to a subsequent and definitive diagnosis D. In this paper the sensitivity (=), specificity (=) and predictive values (=_v) are discussed with reference to the tetrachoric correlation model based on the bivariate normal density (e.g. PEARSON, 1901; KENDALL and STUART, 1972, II, p. 317). The result of HAMDAN (1970) concerning the equivalence of the tetrachoric r_t and the maximum likelihood estimate of the correlation coefficient is utilized in this paper to obtain a test of significance concerning r_t, and also the relation between the tetrachoric function w and the relative risk ψ for the 2X2 table.     

Studies on the Effect of Water-Soluble Polymers on Drug–Cyclodextrin Complex Solubility

The effect of complexation of irbesartan (IRB), a practically water-insoluble drug, with cyclodextrins in presence of different concentrations of water-soluble polymers (PEG 4000 and PVP K-90) on the dissolution rate of the drug has been investigated. Phase solubility studies were carried out to evaluate the solubilizing power of βCD in association with water-soluble polymers towards IRB and to determine the apparent stability constant (K _S) of the complexes. Improvement in K _S value for ternary complexes (IRB–βCD–polymers) clearly proved the benefit on the addition of water-soluble polymer to increase complexation efficiency. The dissolution rate of the drug from ternary systems containing PEG 4000 and PVP K-90 was higher as compared to the binary system. An optimum increase in the dissolution rate of the drug was observed at a polymer concentration of 5% w/w for PVP K-90 and 10% w/w for PEG 4000. DSC, FTIR, SEM, and XRD studies were carried out to characterize the complexes.     

The limitations of heart rate as a predictor of metabolic rate in fish

Although telemetered heart rate (f_H) has been used as a physiological correlate to predict the metabolic rate (as oxygen consumption, V?O₂) of fish in the field, it is our contention that the method has not been validated adequately for fish. If f_H in fish is to be used to estimate V?O₂, a single linear (or log-linear) relationship must be established for each species between the two variables which allows V?O₂ to be predicted accurately under all environmentally relevant conditions. Our analyses of existing data indicate that while a good linear (or log-linear) relationship can be established between f_H and V?O₂, the conditions under which the relationship applies may be quite restricted. Physiological states and environmental factors affect the relationship between f_H and V?O₂ significantly such that several curves can exist for a single species. In addition, there are situations in which f_H and V?O₂ do not covary in a significant manner. In some situations f_H can vary over much of its physiological range while V?O₂ remains constant; in others V?O₂ may vary while f_H is invariate. The theoretical basis for this variability is examined to explain why the use of telemetered f_H in predicting V?O₂ of fish may be limited to certain specified applications.     

Chlorophyll a is the crucial redox sensor and transmembrane signal transmitter in the cytochrome b6f complex. Components and mechanisms of state transitions from the hydrophobic mismatch viewpoint

The cytochrome (cyt) b₆f complex is involved in the transmembrane redox signaling that triggers state transitions in cyanobacteria and chloroplasts. However, the components and molecular mechanisms are still unclear. In an attempt to solve this long-standing problem, we first focused on the unknown role of a single chlorophyll a (Chla) in cyt b₆f with a new approach based on Chla structural properties. Various b₆f X-ray crystal structures were analyzed to identify their differences, which correlate with differences in Chla molecular volume. We found that the distance of the Rieske [2Fe-2S] cluster to Chla correlates with the distance between a pair of residues at the Q_o-site and the distance between a pair of residues at the opposite membrane side. These correlations were accompanied by the rotation of a key peripheral residue and by changes in the hydrophobic thickness of cyt b₆f. Parallel analysis of cyt bc₁ crystal structures allowed us to conclude that Chla acts as the crucial redox sensor and transmembrane signal transmitter in b₆f for changes in the plastoquinone pool redox state. The hydrophobic mismatch induced by the changed hydrophobic thickness of cyt b₆f is the driving force for the structural reorganizations of the photosynthetic apparatus during induction and the progression of state transitions in cyanobacteria and chloroplasts. A mechanism for LHCII kinase activation in chloroplasts is also proposed. Our understanding of the dynamic structural changes in bc-complexes during turnover at the Q_o-site and state transitions is augmented by the time-sequence ordering of 56 bc crystal structures.     

Kneading Technique for Preparation of Binary Solid Dispersion of Meloxicam with Poloxamer 188

The aim of the present study was to enhance the dissolution rate of meloxicam (MLX), a practically water-insoluble drug by preparation of solid dispersion using a hydrophilic polymer, poloxamer 188 (PXM). The kneading technique was used to prepare solid dispersions. A 3² full factorial design approach was used for optimization wherein the drug, polymer ratio (X ₁), and the kneading time (X ₂) were selected as independent variables and the dissolution efficiency at 60 min (%DE₆₀) and yield percent were selected as the dependent variable. Multiple linear regression analysis revealed that for obtaining higher dissolution of MLX from PXM solid dispersions, a high level of X ₁ and a high level of X ₂ were suitable. The use of a factorial design approach helped in optimization of the preparation and formulation of solid dispersion. The optimized formula was characterized by solubility studies, angle of repose, and contact angle; Fourier transform infrared spectroscopy, differential scanning calorimetry, x-ray diffraction studies, and scanning electron microscopy demonstrated that enhanced dissolution of MLX from solid dispersion might be due to a decrease in the crystallinity of MLX and PXM. Analysis of dissolution data of optimized formula indicated the best fitting with Korsemeyer–Peppas model and the drug release kinetics as Fickian diffusion. In conclusion, dissolution enhancement of MLX was obtained by preparing its solid dispersion with PXM using kneading technique.     

Inversion of net ecosystem CO2 flux measurements for estimation of canopy PAR absorption

The fractional absorption of photosynthetically active radiation (f_PAR) is frequently a key variable in models describing terrestrial ecosystem–atmosphere interactions, carbon uptake, growth and biogeochemistry. We present a novel approach to the estimation of the fraction of incident photosynthetically active radiation absorbed by the photosynthetic components of a plant canopy (f_Chl). The method uses micrometeorological measurements of CO₂ flux and incident radiation to estimate light response parameters from which canopy structure is deduced. Data from two Ameriflux sites in Oklahoma, a tallgrass prairie site and a wheat site, are used to derive 7‐day moving average estimates of f_Chl during three years (1997–1999). The inverse estimates are compared to long‐term field measurements of PAR absorption. Good correlations are obtained when the field‐measured f_PAR is scaled by an estimate of the green fraction of total leaf area, although the inverse technique tends to be lower in value than the field measurements. The inverse estimates of f_Chl using CO₂ flux measurements are different from measurements of f_PAR that might be made by other, more direct, techniques. However, because the inverse estimates are based on observed canopy CO₂ uptake, they might be considered more biologically relevant than direct measurements that are affected by non‐physiologically active components of the canopy. With the increasing number of eddy covariance sites around the world the technique provides the opportunity to examine seasonal and inter‐annual variation in canopy structure and light harvesting capacity at individual sites. Furthermore, the inverse f_Chl provide a new source of data for development and testing of f_PAR retrieval using remote sensing. New remote sensing algorithms, or adjustments to existing algorithms, might thus become better conditioned to ‘biologically significant’ light absorption than currently possible.     

Evaluation of methods for estimating soil carbon dioxide efflux across a gradient of forest disturbance

Better understanding of variation in soil carbon dioxide (CO₂) efflux caused by measurement techniques is needed, especially over gradients of site disturbance, to accurately estimate the global carbon cycle. We present soil CO₂ efflux data from a gradient of disturbance to ponderosa pine (Pinus ponderosa C. Lawson var. scopulorum Engelm.) forests in northern Arizona, USA that were obtained using four different techniques: vented static chambers, a Licor 6400‐09, and soil CO₂ diffusion profiles using two different models (Moldrup, Millington–Quirk) to estimate soil gas diffusivity. We also compared soil CO₂ efflux measured by the Moldrup and Millington–Quirk diffusion profile methods to nighttime total ecosystem respiration (TER) data from an eddy covariance tower. We addressed four questions: (1) Does the use of a given method to measure soil CO₂ efflux bias results across a disturbance gradient? (2) Does the magnitude of difference between observed and modeled estimates of soil CO₂ differ between methods and across sites? (3) What is the spatial variability of each method at each site? (4) Which method is closest to the estimate of TER measured by the eddy covariance tower? Although soil CO₂ efflux varied significantly among methods the differences were consistent among sites. Measured and modeled total growing season fluxes were generally higher for the Licor 6400‐09 and Millington–Quirk diffusion gradient methods compared with static chamber and the Moldrup diffusion gradient methods. A power analysis showed that the larger static chamber was the most efficient method at sampling spatial variation in soil CO₂ efflux. Nighttime measurements of soil CO₂ efflux from the Moldrup diffusion gradient method were most strongly related to nighttime TER assessed with eddy covariance. The use of a single, well‐implemented method to measure soil CO₂ efflux is unlikely to create bias in comparisons across a gradient of forest disturbance.     

A predictive model for the growth rate of Bacillus cereus in broth by response surface methodology

A response surface methodology (RSM) was developed for predicting the growth rate of Bacillus cereus in a tryptic soy broth medium as a function of temperature (10 to 40°C), pH (5.5 to 8.5), and the NaCl concentration (0 to 8%). The primary model showed a good fit (r² = 0.920 to 0.999) to a Gompertz equation to obtain growth rates each condition. The quadratic polynomial model was found to be significant (p < 0.0001) and predicted values were found to be in good agreement with experimental values (R² value of 0.9486). The evaluation of RSM for describing the growth rate of B. cereus used the bias factor (B_f) and the accuracy factor (A_f). Both the B_f value (1.11) and the A_f value (1.50) were within acceptable ranges. This model was provided an efficient and accurate method for predicting the growth of B. cereus as a function of the controlling factors.     

Studies on Formulation Development of Mucoadhesive Sustained Release Itraconazole Tablet Using Response Surface Methodology

The purpose of this research was to prepare and evaluate sustained release mucoadhesive tablets of Itraconazole. It is practically insoluble in aqueous fluids hence its solid dispersion with Eudragit E100 was prepared by spray drying. This was formulated in matrix of hydrophilic mucoadhesive polymers Carbopol 934P (CP) and Methocel K4M (HPMC). The formulation was optimized using a 3² factorial design. Amounts of CP and HPMC were taken as formulation variables for optimizing response variables i.e. mucoadhesion and dissolution parameters. The optimized mucoadhesive formulation was orally administered to albino rabbits, and blood samples collected were used to determine pharmacokinetic parameters. The solid dispersion markedly enhanced the dissolution rate of itraconazole. The bioadhesive strength of formulation was found to vary linearly with increasing amount of both polymers. Formulations exhibited drug release fitting Peppas model with value of n ranging from 0.61 to 1.18. Optimum combination of polymers was arrived at which provided adequate bioadhesive strength and fairly regulated release profile. The experimental and predicted results for optimum formulations were found to be in close agreement. The formulation showed C _max 1898 ± 75.23 ng/ml, t _max of the formulation was 2 h and AUC was observed to be 28604.9 ng h/ml     

Heart rate and oxygen uptake relationship: a comparison of loaded marching and running in women

Heart rate (beats · min^–1;f _c) measured during marching with a load is often used to predict the oxygen cost (1·min^–1; VO₂) of the activity. The prediction comes from thef _c/VO₂ relationship determined from laboratory measures off _c and VO₂ during treadmill running. Studies in men have suggested that this may not be appropriate although this has yet to be examined in women. This study, therefore, compared thef _c/VO₂ relationship between loaded marching and maximal running protocols in women. Sixteen female subjects [mean (SD), age 21.9 (2.3) years, height 6 (0.06) m, weight 62.6 (7.6) kg] had theirf _c (from three-lead chest electrodes) and VO₂ measured first during standard treadmill run protocols, and again 1 week later during loaded marching protocols. The slopes and intercepts determined from linear regression off _c on VO₂ for each individual for each protocol were compared as were the maximalf _c(f _cmax), VO₂ and ratings of perceived exertion (RPE) from the last work period of each protocol in pairedt-tests. The VO₂ slopes (P < 0.01) and intercepts (P < 0.05) differed significantly between loaded marching and running.f _cmax for loaded marching were 90% off _cmax for running (P < 0.01) and VO₂ for loaded marching were 80% of those for running (P < 0.01). However, RPE at the final levels for the two protocols were not significantly different. The data suggest that in women the VO₂ relationships for loaded marching and for running are different. This difference is similar to that found in men when speed is held constant and the load and gradient are varied. The results suggest that it would be erroneous to usef _c and VO₂ measured during running protocols in the laboratory to estimate energy expenditure and work intensity during loaded marching in the filed.     

Goodness-of-Fit Testing for the Logistic Regression Model when the Estimated Probabilities are Small

The distribution of the Hosmer-Lemeshow chi-square type goodness-of-fit tests (?_g, ?_g) for the logistic regression model are examined via simulations designed to examine their behavior when most of the estimated probabilities are small or are expected to fall in a few deciles. The results of the simulations show statistic ?_g should be used when the two outcome groups (y = 0, 1) are not well separated, Δ≤2, where Δ² is the Mahalanobis distance. Statistic ?_g should be used when Δ ≥ 8. Either statistic may be used when 2 ≦ Δ ≦ 8. All tests should be used with caution when the proportion in the sample with y = 1 is less than 0.1.     

Comparison of biosimilar CT-P10 and innovator rituximab in patients with rheumatoid arthritis: a randomized controlled Phase 3 trial

This multinational, randomized, double-blind trial, (ClinicalTrials.gov identifier NCT02149121) was designed to demonstrate equivalence in pharmacokinetics and efficacy between CT-P10 and innovator rituximab (RTX) in patients with rheumatoid arthritis (RA). Adults with active RA were treated with CT-P10, United States-sourced RTX (US-RTX; Rituxan®), or European Union-sourced RTX (EU-RTX; MabThera®) at weeks 0 and 2. The co-primary pharmacokinetic endpoints were area under the serum concentration–time curve (AUC) from time zero to last measurable concentration (AUC_0–last), AUC from time zero to infinity (AUC_0–∞), and maximum concentration (C_max) after two infusions. The primary efficacy endpoint was change from baseline to week 24 in Disease Activity Score using 28 joints-C-reactive protein (DAS28-CRP). Pharmacodynamics, immunogenicity, and safety were also assessed. 372 patients were randomly assigned to CT-P10 (n = 161) or RTX (n = 211 [US-RTX, n = 151; EU-RTX, n = 60]). For the co-primary pharmacokinetic endpoints, 90% confidence intervals (CI) for ratios of geometric means (CT-P10/US-RTX, CT-P10/EU-RTX or EU-RTX/US-RTX) all fell within the equivalence margin of 80–125%. Adjusted least squares (LS) mean (standard error) change from baseline in DAS28-CRP at week 24 was ?2.13 (0.175) for CT-P10 and ?2.09 (0.176) for RTX. The 95% CI (?0.29, 0.21) of the estimated treatment difference between CT-P10 and RTX (?0.04) was entirely within the efficacy equivalence margin of ±0.5. Pharmacodynamics, immunogenicity, and safety profiles were similar for CT-P10 and RTX. The pharmacokinetics of CT-P10, US-RTX, and EU-RTX were equivalent. CT-P10 and RTX were also equivalent in terms of efficacy and displayed similar pharmacodynamic, immunogenicity, and safety profiles up to week 24.     

A Distribution-Free Two-Sample Equivalence Test Allowing for Tied Observations

A new testing procedure is derived which enables to assess the equivalence of two arbitrary noncontinuous distribution functions from which unrelated samples are taken as the data to be analyzed. The equivalence region is defined to consist of all pairs (F,G) of distribution functions such that for independent X ∼ F, Y ∼ G the conditional probability of {X > Y} given {X ≠ Y} lies in some short interval around 1/2. The test rejects the null hypothesis of nonequivalence if and only if the standardized distance between the U-statistics estimator of P[X > Y ∣ X ≠ Y] and the center of the equivalence interval (1/2 — ε₁, 1/2 + ε₂) does not exceed a critical upper bound which has to be computed as the α-quantile of a χ²-distribution with one degree of freedom and a random noncentrality parameter proportional to the squared length of that interval. The test is shown to maintain the asymptotic significance level under very weak regularity conditions. Results of an extensive simulation study suggest that its level properties are very satisfactory in small samples as well. The power turns out to be inversely related to the rate P[X = Y] of ties between observations from different samples.     

Aerobic fitness and sympatho-adrenal response to short-term psycho-emotional stress under field conditions

A possible relationship between aerobic fitness (AF), measured by maximal cycle ergometry (CE) and sympatho-adrenal response to acute, short lasting psycho-emotional stress was investigated by monitoring heart rate (f _c) and excretion of catecholamines. The activation of the sympatho-adrenal system was characterised by the noradrenaline : adrenaline ratio. A group of 11 healthy men [22.8 (SD 2.52) years] lived under identical environmental conditions; their mean maximal oxygen uptake ( ) was 47.1 (SD 3.9) ml · min^–1 · kg^–1. After the physiological and psychological laboratory tests had been completed thef _c of the subjects was monitored continuously during the guerilla slide and parachute jump by night, two emotionally stressful military tasks. Maximalf _c (f _{c, max}) attained during these events was 84.5% and 83% off _{c, max} during CE (f _{c, max, CE}), respectively. A significant relationship (r=–0.92,P<0.0002) betweenf _{c, max} reached during the stressful tasks and was found only for the guerilla slide, which was preceded by physical strain, sleep deprivation and energy deficit. One subject with some prior experience in parachuting showed the lowestf _c response and the lowest sympatho-adrenal activation in both events, independent of the degree of AF. In conclusion, AF was found to influence the sympatho-adrenal and fc response to acute, short-lasting emotional stress when the stressful event was aggravated by preceding physical strain, the magnitude of the stress response depending largely on individual experience and effective mechanisms for coping with specific stimuli.