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4 Research Needs Mineral inadequacy and depletion in military personnel can come from many sources. Military personnel who participate in training and operational exercises routinely decrease their food intake and, as a result, may consume inadequate amounts of minerals. Even in the absence of reduced food intake, highly- demanding physical activities may result in mineral depletion, because physical and environmental stressors can increase the turnover and losses of minerals. Dietary or blood biochemical markers of nutritional status may not always reveal mineral deficiencies, which may be explained by a lack of sensitive biochemical markers of mineral status, brief durations of restricted intakes, or mineral mobili- zation from stores into the blood with increased metabolic demands and loss of body weight. Also, determining the independent effect of restricted energy in- takes on specific micronutrient impairments is difficult. Throughout this report, the committee stresses the need to establish Military Dietary Reference Intakes specific for military personnel in situations of extreme weather, intense exercise, and other stressors that might alter the nutrient re- quirements for maintaining or improving health and physical and cognitive per- formance. The committee also recognizes that the appropriate data to establish mineral standards for the military are scarce. Thus, there is a great need to conduct controlled studies on mineral nutrition and physical performance, cogni- tive function, and behavior with military personnel in garrison training and in the field while the soldiers are engaged in support, training, and combat and in the context of moderating variables. These variables include gender, body composi- tion, fitness, task, physical demand, extreme environmental conditions, sleep deprivation, food restriction, and psychological stressors (e.g., depression, anxi- ety, fear). For dietary intervention studies, it is critical that nutrient interactions 191

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192 MINERAL REQUIREMENTS FOR MILITARY PERSONNEL are considered in the design and interpretation of results. Studies should be con- trolled for other potentially limiting nutrients. The data from the recommended studies are needed to evaluate critically the adequacy of current rations provided to and consumed by soldiers, and, if needed, will provide a foundation for developing new rations. Such information also may be useful to better understand the determinants of food intake by soldiers (Hirsch and Kramer, 1993). Therefore, the committee urges that efforts be made to an- swer some of these research questions more accurately by generating data from experimental studies that more closely reflect the military environment. This report is concerned with military personnel in garrison training; therefore, the research questions are targeted at soldiers in that environment. However, similar research questions could (and should) be posed about mineral requirements in soldiers who face other extreme field environments, such as those encountered during sustained operations. The committee agreed that ongoing research in a number of nutritional areas is critical and discussed several strategies for addressing unresolved problems relating to military nutrition. In fact, it examined the value of a cohort longitudi- nal study that would collect data on a broad array of nutritional indexes through- out the military careers of a cohort of recruits or enlisted soldiers and determined that even though the study might be of some value its feasibility is questionable, especially considering the tremendous use of resources when the relevance of the data collected is unclear. Also, resources may be tapped for other more pressing needs, making a longitudinal study a long-term, difficult goal to accom- plish. A more useful, reasonable, and economical strategy would be to conduct focused studies with clearer objectives that are more comprehensible to com- manders. In any event, the committee recommends that attention is paid to exist- ing nutrition-related research questions in a manner best suited to military circumstances. ORGANIZATION AND PRIORITIZATION This chapter focuses on research needs--organized according to prioritiza- tion criteria--that would assist the military with answering questions related to mineral requirements for soldiers in garrison training, including design details that might be helpful when developing a research agenda. In the following section on research priorities, the committee describes design details of two overall, cross-cutting studies (i.e., they apply to more than one mineral); the two studies are considered the highest priority, because they will provide criti- cal information regarding mineral losses and mineral status of military person- nel in garrison training. Then, the subsequent sections on specific minerals list and prioritize (the first study is the highest priority, the last is the lowest priority) the most important studies to pursue, according to the strength of the current available evidence. In some of those sections, the committee lists a

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RESEARCH NEEDS 193 category under the heading Other Research Needs; these are studies that address interesting research questions, but for which less evidence has been collected. The committee recognizes the high cost of the studies proposed; therefore, if resources are slim or there are other pressing needs--in addition to addressing the two highest-priority studies--the following research questions for specific minerals should be explored: Does iron supplementation prevent iron deficiency? What is the best strat- egy to prevent iron deficiency? How does physical activity influence calcium requirements? Does iron intake above the levels recommended in this report have ben- eficial effects in cognitive functions? Does magnesium intake above the levels recommended in this report offer protection from sleep deprivation disturbances? Does zinc intake above levels recommended in this report result in im- proved physical and cognitive performance? RESEARCH PRIORITIES 1. Study the Effects of Military Garrison Training on Mineral Losses and Performance A study is needed to assess the effects of environmental, physical, and psychological stressors encountered by military personnel--including heat, physical activity, and possibly sleep restriction--on the mineral losses and resulting effects on physical and mental performance using modern analyti- cal capabilities. This study also should determine how much of each mineral is required to replenish losses and to optimize performance. It is expected that the primary increases in mineral losses during garrison training will be through sweat and urine, but endogenous fecal losses also may occur, either increasing with the stressful conditions or possibly decreasing to compensate for the sweat and urinary losses. Consequently, the committee sug- gests using a model environment that emulates conditions similar to military garri- son training (the committee also suggests that, even though sustained operations are not the focus of this report, using a model environment similar to those condi- tions could assess the related effects). Following are the details of a proposed study that will provide answers to questions about mineral requirements during garrison training (see Figure 4-1). An account of the general design is followed by descrip- tions of the dietary interventions, exercise interventions, and sleep distortion inter- vention; outcome measurements related to sweat losses, physical performance, and cognitive performance also are described or referenced.

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194 mental and physical and losses mineral on exertion physical and sweat of effect the load. lb Ready-to-Eat. determine 4050 to Meal, = with study MRE hours/day Proposed exercise; 4 4-1 = for Ex FIGURE performance. NOTE: *Exercise

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RESEARCH NEEDS 195 General Design A proposed study design to determine the effect of sweat and physical exer- tion on mineral losses and on physical and mental performance is illustrated in Figure 4-1. Study subjects should represent the genders and age ranges of sol- diers in the military population. Each subject would participate in a baseline phase (versus a stress phase with heat and heavy exercise) while consuming a controlled diet, based on either the current typical meals, ready to eat (MREs) or on the mineral values of the nutritional standards of operational rations (NSORs) [e.g., based on Recommended Dietary Allowance (RDA) for military garrison training], and would receive all subsequent treatments in randomized order while being subjected to heat and the physical demands typically experienced by mili- tary personnel. Controlled conditions would be achieved using an environmental chamber with supervised physical activity. Because training and operational ex- ercises may involve significant reductions in total sleep and disruptions of the normal sleepwake cycle, both of which reliably result in severe decrements in cognitive function and mood (Belenky et al., 1994; Lieberman et al., 2005), sleep restriction or disruption could be considered as an additional intervention. Dietary Interventions For all dietary interventions, studies should be controlled for other poten- tially limiting nutrients. One of the dietary periods during the stress phase should be identical to the baseline diet to determine the effect of heat and exercise on increases in mineral losses. There are several options for the dietary treatments. If the baseline dietary intake consists of typical MREs (usually with mineral levels higher than the NSOR), then other dietary treatments could include a lower and higher level of key minerals in "mineral cocktails" that include cal- cium, magnesium, iron, zinc, copper, and selenium. If the MRE diet does not meet the NSOR for any of the minerals, the lower dose of the intervention should be brought to at least the NSOR level. An alternative design is that the baseline diet could be set on the NSOR values, and the intervention could consist of one or two higher doses of minerals. In any case, the higher dose of minerals in the cocktail should not exceed the Tolerable Upper Intake Level (UL) for any nutrient. A one-week adaptation to each dietary intake is reasonable since the initial mineral status of subjects is not one of defi- ciency. Washout periods between treatments should be at least seven days. Exercise Intervention The exercise intervention should mimic garrison training (or the military situation of interest). The amount of energy expended in physical activity per day for a male in garrison training can be estimated from Tharion et al. (2005).

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196 MINERAL REQUIREMENTS FOR MILITARY PERSONNEL The total energy expenditure is estimated at 16.5 MJ/day (~ 3,900 kcal/day). For a 70-kg man, the resting metabolic rate should be about 1,680 kcal/day and the digestion and metabolism of food is approximately 10 percent of the food's energy content. Assuming energy intake is equal to energy expended (IOM, 2002/2005), a little more than 1,800 kcal/day will be expended in physical activ- ity. Using metabolic equivalent values (multiples of an individual's resting oxy- gen uptakes) to determine the kilocalories burned, walking at 3.5 mph on a level surface is the equivalent of about 4 kcal/min, and adding a 40-lb (18.14 kg) load should increase the energy expenditure by 12 kcal/min (Ainsworth et al., 2000); thus, the amount of energy expended from continuous exercise would be 300 360 kcal/h or about 3,0003,600 kcal/day. Therefore, the exercise intervention to employ in the study could be walking (possibly using a treadmill) at 3.5 mph carrying a 40-lb (18.14 kg) load. The work can be intermittent so as to provide a recovery period between exercise bouts. An environmental temperature of ap- proximately 30C and 70 percent of relative humidity would simulate summer conditions in the southeastern United States. Sleep Deprivation Intervention If sleep deprivation is a concern and included in the intervention, then en- suring that the soldiers are reasonably well rested before participating in the study is critical. A representative scenario could be achieved during the five-day intervention phase by limiting sleep to 4 h/day (prolonged-moderate restriction), a single 24-hour episode without sleep (acute-severe restriction), or a combina- tion of the two. This intervention's effectiveness can be verified by assessments of slow-wave activity in the electroencephalogram (EEG), sustained attention, and mood states. Sleep disruption (i.e., intermittent interruptions of sleep) and shifting the sleepwake cycle (i.e., circadian disruption) are alternatives for re- stricting total sleep time that better represent real situations. The Committee on Military Nutrition Research (CMNR) [IOM, 2004] recently reviewed the impact of sleep loss and disruption on cognitive performance in the military and the currently available measures for assessing sleepiness in military settings. Measurement of Mineral Losses This experimental design provides evaluation of a doseresponse effect of minerals on ameliorating mineral losses due to heat and sweat and can be used to evaluate a doseresponse effect of minerals on physical and mental performance. Daily, 24-hour whole-body sweat analysis during each period will best quantify mineral losses and, if conducted over several days (e.g., five days), will indicate adaptation due to acclimatization. Analyzing sweat losses during one extended period (two to three weeks) to determine longer-term adaptation also could be valuable. The wash-down procedure for dermal collections is described by

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RESEARCH NEEDS 197 Palacios et al. (2003). Because the increased sweat losses associated with high temperatures and heavy work are likely to occur without additional cellular losses, the mineral losses in sweat should be evaluated with and without proce- dures to exclude cellular debris, as described by Brune et al. (1986). Complete balance data are needed to evaluate all routes of potential loss as well as the mineral intakes needed to correct the losses. A quantitative fecal marker should be used (e.g., polyethylene glycol, chromium, or 57Cr). Stable isotopic tracers could be used to determine interactions among minerals on ab- sorption of calcium, zinc, and iron. Cognition and Behavior Measurements Outcome measures in all study phases (including baseline) could include assessments of sleep patterns, mood states, and cognitive function (e.g., atten- tion, memory, decision-making, and psychomotor skills). A wide variety of potentially useful biochemical, physiological, and behav- ioral markers for mental performance in military personnel were identified and reviewed briefly in a recent IOM report, Monitoring Metabolic Status (IOM, 2004). Future research on mineral requirements to maintain and optimize cogni- tive function and behavior should include careful review and consideration of the markers in designs modeling the full range of complexities and demands common to research in military settings. This battery of tests should be devel- oped in collaboration with Dr. Harris Lieberman (Lieberman, 2005; see Lieberman in Appendix B). The methods used to monitor and evaluate the effects of nutrition on cogni- tive function and behavior are similar regardless of the particular mineral nutri- ent of interest. Because successful performance of mental tasks typically in- volves the coordinated operation of several distinct cognitive and psychomotor processes, assessing the effects of an experimental intervention also involves measuring performance in several cognitive and psychomotor tasks--tasks em- phasizing sensation (i.e., the processes involved in detecting stimuli); attention [i.e., the processes of focusing on (selective) and allocating resources (intensive) to one aspect of the external or internal environment]; perception (i.e., the pro- cesses of interpreting or attaching meaning to stimuli); learning and memory (i.e., the processes of acquiring, storing, and retrieving information); reasoning (i.e., the processes of concept formation, problem solving, and decision making); and response selection and execution. An individual's knowledge base (e.g., general information, rules, and specific past events) is a critical component of cognitive function and, thus, also could be evaluated. Task failure can result from the breakdown of any one or more of the outlined component processes. The many other factors that directly or indirectly affect mental performance may need to be addressed by ensuring that they are comparable among all treat- ment and control groups, included as covariates in statistical analyses, or ma-

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198 MINERAL REQUIREMENTS FOR MILITARY PERSONNEL nipulated as part of the intervention. Important moderating factors to consider when assessing mental performance in military personnel are motor skills (in- cluding the availability and limitations on the skills required to execute the be- havior being recorded), training, and stressors. Improvements in task (physical or mental) performance result in part from the fact that some components in the process have become less effortful and more automated with training. Automat- ing task performance, usually achieved through repetition, requires fewer re- sources to successfully complete the task but small, important errors may go unnoticed for a longer period of time than if the task were not automated. The proposed study might include an assessment of whether and how quickly com- plex tasks relevant to the military are automated, and how this relationship is mediated by environmental, physical, and psychological stressors. Mental performance also is affected by chronic as well as physical and psychological states and stressors, which must be taken into account. Stressors directly related to physical and environmental factors common in the military include exercise-induced physical exhaustion, sleep restriction, food restriction, and extreme temperatures. Psychological states to be studied include mood states--such as anxiety, depression, hostility, and vigor--which also should be evaluated as outcomes in determining the effects of high-demand activities com- mon on mental performance. Measurements of Physical Performance The most important physical performance measurements are for aerobic and muscular endurance. There are many different tests that could measure these two variables, and the military should select the ones that have been proven to reflect the reality of military environments and physical performance demands. Because endurance exercise tests to exhaustion can be highly variable (Vogel, 1994), researchers recommend that an exercise time trial, where time will be the out- come measure, be used to determine aerobic endurance. A 15-km time trial on a cycle ergometer could be used to measure aerobic endurance as long as the study is conducted in a laboratory facility (Hinton et al., 2000). This test will require the subjects to first complete a maximal oxygen uptake (VO2max) test, where VO2max and associated responses (maximal heart rate or ventilation) are mea- sured. One test option is the bicycle test used by McArdle et al. (1973). An alternative time-trial method for measuring aerobic endurance in the field is a 20-km road march carrying a load (Vogel, 1994). Several methods can be used for measuring muscular strength and endur- ance. The preferred methods measure dynamic strength using isotonic muscle contractions in lifting weights to determine the one repetition maximal weight or an isokinetic dynamometer to determine the peak torque at different velocities (Howley and Franks, 1997). When an isokinetic dynamometer is available, mus- cular endurance can be measured by having subjects repeat maximal contrac-

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RESEARCH NEEDS 199 tions for 60 seconds (Vogel, 1994) and determining the rate of fatigue (the dif- ference between peak and minimum torque). Alternative methods of measuring muscular endurance include subjects doing either as many push-ups as they can within a one-minute time period or the maximum number that they can do (Howley and Franks, 1997). As stated, among the available test options for evalu- ating performance, scenarios that most closely simulate and measure military physical performance activities are optimal (Vogel, 1994). 2. Determine Mineral Status and Food and Dietary Intakes The committee recommends that the military conducts periodic surveys to determine the mineral intakes and status of soldiers at various times from entry to training, deployment, or combat, especially for calcium and iron. If mineral status cannot be measured, then the intake levels, at least should be surveyed. The following paragraphs describe potential studies to address calcium in- take and iron status determinations. Iron Justification. Approximately 1114 percent of women in the United States are iron deficient, according to the latest dietary survey (IOM, 2001). It is likely that the prevalence of iron deficiency among women joining the military is similar to that of women in the general population, although this fact is unproven. The committee concluded that women's iron status is an important criterion in deter- mining the necessity of a strategy for increasing iron intake. If a strategy is needed, it is critical to identify the most efficient one for decreasing the deficiencies. In addition to the potential iron deficiencies that may exist on entry into military service, an increase in iron requirements caused by active training could raise the prevalence of women's iron deficiency to > 50 percent; in fact, some women may become anemic. The adverse consequences on immune function, emotional and cognitive performance, and physical capacity may limit the effec- tiveness of women's training and performing duties. There are no systematic data available at this time on the true prevalence of iron deficiency and anemia in female troops either at service entry or during active military duty. The committee concluded that, in addition to a study on the feasibility of monitoring women's status on entry into military service, surveillance programs should be established to monitor iron status at the end of all intensive training phases as well as periodically thereafter during military service, including during garrison training. Question. What is the prevalence of iron deficiency and anemia in females in the military at service entry and during active service, including garrison training?

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200 MINERAL REQUIREMENTS FOR MILITARY PERSONNEL Study design. The study design is straightforward. The sampling should be statistically valid to represent military personnel's ages and races. Samples should be collected at the start of training or deployment as well as regularly throughout active duty to monitor the stability of the iron (or other minerals) nutritional status. Outcome measurements. The outcome measures for iron status should in- clude a complete blood count (CBC) (i.e., hemoglobin [Hb], hematocrit, plate- lets, etc.) plus serum ferritin, soluble serum transferrin receptor (sTfR), and per- haps erythrocyte protoporphyrin concentration (see Chapter 3). Calcium Justification. Calcium could be important for optimizing military perfor- mance by potentially helping to prevent stress fractures during training or com- bat and to modulate emotional health. Unfortunately, there is no biomarker that can indicate calcium nutrition status; instead, an indication of its status can be suggested from the total dietary intake. Researchers should conduct periodic surveys of calcium intake from food beverages, dietary supplements, and calcium-containing medications (e.g., antacids). Very little is known currently about dietary supplement use among enlisted military personnel in garrison training; however, there is anecdotal evidence that it is common, probably at a level higher than in the U.S. general population and similar to the athletic community. Using mineral supplements--such as multivi- tamin mineral supplements and single supplements (e.g., calcium and iron)--is fairly common and often contributes to total nutrient intakes that might other- wise be inadequate. In addition to anecdotal evidence, researchers are using surveys--including an anticipated 2005 edition of the Health Behaviors survey through the Department of Defense Military Health System and pre- and post- deployment surveys on the use of dietary supplements (Corum, 2004)--to gauge dietary supplement use in the military. It is imperative that military health ser- vices and commanders be provided with the results. Continuing the periodic surveys of food intake is important; however, given the use of dietary supplements, these surveys no longer provide enough informa- tion to describe accurately the total dietary intake of minerals by military person- nel in the field, and the food surveys need to include questions on dietary supple- ment use. The assessment of total dietary intakes for many nutrients requires that food and beverage, as well as dietary supplements and certain medications con- taining nutrients (e.g., calcium-containing antacids), be included in the surveys, since sizeable numbers of enlisted personnel use nutrient-containing dietary supplements and over-the-counter medications containing calcium. Determining the content of calcium in rations is necessary for assessing intakes; however, food composition tables often may not reflect accurately that information for military rations. Therefore, to gain useful estimates of actual calcium intakes,

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RESEARCH NEEDS 201 composite military diets should be analyzed by atomic absorption spectropho- tometry using acid-digested food samples. The same process, incurring only minimal additional resources, could be used to determine intake levels from rations for copper, magnesium, selenium, and zinc. Coordinating survey activities within the military is important to avoid du- plication of activities and to conserve limited resources. Because of the concerns relevant to iron and calcium, a more thorough knowledge of related supplement use among military personnel is a priority. Question. What is the dietary intake of calcium from food, dietary supple- ments, and calcium-containing medications? Study design and outcome measurements. Design and outcome measures for calcium status should include dietary surveys on food, dietary supplements, and medications containing calcium. Other Research Needs In addition to minerals, other dietary supplement products of unknown or unproven efficacy may be taken by enlisted personnel with the hope that they will improve performance or weight loss, or both. Although not a part of the current task, the committee acknowledges that the dietary intake surveys should extend to other supplements, including performance-enhancing supplements. The anecdotal evidence and the limited number of available surveys should provide insight as to which supplements the surveys should include. Among the various strategies for increasing nutrient intakes from foods, the use of dietary supplements may be warranted for some individuals, particu- larly women (e.g., to increase iron intake to adequate levels and folic acid intake, among women of child-bearing age, to minimize the risk of spina bifida in new- borns) and individuals on weight-loss diets (e.g., to increase mineral intakes to adequate levels on hypocaloric regimens). Research is needed to determine the best strategy for increasing necessary dietary intakes. There is still a need for research that addresses the most appropriate ways to disseminate to professionals as well as to soldiers nutrition-related informa- tion on food and dietary supplements, including the findings from the CMNR studies. RESEARCH NEEDS Calcium Calcium Losses Justification. Daily, whole-body calcium losses through sweat and excreta under conditions relevant to military personnel are poorly understood. Sweat

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208 MINERAL REQUIREMENTS FOR MILITARY PERSONNEL during periods of high physical stress and exhaustion as well as in the ideal laboratory conditions). The battery described by Lieberman et al. (2005, Lieberman, 2005; see Lieberman in Appendix B), with minor additions, is recommended for use in the proposed study because it is comprised of standardized and validated tasks that measure critical cognitive processes and psychomotor skills, also it has been used successfully in previous studies with military personnel. Included in the battery is a visual scanning task that measures sustained attention or vigilance; a matching-to-sample task that measures short-term memory and pattern recogni- tion; a repeated acquisition task that measures motor learning, attention, and short-term memory; a grammatical reasoning task that measures logical reason- ing; and a four-choice reaction-time task that measures psychomotor function. Data from several of these tasks are suitable for signal detection analysis as well as for traditional statistical analysis for intervention effects. This battery, which is based on tasks requiring responses to visual stimuli, could be enhanced by including tasks that require responses to auditory stimuli, such as Bakan's auditory vigilance task (Bakan, 1959), and by extending the reaction-time task to include a single-choice condition for measuring simple motor fatigue. Finally, this battery of cognitive tasks will address directly the possible impact of iron nutrition on memory function. Intervention effects on mood states, particularly depression and anxiety, can be determined by administering the POMS-BI. Alternatively, if a measure spe- cific to depression is desired so that cognitive and somatic aspects of depression can be evaluated independently, then researchers recommend using a standard- ized and validated test, such as the Beck Depression Inventory II (Beck et al., 1996), in conjunction with the POMS-BI. Supplement Use and Prevention of Iron Deficiency Justification. Iron supplementation can be a highly effective approach to treating iron deficiency, but iron can be toxic in large amounts. Hence, it is important to determine if supplementation is a viable route for protecting women who participate in heavy training from a decrease in iron status. Studies in the exercise literature demonstrate that small doses of ferrous sulfate can be effec- tive in eliminating the decline in iron status that occurs with heavy training (Brownlie et al., 2002; Hinton et al., 2000). It is important to determine if iron supplements are necessary or if fortified foods or dietary recommendations, or both, can effect a change. Question. Can supplemental iron and dietary intervention approaches alle- viate the drop in iron status of female soldiers during garrison training or even during field missions? Study design. The intervention could be a single pill, fortified foods plus a dietary recommendation, dietary recommendations, or a combination of supple-

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RESEARCH NEEDS 209 mentation plus fortification and dietary recommendations. Female recruits would be assigned randomly to the four intervention groups as well as to a placebo group (no intervention). Further evaluation is needed to assess the possible advantage of routine iron supplementation for all female soldiers versus iron supplementation only for those who are iron deficient after screening, so stratification may be necessary. Outcome measurements. Traditional iron status measures such as ferritin, sTfR, TSAT, erythrocyte protoporphyrin concentration, and CBC would be mea- sured as an index of the test subjects' iron status (see Chapter 3). Dietary intake data (obtained by either a questionnaire or weighed food intakes), dietary supple- ment use, and measures of iron status would provide evidence of each inter- vention's effectiveness. Other Research Needs Perform field testing of current filter paper technology to evaluate the feasibility of iron status biomarkers (i.e., ferritin or sTfR) as indicators of iron nutrition during long deployments. Develop methods to test field-friendly feel cognitive tasks (e.g., finger- tapping, memory tasks, etc.) that can be used without computers as a means of assessing of alterations in cognitive functioning. Magnesium Magnesium Losses Justification. Daily, whole-body magnesium losses through sweat and ex- creta under conditions relevant to military personnel are poorly understood. Sweat losses due to heat and physical exertion are the most probable route of magnesium losses. A short-term study could increase substantially the under- standing of the potential magnesium losses that military personnel could experi- ence and the magnesium levels that might be necessary to correct those losses. Question. What is the effect of environmental, physical, and psychological stressors and other conditions relevant to the military, including heat and physi- cal activity, on daily, whole-body magnesium losses? Study design and outcome measurements. See research priority 1, Study the Effects of Military Garrison Training on Mineral Losses and Performance for information on the recommended study design and outcome measurements (also see Figure 4-1). Magnesium and Cognition and Behavior Justification. Although there are no data specific to military personnel that address a possible relationship between magnesium nutrition and sleep, experi-

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210 MINERAL REQUIREMENTS FOR MILITARY PERSONNEL mental studies with civilians have shown that magnesium is involved in regulat- ing brain activity (Penland, 1995), plasma markers of magnesium status change with sleep deprivation (Takase et al., 2004), and magnesium supplementation decreases sleep disturbances, at least in the elderly (Held et al., 2002). Civilian studies also have demonstrated that magnesium nutrition may be related to de- pression (Murck, 2002). Further, several putative mechanisms for effects of mag- nesium on brain function relevant to sleep, cognition, and behavior have been identified. Given that sleep deprivation and disruption could occur common dur- ing military training and especially during sustained operations and produce se- vere decrements in cognitive function and mood states (Belenky et al., 1994; Lieberman et al., 2005), there is a need to determine whether increasing magne- sium intake will improve sleep, protect against the effects of sleep deprivation, or regulate mood states of military personnel in garrison training and during sustained operations. Question. Does increased magnesium intake improve sleep, protect against sleep deprivation, and regulate mood states of military personnel? Study design. This study can be conducted as part of the priority research 1. Study the Effects of Military Garrison Training on Mineral Losses and Perfor- mance, in which the dietary treatments consist of MREs supplemented with 0, 150, 300 mg/day of magnesium. These amounts were used in previous civilian studies and pose minimal risk of exceeding the UL when combined with typical dietary intakes. Assessments of sleep quantity and quality and of mood states should be conducted during the five-day periods of intervention with physical, environmental, and psychological stressors. Outcome measurements. The gold standard for measuring sleep architec- ture is polysomnography (PSG), which includes measurements of brain electri- cal activity (by electroencephalography), eye movements (by electrooculargram), and muscle tone (by electromyogram) was reviewed by the report Monitoring Metabolic Status (IOM, 2004). Data collected during the PSG can be analyzed spectrally and collated to permit identification and quantification of sleep stages and, thus, sleep patterns and should be used in the proposed laboratory simula- tion where there will be no practical equipment constraints. Wrist-worn acti- graphy would be the recommended practical alternative to PSG in a field study. Assessment of slow-wave (13.9 Hz delta and 47.9 Hz theta) activity in the EEG can be used to determine physiological changes in brain arousal and sleep quantity. Analysis of the PSG should be complemented by performing a sleep latency test (Wesensten, 2004) and administering a subjective measure of sleepi- ness (IOM, 2004), to assess sleep quality. Intervention effects (Lorr and and McNair, 1984) on mood states, particu- larly depression, can be determined by administering the POMS-BI (see section above Calcium and Cognition and Behavior). Alternatively, if a measure spe- cific to depression is desired so that cognitive and somatic aspects of depression can be evaluated independently, then using a standardized and validated test, such as the Beck Depression Inventory (Beck et al., 1996), in conjunction with

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RESEARCH NEEDS 211 the POMS-BI is recommended. Electrophysiological measurements are likely to respond to changes in magnesium status and when resources permit, should be included as outcome measures. The consequences of sleep deprivation and disruption on mental perfor- mance can be determined by administering a battery of cognitive tasks, includ- ing those that assess attention (particularly vigilance), perception (including pat- tern recognition), learning and memory, reasoning, and decision making. Effects on psychomotor performance can be determined with single and multiple-choice reaction-time tasks. The battery described by Lieberman et al. (2005), with mi- nor additions, is recommended for use in the proposed study (see the section on research needs for iron). The Committee on Military Nutrition Research (IOM, 2004) recently reviewed the impact of sleep loss and disruption on cognitive performance in the military and the currently available measures for assessing sleepiness in military settings. Magnesium Dietary Intake Levels Justification. The amount of magnesium consumed by military personnel is unknown, and there are only estimates of magnesium content in military rations, calculated mostly from food composition databases. In order to verify if the dietary intake level of magnesium is adequate the level in the diets, including dietary supplements, should be determined. Question. How much magnesium is consumed by military personnel under garrison training? Study design. Food composition tables often are inaccurate to estimate the magnesium contents of foods in military rations, and estimates of magnesium intake using existing food composition data tables are approximate at best. Therefore, com- posite military diets should be analyzed (by atomic absorption spectrophotometry using acid-digested samples) for magnesium to gain useful estimates of actual mag- nesium intakes. Researchers also should determine the frequency with which mili- tary personnel use supplements containing appreciable amounts of magnesium. This analysis could be done jointly with the calcium analysis described in research prior- ity 2. Determine Mineral Status and Food and Dietary Intakes. Outcome measurements. Measures will reflect the amount of magnesium in the military diets and rations using existing food composition data and actual magnesium analysis. Selenium Selenium Losses Justification. Daily, whole-body selenium losses through sweat and excreta in conditions relevant to military personnel are poorly understood. Sweat losses due to heat and physical exertion are the most probable route of excess losses. A

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212 MINERAL REQUIREMENTS FOR MILITARY PERSONNEL short-term study could increase substantially the understanding of the potential selenium losses that military personnel could experience and the levels of sele- nium that might be necessary to correct those losses. Question. What is the effect of conditions relevant to the military, including heat and physical activity, on daily, whole-body selenium losses? Study design and outcome measurements. See the study design in re- search priority 1, Study the Effects of Military Garrison Training on Mineral Losses and Performance. Selenium and Immune Function Justification. There are data that suggest that selenium deprivation can im- pair immune function. Question. Does selenium supplementation of nondeficient subjects improve immune function? Study design. Military men and women should be placed randomly in pla- cebo groups (soldiers eating typical diets, using no supplementation) and in a group taking a selenium supplement of 200 g/day. The total dietary intake would fall below the IOM UL for selenium (400 g/day). Outcome measurements. Following four weeks of supplementation, im- mune status improvements would be measured by postvaccination immune re- sponses [e.g., antibody production and T-cell number and function (phytohe- magglutinin and pokeweed mitogen) studies]. Selenium Dietary Intake Levels Justification. The amount of selenium consumed by military personnel is unknown, and there are only estimates of the selenium content in military ra- tions, calculated mostly from food composition databases. In order to verify if the dietary intake level of selenium is adequate the level in the diets, including dietary supplements, should be determined. Question. How much selenium is consumed by the military personnel under garrison training? Study design. Food composition tables often are inaccurate for estimating the selenium contents of military rations, estimates of selenium intake using existing food composition data tables are approximate at best. Therefore, com- posite military diets should be analyzed (by atomic absorption spectrophotom- etry using acid-digested samples) for selenium to gain useful estimates of actual selenium intakes. Researchers also should determine the frequency with which military personnel use supplements containing appreciable amounts (> 50 g/ day) of selenium. This selenium diet analysis could be done jointly with the calcium analysis described in research priority 2, Determine Mineral Status and Food and Dietary Intakes.

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RESEARCH NEEDS 213 Outcome Measurements. The measures should reflect the amount of sele- nium in military diets and rations using existing food composition data and actual selenium analysis. Selenium and Cognitive Function Justification. Data from several civilian studies have shown that increasing selenium intake may improve mood states, including depression, anxiety, and confusion (see Chapter 3). There have been no studies of selenium nutrition and cognition and behavior with military personnel, and civilian studies were con- ducted under minimal stress conditions. Thus, there is a need to determine the relationship between selenium intake and status and mood states in military personnel engaged in support, training, and combat operations and in the context of moderating variables, including physical, environmental, and psychological stressors. Question. Does increasing the selenium intakes of soldiers undergoing mili- tary garrison training improve their mood states, particularly depression, and therefore affect military performance? Study design. The research question can be addressed as part of research priority 1, Study the Effects of Military Garrison Training on Mineral Losses and Performance. The dietary treatment should consist of MREs supplemented with 0, 50, and 100 g/day of selenium. These amounts were used in previous civilian studies and pose minimal risk of exceeding the UL for selenium (400 g/ day) when combined with typical dietary intakes. If there is no indication of mood improvement after two weeks of supplementation, then supplementations and mood assessments should continue for a minimum of 2 months and prefer- ably 46 months. Outcome measurements. The POMS-BI (Lorr and McNair, 1984) is rec- ommended for use to efficiently measure multiple mood states (see section above Calcium and Cognition and Behavior). Other Research Needs Screen major water sources for selenium content. Zinc Zinc Losses Justification. The increase in zinc requirement for military personnel under garrison training conditions is based primarily on data that show increased sweat losses under conditions of heat exposure and exertion. Such increases are based on experiments lasting up to 12 days, with few subjects, and without measure-

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214 MINERAL REQUIREMENTS FOR MILITARY PERSONNEL ments that could assess possible adaptation in the intestinal absorption or endog- enous excretion of zinc (See Chapter 3). A short-term study could increase sub- stantially the understanding of the potential zinc losses that military personnel could experience and the levels of zinc that might be necessary to correct those losses. Question. Is there long-term adaptation in sweat losses or adaptation in other aspects of zinc balance that counteract the short-term increase in sweat losses with heat exposure and exertion? Study design. See the study design in research priority 1, Study the Effects of Military Garrison Training on Mineral Losses and Performance. Outcome measurements. The measures will address zinc levels in diet, stools, urine, and sweat (elemental balance). Isotopic measurements of zinc ab- sorption and endogenous intestinal excretion are recommended. Zinc Supplementation and Physical Performance Justification. Physical and mental performance measurements can provide the best evidence for setting dietary recommendations, especially since there are no reliable biochemical indexes of marginal zinc status. Performance data to justify increases in zinc intake need to be updated as newer, more sensitive measurements become available. An improvement in dynamic isokinetic strength was observed with 135 mg/day of zinc tablets (composition unknown) for 14 days (Krotkiewski et al., 1982); this outcome should be tested with zinc supple- mented in more moderate amounts, such as 1015 mg/day. Assuming that base- line intakes from MREs will be approximately 1015 mg/day (placebo), a supple- mentation of 1015 mg/day will amount to a total dietary intake of 20 and 30 mg/day for women and men, respectively, which is below the IOM UL of 40 mg/ day. Question. Does zinc supplementation enhance physical functions that may positively influence a soldier's performance? Study design. The randomized, placebo-controlled trials in the study design for Research Priority 1. Study the Effects of Military Garrison Training on Min- eral Losses and Performance could be followed to test physical performance with moderate amounts of supplemental zinc (e.g., 1015 mg/day) added to placebo dietary intakes (approximately 1015 mg/day) for approximately two weeks. For efficiency, zinc may be tested along with other nutrient supplements, with positive results leading to further testing for identification of the effective nutrients. Outcome measurements. Measures should address dynamic isokinetic strength, similar to that measured by Krotkiewski et al. (1982). Additional sensi- tive outcome measurements should be used as new testing conditions become available (see measurements of physical performance in Research Priority 1, Study the Effects of Military Garrison Training on Mineral Losses and Performance).

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RESEARCH NEEDS 215 Zinc and Cognition and Behavior Justification. Although there are no previous studies of military personnel, data from the few available civilian studies suggest that zinc nutrition may have a role in cognitive function, particularly memory and mood states (e.g., depres- sion). However, previous studies were conducted under laboratory conditions (i.e., minimal stress). Several putative mechanisms for effects of zinc on brain function and cognition have been identified. Thus, there is a need to determine the relationship between zinc intake and status and cognitive function and be- havior in military personnel engaged in support, training, and combat operations and in the context of moderating variables, including physical, environmental, and psychological stressors. Question. Does increasing the zinc intakes of soldiers undergoing military garrison training benefit cognitive function, particularly memory, or mood states (e.g., depression), and therefore affect military performance? Study design. The research question can be addressed as part of the overall Research Priority 1, Study the Effects of Military Garrison Training on Mineral Losses and Performance. The dietary treatment should consist of MREs supple- mented with 0, 10, and 20 mg/day of zinc. These amounts were used in previous civilian studies, including three conducted on children, and pose minimal risk of exceeding the IOM UL (40 mg/day) when combined with typical dietary intakes. Outcome measurements. The battery described by Lieberman et al. (2005) (see section above Iron and Cognition and Behavior) is recommended to assess the relationship between zinc nutrition and mental performance. Possible effects of zinc on mood states, including depression, can be assessed with the POMS- BI, as described previously (see section above Calcium and Cognition and Be- havior). Alternatively, if a measure specific to depression is desired so that the cognitive and somatic aspects of depression can be evaluated independently, then using a standardized and validated test, such as the Beck Depression Inven- tory II (Beck et al., 1996), in conjunction with the POMS-BI is recommended. Zinc Dietary Intake Levels Justification. The amount of zinc consumed by military personnel is un- known, and there are only estimates of zinc content in military rations, calcu- lated mostly from food composition databases. In order to verify if the dietary intake level of zinc is adequate the level in the diets, including dietary supple- ments, should be determined. Question. How much zinc is consumed by military personnel under garri- son training? Study design. Food composition tables often may not accurately reflect the zinc contents in the military rations, estimates of zinc intake using existing food composition data tables are approximate at best. Therefore, composite military

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216 MINERAL REQUIREMENTS FOR MILITARY PERSONNEL diets should be analyzed (by atomic absorption spectrophotometry using acid- digested samples) for zinc to gain useful estimates of actual zinc intakes. Re- searchers also should determine the frequency with which military personnel use supplements containing appreciable amounts of zinc. This zinc diet analysis could be done jointly with the calcium analysis described in Research Priority 2, De- termine Mineral Status and Food and Dietary Intakes. REFERENCES Ainsworth BE, Haskell WL, Whitt MC, Irwin ML, Swartz AM, Strath SJ, O'Brien WL, Bassett DR Jr, Schmitz KH, Emplaincourt PO, Jacobs DR Jr, Leon AS. 2000. Compendium of physical activities: An update of activity codes and MET intensities. Med Sci Sports Exerc 32(9 Suppl): S498S504. American Psychiatric Association. 1994. Diagnostic and Statistical Manual of Mental Disorders: DSM-IV. 4th ed. Washington, DC: American Psychiatric Association. Bakan P. 1959. Extraversion-introversion and improvement in an auditory vigilance task. Br J Psychol 50:325332. Beck AT, Steer RA, Brown GK. 1996. BDI-II Beck Depression Inventory: Manual. 2nd ed. San Antonio, TX: Psychological Corporation. Belenky G, Penetar DM, Thorne D, Popp K, Leu J, Thomas M, Sing H, Balkin T, Wesensten N, Redmond D. 1994. The effects of sleep deprivation on performance during continuous combat operations. In: Institute of Medicine, Marriott BM, ed. Food Components to Enhance Perfor- mance: An Evaluation of Potential Performance-Enhancing Food Components for Operational Rations. Washington, DC: National Academy Press. Pp.127135. Brownlie T 4th, Utermohlen V, Hinton PS, Giordano C, Haas JD. 2002. Marginal iron deficiency without anemia impairs aerobic adaptation among previously untrained women. Am J Clin Nutr 75(4):734742 Brune M, Magnusson B, Persson H, Hallberg L. 1986. Iron losses in sweat. Am J Clin Nutr 43(3): 438443. Corum SJC. 2004. Dietary supplement use in the military: Do Army health care providers know enough? AMEDD PB 8-04-1/2/3:3638. Curran SL, Andrykowski MA, Studts JL. 1995. Short form of the Profile of Mood States (POMS- SF): Psychometric information. Psychol Assess 7(1):8083. Held K, Antonijevic IA, Kunzel H, Uhr M, Wetter TC, Golly IC, Steiger A, Murck H. 2002. Oral Mg2+ supplementation reverses age-related neuroendocrine and sleep EEG changes in humans. Pharmacopsychiatry 35(4):135143. Hinton PS, Giordano C, Brownlie T, Haas JD. 2000. Iron supplementation improves endurance after training in iron-depleted, nonanemic women. J Appl Physiol 88(3):11031111. Hirsch ES, Kramer FM. 1993. Situational influences on food intake. In: Institute of Medicine, Marriott BM, ed. Nutritional Needs in Hot Environments: Applications for Military Personnel in Field Operations. Washington, DC: National Academy Press. Pp. 215243. Howley ET, Franks BD. 1997. Health Fitness Instructor's Handbook. Champaign, IL: Human Kinetics. IOM (Institute of Medicine). 1997. Dietary Reference Intakes for Calcium, Phosphorus, Magnesium, Vitamin D, and Fluoride. Washington, DC: National Academy Press. IOM. 2001. Dietary Reference Intakes. Vitamin A, Vitamin K, Arsenic, Boron, Chromium, Copper, Iodine, Iron, Manganese, Molybdenum, Nickel, Silicon, Vanadium, and Zinc. Washington, DC: National Academy Press. IOM. 2002/2005. Dietary Reference Intakes for Energy, Carbohydrate, Fiber, Fat, Fatty Acids, Cholesterol, Protein, and Amino Acids. Washington, DC: The National Academies Press.

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218 MINERAL REQUIREMENTS FOR MILITARY PERSONNEL Vogel JA. 1994. Evaluation of physical performance. In: Institute of Medicine, Marriott BM, ed. Food Components to Enhance Performance: An Evaluation of Potential Performance- Enhancing Food Components for Operational Rations. Washington, DC: National Academy Press. Pp. 113126. Wastney ME, Martin BR, Peacock M, Smith D, Jiang XY, Jackman LA, Weaver CM. 2000. Changes in calcium kinetics in adolescent girls induced by high calcium intake. J Clin Endocrinol Metab 85(12):44704475. Weaver CM. 2006. Clinical approaches for studying calcium metabolism and its relationship to disease. In: Weaver CM, Heaney RP, eds. Calcium in Human Health. Totowa, NJ: Humana Press. Pp. 6593. Weaver CM, Liebman M. 2002. Biomarkers of bone health appropriate for evaluating functional foods designed to reduce risk of osteoporosis. Br J Nutr 88(Suppl 2):S225S232. Wesensten N. 2004. Biomarkers for brain hypometabolism due to sleep deprivation. In: Institute of Medicine, Monitoring Metabolic Status. Predicting Decrements in Physiological and Cognitive Perfomance. Washington, DC: The National Academies Press. Pp. 381391.