United States

Environmental Protection Agency Office of Chemical Safety and Pollution Prevention (7101) EPA 712-C-10-001 July 7, 2010



# Product Performance

Test Guidelines

OPPTS 810.3700: Insect Repellents to be Applied to Human SkinNOTICE

This is one of a series of test guidelines established by the Office of Chemical Safety and Pollution Prevention (OCSPP) (formerly the Office of Prev ention, Pesticides and Toxic Substances (OPPTS) prior to April 22, 2010), United States Environmental Protection Agency for use in testing pesticides and chemical substances to develop data for submission to the Agency under the Toxic Substances Control Act (TSCA) (15 U.S.C. 2601, et seq. ), the Federal Insecticide, Fungicide and Rodenticide Act (FIFRA) (7 U.S.C. 136, et seq. ), and section 408 of the Federal Food, Drug and Cosmetic Act (FFDCA) (21 U.S.C. 346a) , referred to hereinafter as the harmonized test guidelines

.

The harmonized test guidelines serve as a comp endium of accepted scientific methods for research intended to provide data to inform regulatory decisions under TSCA, FIFRA, and/or FFDCA. This document provides guidance for conducting appropr iate tests, and is also used by EPA, the public, and the companies that are required to submit data under FIFRA. These guidelines are not binding on either EPA or any outside parties, and EPA may depart from them where circumstances warrant and without prior notice. The methods described in these guidelines are strongly recommended for generating the data that are the s ubject of the guidelines, but EPA recognizes that departures may sometimes be appropriate. You may propose and alternatives to the methods described in these guidelines, with your supporting rationale . The Agency will assess such proposals on a caseby-case basis.

For additional information about the harm onized test guidelines and to access the guidelines electronically, please go to http://www.epa.gov/oppts and select “Test Methods & Guidelines” from the navigation menu at the top of the screen. You may also access the guidelines in http://www.regulations.gov grouped by Series under Docket ID #s: EPA-HQ-OPPT-2009-0150 through EPA-HQ-OPPT-2009-0159, and EPA-HQ-OPPT-2009-0576.OPPTS 810.3700: Insect Repellents to be Applied to Human Skin

(a) Introduction.

(1) Scope and Purpose. This guideline provides r ecommendations for the design

and execution of studies to evaluate the performance of pesticide products intended to repel insects and other arth ropods in connection with the products’ registration under the Federal Insectic ide, Fungicide, and Rodenticide Act (FIFRA) (7 U.S.C. 136, et seq. ). This guidance applie s to products in any formulation—such as lotion, liquid, or spra y—intended to be a pplied directly to human skin. It does not apply to products applied to or impr egnated into clothing or fabric, or used to repe l insects from indoor or out door spaces. This guidance recommends appropriate study designs and methods for selecting subjects, statistical analysis, and reporting.

(2) General Considerations. Any protocol developed us ing this guidance must meet the requirements set forth in several st atutes and regulations , including, but not limited to, the Federal Insecticide, F ungicide, and Rodenticide Act (FIFRA, 7 U.S.C. 136, et seq. ) under which EPA regulates repellents, and EPA’s rules for the protection of human s ubjects of research, 40 CF R part 26, subparts K through Q. Because these studies would s upport a FIFRA registration and involve intentional exposure of huma n subjects to the test re pellents, review of each protocol by an Institutiona l Review Board (IRB), by EPA, and by EPA’s Human Studies Review Board (HSRB) is require d before the study is initiated. (See 40 CFR §26.1109, §26.1125, and §26.1601.)

This guideline does not supersede or ove rrule the regulations governing research conducted on human subjects contained in 40 CFR part 26, subparts K through Q, or any other Agency regulations. To the extent there are any unintended conflicts between this guideline and any EPA regula tion, the regulation at issue governs.

(3) Related Requirements.

(i) FIFRA Informed Consent Requirement. Any research conducted under this guideline is subject to §12(a)(2)( P) of FIFRA, which defines it as an unlawful act for any person “to use any pesticide in tests on human beings unless such human beings (i) are fully informed of the nature and purposes of the test and of any physical and me ntal health consequences which are reasonably foreseeable therefrom, and (i i) freely volunteer to participate in the test.” Regulations implementing this statutory provi sion and defining associated record-keeping requireme nts can be found at 40 CFR §169.2(j) and (k).

(ii) EPA’s Rule for the Protection of Human Subjects of Research. Any research conducted under this guidelin e is covered by the requirements of EPA regulations for the protection of hu man subjects of research set out at

140 CFR part 26, subparts K, L, and M. Persons conducting and submitting topical repellent efficacy studies sh ould ensure compliance with all applicable requirements of that rule; the following paragraphs highlight a

few of them:

(A) Applicability. Subparts K and L of 40 CFR part 26 apply to regulated third parties who conduct or sponsor research involving intentional exposure of human s ubjects which is intended for submission to EPA for consideration under the pesticide laws— FIFRA and §408 of the Federal Food, Drug, and Cosmetic Act (FFDCA) (21 U.S.C. 346a). (40 CFR §26.1101) Efficacy testing of topically applied repellents typically meets the regulatory definitions of “research” with “human subjects” involving “intentional exposure,” and must therefore be conducted and submitted to EPA in compliance with the requirements of these subparts. (See definitions at 40 CFR §26.1102.) Subpart M applies to any person who submits data from research with human subjects for consideration in c onnection with any action that may be performed by EPA under FIFR A or section 408 of FFDCA.

(B) Prohibition of research involving pregnant or nursing women

or children. 40 CFR §26.1203 as amended effective August 22, 2006, provides:

“Under no circumstances shall a person conduct or sponsor [covered] research that involves in tentional exposure of any human subject who is a pregnant woma n (and therefore her fetus), a nursing woman, or a child.”

(C) Required review by Institutional Review Board (IRB). 40 CFR §26.1109(a) provides:

“An IRB shall review and have authority to approve, require modifications in (to secure approv al), or disappro ve all research activities covered by this subpart.”

(D) Required Pre-testing submissions to EPA. 40 CFR §26.1125 provides:

“Any person or inst itution who intends to conduct or sponsor human research covered by §26.1101( a) shall, af ter receiving approval from all appropriate Inst itutional Review Boards (IRBs), submit to EPA prior to initiating such research all information relevant to the proposed resear ch specified by §26.1115(a), and, to the extent not already included:

2(a) A discussion of: (1) The potential risks to human subjects; (2) The measures proposed to minimize risks to the human subjects; (3) The nature and magnitude of all expected benefits of such research, and to whom they would accrue; (4) Alternative means of obtaining information comparable to what would be collected through the proposed research; and (5) The balance of risks a nd benefits of the proposed

research.

(b) All information for subjects and written informed consent agreements as originally pr ovided to the IRB, and as approved by the IRB.

(c) Information about how subj ects will be recruited, including any advertisements proposed to be used.

(d) A description of the circumstances and methods proposed for presenting information to potential human subjects for the purpose of obtaining th eir informed consent.

(e) All correspondence between th e IRB and the investigators

or sponsors.

(f) Official notification to the sponsor or investigator, in accordance with the requiremen ts of this subpart, that research involving human subjects has been reviewed and approved by an IRB.”

(E) Required post-testing submissions to EPA. 40 CFR §26.1303 provides:

“Any person who submits to EPA data derived from human research covered by this subpart shall provide at the time of submission information concerning the ethical conduct of such

research. To the extent available to the submitter and not previously provided to EPA, such information should include:

(a) Copies of all of the reco rds relevant to the research specified by §26.1115(a) to be prepared and maintained by an IRB.

(b) Copies of all of the records relevant to the information identified in §26.1125(a) through (f).

3(c) Copies of sample records used to document informed consent as specified by §26.1117, but not identifying any subjects of the research.”

(F) Prohibition of EPA reliance on unethical human research. 40 CFR §26.1705 provides:

“ . . . EPA shall not rely on data from any research initiated after April 7, 2006, unless EPA has adequate information to determine

that the research was conducted in substantial compliance with subparts A through L of this part , or if conducted in a foreign country, under procedures at least as protective as those in subparts A through L of this part.”

(iii) Good Laboratory Practice Standards. Good Laboratory Practice Standards (GLP) as defined in 40 CF R part 160 apply to both laboratory and field studies of repellent efficacy. According to 40 CFR §160.17: “EPA may refuse to consider reli able for purposes of supporting an application for a research or marke ting permit any data from a study which

was not conducted in accordance wi th this part.” 40 CFR §160.12(b) requires with any submitted research data “[a] statement describing in detail all differences between the pr actices used in the study and those required by this part.” Additiona lly, 40 CFR part 158 specifies that “applicants must adhere to the good laboratory practice (GLP) standards described in 40 CFR part 160 when conducting studies.” (40 CFR §158.70(b)).

(iv) State Requirements. Investigators and Sponsors should ensure research is conducted in compliance with any a pplicable state laws or regulations, which are independent of and additional to those cited here.

(4) Organization of the Guideline. This guideline begins with definitions of special importance in understanding this guideline (Section b). Sections (c) through (h) provide general guidance applicable to all topical repellen t efficacy testing,

whether conducted in the laborat ory or in the field, and to all target species. Each

of these six sections discusses one of th e primary stages of repellent testing.

(c) Development of protocols for repellent studies. (1) Scientific design of repellent studies. (2) Ethical justification for repellent studies. (3) Subject selection and informed consent. (4) Protection of subject privacy and confidentiality (d) Review of protocols for repellent studies. (e) Changes to IRB-approved research before execution. (f) Execution of repellent studies.

4(g) Reporting of completed repellent studies. (h) Retention of records.

Sections (i) through (l) provide guidance specific to diffe rent kinds of testing of repellents. Section (i) addresses empirical estimation of a typical consumer dose, and sections (j) through (m) address testing of repellency in the laboratory or field to particular species of arthropods.

(i) Specific guidance for dose-determination studies. (j) Specific guidance for laboratory st udies of mosquito or biting fly repellency. (k) Specific guidance for field studies of mosquito or biting fly repellency. (l) Specific guidance for laboratory st udies of tick or chigger repellency.

The final section (m) lists references considered in the development of this guideline.

Three appendices are attached as well:

Appendix A: Checklist of Elements Required by 40 CFR §26.1125. Appendix B: Framework for Scie nce and Ethics Reviews of Proposed

Human Research. Appendix C: Checklist of Elements Required by 40 CFR §26.1303.

(b) Definitions. The following definitions are of spec ial importance in understanding this guideline. They apply only in the context of this guideline and ar e not intended to be more generally applicable.

(1) The following events may indicate a failure of repellent efficacy.

(i) A landing is the act of a flying or jump ing insect or other arthropod alighting on human skin w ithout probing or biting.

(ii) A probe is the act of penetrating huma n skin by the mouthparts of an insect or other arthropod without ingestion of blood.

(iii) A bite is the act of penetrating human sk in by the mouthparts of an insect or other arthropod with ingestion of blood, typically associated with abdominal swelling and color change.

(iv) A crossing is the act of passage by a tick or chigger from an area of

untreated skin to an area of treated skin. A crossing may be quantified either or both by the distance the tic k or chigger moves onto treated skin or by how long the tick or chigger remains on treated skin.

5(2) An unconfirmed event is a landing, probe, bite, or crossing not followed by

another similar event within 30 minutes.

(3) A confirmed event is one landing, probe, bite, or crossing followed by another

similar event within 30 minutes. The firs t event is confirme d by the second; the

second event is the confirming event

.

(4) A human subject is a living individu al about whom an investigator conducting research obtains either data through interv ention or interaction with the individual or identifiable private information. By this definition, both untreated control subjects and treated subjects are considered human subjects of repellent efficacy testing.

(5) Questing is the behavior of ticks or ch iggers actively seeking a host.

(6) A repellent is a product intended to disrupt the host-seeking behavior of insects or other arthropods, driving or keeping th em away from treated human skin.

(7) Complete Protection Time (CPT) is the time from application of a repellent until efficacy failure as it is defined in each study—for example, the time from application until the first efficacy failur e event confirmed within 30 minutes by a

second similar event.

(8) Dose determination is a testing procedure used to estimate a “typical consumer dose” of a topical repellent.

(c) Development of protocol s for repellent studies. The first major stage of repellent testing is development of a protocol. Under EPA’s amended rules for the protection of human subjects—40 CFR part 26—protocol de velopment is the focus of far more

attention than was the case before that rule took effect in April 2006. A detailed protocol must be approved by an Institutional Review Board (IRB) and, accompanied by supporting documentation meeting the requi rements of 40 CFR §26.1125, must then be reviewed by EPA and the HSRB befo re research is initiated.

It is critical to all later stages of repellen t testing that protocol s contain the elements required by EPA’s Good Labor atory Practices regulations at 40 CFR §160.120 and meet all other applicable scie ntific and ethical standards. The broad topics of scientific design, ethical justification, subject selection and in formed consent, and protection of subject privacy and confidentiality are discussed in detail below.

(1) Scientific Design of Research. To be scientifically justified, the proposed research should address an important re search question that cannot be answered by existing data. In addition, the design should be such as to be likely to provide a definitive answer to the research ques tion. The design should include a detailed description of the experimental desig n, addressing topics (i) through (x).

6(i) Objectives. The objective of most repellent efficacy testing to support registration is to estimate how l ong after treatment a repellent will continue to protect users from the ta rget pest; this period of effective repellency is measured as CPT. Sometimes the objective of repellent efficacy testing is to compare the efficacy of one treatment to another treatment or to an untreated control; this ratio is expressed as Relative Protection (RP). In dose-determinati on testing associated with repellent testing the objective is typically to estimate a “typical consumer dose” for use in later repellent testing. In all cases the scientific objective should be stated clearly.

(ii) Test materials. Repellent efficacy should be tested using the end-use formulation as registered or as propo sed for registration. Test materials should be stored at ambient temp erature and humidity before use.

(iii) Choice of endpoints and measures. Endpoints chosen for the study should be appropriate for the specific objectives of proposed research and likely to provide a robust answer to the research question while minimizing the risks to subjects. Cons iderations in endpoi nt selection for dose determination and repellency include:

(A) Dose determination. To estimate a “typical consumer dose” the endpoint of concern is a rate of application, typically expressed either by weight as mg/cm 2 of treated skin surface or volumetrically as ml/cm 2 of treated skin surface. Because each subject is likely to apply a di fferent amount of repellent in uncontrolled trials, the “typical dose ” should be calculated as the mean of multiple applicati ons by each of many subjects.

(B) Repellency. The endpoint of repellency testing should be selected to show a failure of repellent e fficacy for subjects treated with a “typical consumer dose.” Efficacy failure in a test to determine Complete Protection Time (CPT) may be defined either as the first event—i.e., the first landing, bite, or crossing—or as the first confirmed event—i.e., the first la nding, bite, or crossing confirmed within 30 minutes by another similar event. The Agency encourages the use of the first c onfirmed failure event to estimate complete protection time for indi vidual subjects, and the median CPT across all subjects in each treatment arm as the summary

measure of CPT.

( 1 ) In repellency testing with mo squitoes or biting flies the study design should choose either landings or bites as the event showing failure of repe llent efficacy. The Agency encourages the choice of landings for field testing to reduce

7the risk of exposing subjects to vector-borne diseases. Even in laboratory testing us ing laboratory-reared, diseasefree insects, the choice of landi ngs reduces the risk of an

adverse reaction to b ites. A proposal to use bites as an endpoint in either field or laboratory testing should be justified.

( 2 ) In repellency testing with ticks or chiggers the appropriate event to show failure of repe llent efficacy is a “crossing”

from untreated skin onto treated skin.

(iv) Duration. Repellency testing should continue long enough to fairly assess the duration of protection pr ovided by the repellent, and long enough that all or nearly all subjec ts experience efficacy failure.

If testing ends before some subjects experience a failure of efficacy the resulting data-set is said to be “right-censored.” Right-censorship of data poses challenges for meaningful and accurate analysis. Right-censorship can be reduced through appropriate ca re in study design—for example, by planning a test of longer duration or , when appropriate, by pre-treating subjects well before their first expo sure to target pests. Methods for analyzing right-censored datasets ar e discussed in paragraph (c)(1)(xi)

below.

(v) Sample size. The sample should be large e nough to be likely to yield a definitive answer to the research question being addressed, and its size should be justified statistically in ea ch protocol, taking into account the specific characteristics of the proposed research and the desired accuracy and precision of the resu lts. Researchers are encouraged to consult a statistician to help determine appropriate sample size.

Withdrawal of test subjects from the study before failure of efficacy decreases the sample size and may co mpromise the validity or utility of test results. The protoc ol should fully describe how the proposed sample size was determined, and how data fo r subjects who withdraw from the test prematurely will be treated.

Other factors which may affect sample size are the numbe r of treatments, the experimental design, and the hete rogeneity of the target population (e.g., by age, sex) and of the environm ent (different habitats, different conditions, different species a nd population densities).

(vi) Allocation of subjects to treatments. Subjects should be allocated to treatments randomly and, to the extent possible, blindly.

8Multiple treatments per subject may be permissible if all treatments contain the same active ingredient at comparable concentrations, but caremust be taken in the study design and execution to prevent crosscontamination or confounding interference. It is particularly important for subjects receiving different treatment s on different limbs not to rub their limbs together or otherwise contaminate the repellent treatments, to keep results independent. The number of treatments per subject should be limited by the feeding behavior of the target species. For example, for mosquito species feedin g close to the ground on lower limbs, treatments should be applied only to subjects’ lower legs, an d only two treatments per subject will be possible. Treating th e same subject more than once with

the same treatment does not increase th e sample size; a single subject can only be counted once in determining the sample size.

When assessing relative protecti on provided by more than one product tested simultaneously, a Latin square design, in which each subject is

tested with each treatment over the course of the study, is often appropriate.

(vii) Untreated controls. To minimize risks to untr eated control subjects, both the number of untreated subjects and the duration of their exposure should be minimized. Untreated controls should be exposed intermittently, and then only long enough to confirm ade quate pest pressure throughout the study. Recent research (Barnard, et al. 2002) has shown that treated subjects should not serve as their own untreated controls.

(viii) Positive controls. Positive controls are de sirable to support comparison of results from different repellency studies or from testing on different days or at different locations. The recommended positive control material is 20% deet in ethanol, applied at a standard dose rate of 1 ml per 600 cm 2

. Positive controls are particularly valu able when the objectives of the study include comparisons between formul ations, and when testing spans different days. To minimize the like lihood of interference, subjects treated

with the test material should not also serve as concurrent positive controls.

(ix) Preparation of subjects. Before treatment with a test repellent or use as an untreated or positive c ontrol, subjects’ limbs s hould be washed with an unscented detergent and carefully rinsed and dried. Subjects should avoid alcohol, tobacco, and scented product s (perfume, cologne, hair spray, lotion, soap, etc.) for at least twelve hours before and throughout the test. Subjects should avoid activities th at increase perspiration, and avoid abrading, rubbing, touching, or we tting the treated area.

(x) Treatment of subjects. Subjects in trials desi gned to estimate a typical consumer dose should self-treat with the repellent, which should be provided in the type of container and delivery system (e.g., pump spray,

9aerosol spray, towelette, or lotion) a nd with the directi ons for use intended

for commercial distribution. Subjects in repellency trials should all be treated at the same standard dose rate, and treatment should be delivered

so as to ensure consistent a pplication and uniform coverage.

(xi) Statistical analysis plan. Protocols should include a full description, explanation, and justific ation for the statistical methods proposed to analyze both dose determination and re pellency test results, taking into account the specific study object ives and variables.

The statistical analysis plan should provide for testing results for normal

distribution. When results are normall y distributed, it may be appropriate to report the mean CPT across all treate d subjects with its standard error.

When the data do not fit a normal dist ribution—more typical of repellency datasets—it may be possible to transf orm them to fit a distribution for which a parametric method of analysis can be employed. When the data

do not fit and cannot be transformed to fit an underlying distribution, nonparametric analyses, such as Kaplan-Meier survival analysis 1 , are suggested.

If the study objective is to determin e CPT across all subjects, any rightcensorship of repellency data will lead to underestimation of both the

mean and the variance around it. Beca use right-censorship is common in repellency testing, EPA recommends use of the median CPT with its 95% confidence limits as the summary measure of CPT.

(xii) QA/QC plan. Protocols should provide for periodic quality assurance inspections adequate to ensure the integrity of the st udy and consistent with the requirements of EPA’s Good Laboratory Practices regulations (40 CFR part 160.)

(2) Ethical Justification for Research. Because repellent efficacy testing is never directly beneficial to the subjects of the research, th e risks to subjects must be minimized for the research to be ethically justified. The expe cted benefits to society from the knowledge likely to be gain ed in the research must also outweigh the minimized risks to subjects. (40 CFR §26.1111)

(i) Alternatives to research with human subjects. If the research question can be answered without conducting ne w research with human subjects, human research is not justifiable. Investigators should consider possible alternatives to human research; protocols should discuss possible

alternatives to human research and explain why they are infeasible or would not answer the research question.

1 For guidance see, for example, WHO (2009) Guidelines for Efficacy Testi ng of Mosquito Repellents for Human Skin: WHO/HTM/NTD/WHOPES/2009.A. Annex 3: Estimation of Median and Confidence Interval of Complete Protection Time Using the Kaplan-Meier Survivor Function.

10(ii) Prerequisite research. Before testing a repellent’s efficacy with human subjects, acute toxicity to animals of the active ingredients should be tested and reported in the protocol . Acute dermal toxicity, dermal irritation, eye irritation, and skin se nsitization studies are needed to estimate the margin of exposure (MOE ) for subjects participating in a repellent test. All additional componen ts in the materials tested should be cleared by EPA for use in repellent formulations.

(iii) Risks to subjects. Protocols should include a complete discussion of risks to subjects associated with thei r participation in the research, characterizing all risks and their likelihood, and describing steps proposed

to minimize each of them.

(A) Risk characterization. The nature of all risks to subjects should be described and the probabilities of occurrence of each type of risk should be estimated in the prot ocol. Potential risks in repellent testing typically include (but may not be limited to) reactions to the test substance or to arthropod b ites, acquisition of vector-borne illness, physical risks (e.g., stresses imposed by the requirements or conditions of the test), or po ssible psychological risk associated with a breach of confidentiality in handling the results of pregnancy testing.

(B) Risk minimization. As a condition of approval of proposed research, an IRB must determine th at risks to subjects have been minimized by using procedures wh ich are consistent with sound research design and which do not unnecessarily expose subjects to risk. (See 40 CFR §26.1111(a)(1).) Different actions are effective in minimizing different risks; pr otocols should id entify specific steps taken to minimize each identified risk to subjects.

Protocols should discuss sp ecific steps proposed to prevent research-induced harm—for exampl e, by training subjects, using appropriate eligibility criteria, ensuring adequate supervision of subjects, and having a physician on call during testing. In addition, protocols should specify how any research-induced harm will be

reversed —for example, by the inve stigator’s or sponsor’s committing to pay for uninsured costs of medical treatment of subjects for injuries or illnesses re sulting from their participation in the research, and for longer-term treatment, if needed, of any arthropod-borne disease cont racted during testing.

Study protocols should include pr ocedures for monitoring the safety of subjects, stopping ru les specifying conditions under which a subject would be withdr awn from the research or the

11research would be terminated to protect subjects, and a medical management plan covering foreseeable contingencies.

Methods of risk minimization are ultimately study-specific, but ensuring that risks to subjects ar e minimized in repellent testing may involve some or all of the following:

• Monitoring of potential field te sting sites for the presence

of disease vectors at least w eekly during the month before testing. Sites at which inf ected vectors are known to be present should not be used for field testing;

• Serological or DNA-based assays of insects collected at the site of field testing to determ ine the presence or absence of disease organisms. Results of such testing should be reported to subjects after completion of the field test;

• Using the minimum number of untreated controls consistent with statistical soundness;

• Exposing untreated control subjects intermittently for only the minimum time required to confirm continued pest

pressure;

• Training subjects to use an aspirator to capture landing insects before they have time to bite;

• Using only pathogen-free labor atory-reared insects in laboratory tests;

• Excluding subjects known to be sensitive to repellents or to insect bites;

• Using bites as the endpoint in either field or laboratory studies only when fully justified;

• Ensuring that enough sub-inves tigators are present at all times so that the Principal Investigator can, if needed, attend to the safety of a s ubject without compromising the integrity of the research or endangering other subjects;

• Initiating post-study contact wi th subjects to inquire about any signs of study-related injury or illness.

(iv) Expected benefits of research. Participation as a subject in repellent testing has no direct benefit for indi vidual subjects, yet is not risk-free,

even after risks have been minimized. Thus the ethical justification for the research ultimately depends on the e xpected benefits to society of the information likely to be gained fr om the research. Protocols should characterize all anticipated benefits of the information to be gained from the research, and to whom or to what segment in society each identified benefit is likely to accr ue. In addition, protocols should estimate the likelihood of achieving each identified societal benefit.

12Societal benefits include only thos e directly resulting from the knowledge likely to be gained from the research . Benefits potentially resulting from use of effective repellents should not be attributed to the proposed research. Payments to subjects or othe r incentives for participation should not be treated as benefits, either to the subjects or to society.

(v) Relation of benefits to risks. As a condition of approval of proposed research, an IRB must also determine th at minimized “risks to subjects are reasonable in relation to anticipated be nefits, if any, to subjects, and the importance of the knowledge that may r easonably be expected to result. In evaluating risks and benefits, the IRB should consider only those risks and benefits that may result from the research. . . . The IRB should not consider possible long-range effects of applying knowledge gained in the research . . . [to be] within the purview of its responsibility.” (See 40 CFR §26.1111(a)(2).) The protocol should discuss explicitly the balance of risks to subjects and anticipated so cietal benefits of the research.

(3) Subject selection and informed consent. Subject selection is important both to

the scientific merit of research and to its ethical acceptability. Selection of representative subjects is cr itical to the generalizability of the results of research, and to be ethically acceptable, subject selection must be e quitable. (40 CFR §26.1111(a)(3)) Fully informed and fully voluntary consent of subjects is

fundamental to ethical human resear ch. (FIFRA §12(a)(2)(P) and 40 CFR §26.1116)

(i) Representative sampling. The results of repellent testing should be as generalizable as possible to the targ et population of repellent users. Samples should include adults of vari ous ages, of both sexes, and of a variety of races and ethnicities, and protocols should describe the demographic characteristics of the pool from which subjects will be

recruited. Attractiveness of recruite d candidates to the target species should be verified before they par ticipate in repellency testing.

Testing with a sample known to be unrep resentative is discouraged; if it is proposed, it should be fully jus tified. Arguments based on the convenience of the investigators or on the difficulties associated with recruiting a more representative samp le do not justify testing with an unrepresentative sample.

(ii) Inclusion/Exclusion factors. All exclusion/incl usion criteria proposed should be supported by a clear and exp licit rationale. Because testing a topical repellent necessarily involve s intentional exposure of subjects, children under 18 and pregnant or nurs ing women must be excluded. (See 40 CFR §26.1203.) Because older adults are more susceptible to arthropod-borne diseases, candidate s over 55 should be excluded as subjects in field testing, or their inclusion should be specifically justified.

13Members of certain vulnerable popula tions, including people of limited mental capacity, those not in good hea lth or with compromised immune systems, those sensitive to chemicals, and students or employees of the investigators or sponsor s should always be excluded. Other potential subjects who may be in a vulnerabl e position, including the educationally or economically disadvantaged, or th ose who communicate with difficulty because of language differences or disabilities, should not be excluded arbitrarily if appropriate specific provision can be made to ensure their safety and welfare.

Subjects should generally be recruite d from populations in the area where testing will be conducted. An of fering of distant travel may unduly

influence a candidate’s choice to en roll, and a subject who has accepted long-distance transportation may feel less than free to withdraw from a study. If it is proposed, transportation of subjects to distant locations should be justified, and specific m echanisms should be proposed to prevent any coercion or undue influence.

(iii) Methods of recruiting. Protocols should describe in detail the proposed recruiting process, from the firs t contacts with potential candidates through all discussions of the research and the subjects’ enrollment. Any advertisements or flyers proposed for use in recruiting should be appended to the protocol, and should reflect IR B review and approval. If recruiting will be done through telephone calls, the script for recruiting calls should be appended to the protocol, and should reflect IRB review and approval. If any candidates may prefer to sp eak or read a language other than English, procedures for accommodati ng their needs in the recruiting process and in the conduct of the re search itself should be described in

detail.

(iv) Compensation of subjects. It is reasonable to compensate participants in repellent efficacy research for thei r time and trouble. The level of compensation should not be so high as to constitute an undue influence in the choice to participate, nor should it be so low as to make participation in the research attractive only to the economically disadvantaged. Compensation should not be used or admi nistered so as to compromise the freedom guaranteed to subjects to withdr aw from participation at any time for any reason, without sacrificing be nefits to which they are entitled.

(v) Informed consent. It is a fundamental requi rement for ethical research with human subjects that participatio n of subjects be both fully informed and fully voluntary.

(A) Process. Informed consent is an extended process, involving much more than simply obtaining a subject’s signature on a form.

14It begins with recruiting, and continues after signature of the consent form throughout the conduct of the research. A single discussion in the protocol of th e recruiting and consent processes

as a continuous whole is recommended.

As a condition of approval of proposed research, an Institutional Review Board (IRB) must determin e that informed consent will be sought from each prospective subject in accordance with, and to the extent required by §26.1116, and that informed consent will be appropriately documented, in accordance with and to the extent required by §26.1117. (See 40 CF R §26.1111(a)(4) and (5).)

The rule at §26.1116 requires that consent be sought “only under circumstances that provide the prospective subject . . . sufficient opportunity to consider whether or not to participate and that minimize the possibility of coercion or undue influence. The information that is given to th e subject shall be in language understandable to the subject.” EPA recommends that consent

discussions with indivi dual candidates be conducted in private, and

that all consent documents be written at or below an 8 th grade reading level, as measured by the Flesch-Kincaid Grade Level

score or another standard tool for assessing readabilit y. (A tool to

calculate the Flesch-Kincaid Grade Level score is a standard

feature available to users of MS-W ord.) If consent materials are translated into another language, it is important to ensure the

translations are also at an appropriate reading level.

The informed consent process should describe how the investigator will confirm each candidate’s understanding of the research and of

what it will involve before he or she is enrolled.

The rule at §26.1116 further define s the general requirements for informed consent, including at §26.1116(a) the basic elements always required, at §26.1116(b) other elements conditionally required, and at §26.116(e) a requirement to identify the pesticide

and the nature of its pesticidal function.

(B) Documentation of informed consent. The rule at §26.1117 provides two options for documenting informed consent. The first option is a “written consent docum ent that embodies the elements of informed consent required by § 26.1116.” The second is “a

short form written consent documen t stating that the elements of informed consent required by § 26.1116 have been presented orally to the subject.” The short form requires that a witness be present in the consent interview. In repellent efficacy testing the short form is generally inappropriate; if use of a short form is proposed,

15it should be justified, and prior di scussion with EPA is strongly encouraged.

Ensure that any elements appear ing in both the protocol and the consent form are in harmony. Elements common to both documents may include, for example, descriptions of the purpose of the research, subject inclusio n/exclusion factors, risks to subjects and how they will be minimized, and what will happen in the course of the research. The protocol and the consent form

should be consistent in substa nce, although the presentations should usually not be the same, because the readers and purposes

differ. Consent forms should addre ss candidates consistently in the second person—as “you”—and shou ld present all information from the subject’s point of view. By contrast protocols should be written in the third person, and s hould present all information from the investigators’ point of view.

In repellent testing, the consen t form should tell subjects how many bites, if any, they are likely to get, and what symptoms of arthropod-borne disease they should be alert for after participating in a field study.

(4) Protection of subj ect privacy and confidentiality. It is important to protect the privacy and confidentiality of subjects in repellent efficacy research, and to

inform candidates in the recr uiting and consent processes of the extent, if any, to which confidentiality of records identifying them will be maintained.

(i) Subject identification. Subjects should not be identified by name in data collection forms or study reports. S ubject names unavoidably must appear

on consent forms and on administrativ e documents, but subjects should be identified only by an arbitrary code on other study documents. The key linking identifying codes to subjects should be stored securely, away from other study records.

(ii) Photographic images. If photographs or videos are made to document the research, care should be take n to minimize making recognizable images of subjects. If faces or id entifying marks cannot be excluded from a photograph or video image, the imag e should be altered to protect the identity of the subject(s).

(iii) Pregnancy testing. Testing of female candidates for pregnancy should be handled with care and discretion. The investigator is responsible for ensuring that pregnant or nursing wo men are excluded as subjects (See 40 CFR §26.1203). It is not necessary to meet this responsibility by testing all female candidates for pregnancy. Fo r example, it is neither respectful nor informative to require women wh o are post-menopausal or surgically

16incapable of pregnancy to take a pr egnancy test. Investigators should

consider how best to meet their re sponsibilities while fully respecting female subjects.

When pregnancy testing is conducted, the circumstances of testing should protect the privacy of the candidate, bo th during the testing itself and in handling the results. Unexpected news of a pregnancy can cause significant psychological distress, wh ich can be increased by any breach of discretion in handling the informa tion. It is a good practice to recruit more candidates than are required by the study design, so that the design would not be compromised by withdrawal of one or more subjects. This approach permits a candidate with a positive pregnancy test to withdraw without stating a reason. No record s of a positive pregnancy test should

be retained.

(d) Review of protocol s for repellent studies. EPA’s Rule for the Protection of Human Subjects of Research requires ex tensive review of protocols be fore research is initiated. The rule requires that the complete protoc ol, consent form, and supporting materials be reviewed and approved by an IRB (See 40 CFR §26.1109), and having been approved by the IRB, that it be submitted, along with specified additional supporting documentation, to EPA for review by EPA and by the HSRB (See 40 CFR §26.1125 and §26.1601.)

(1) Review by IRB. The membership, functions, ge neral procedures, and decision

criteria for IRBs are defined in the rule at 40 CFR §26.1107-§26.1111. Records to be maintained by IRBs are define d at 40 CFR §26.1115. Most IRBs review many other kinds of human research in addition to repellent efficacy studies. Extensive guidance to and about IRBs can be found on the website of the Office of Human Research Protections (OHRP), at www.hhs.gov/ohrp

.

(i) Required elements in submission to IRB. Each IRB has its own application forms and procedures. Typical requirements are for a complete protocol, a consent form, and all materials intended for use in the recruiting process or to be provided to candidates or subjects. A protocol responsive to section (c) of this guidelin e is likely to satisfactorily address most requirements of a reviewing IRB.

(ii) Criteria for IRB approval. The criteria for IRB approval of proposed

research are defined in the rule at 40 CFR §26.1111. In summary, this passage of the rule requires that the IRB determine that proposed research

meets all these criteria:

• Risks to subjects are minimized;

• Risks to subjects are reasonable in relation to the importance of the knowledge that may be reasonably expected to result from the research;

17• Selection of subjects is equitable;

• Informed consent will be sought from each prospective subject;

• Informed consent will be appropriately documented;

• When appropriate, adequate provision is made for monitoring

collected data to ensure th e safety of subjects; and

• When appropriate, adequate provision is made to protect the privacy of subjects and to mainta in the confidentiality of data.

In addition, the IRB ensures that when any subjects are likely to be vulnerable to coercion or undue influence, additional safeguards are included in the study design to pr otect their rights and welfare.

(iii) Documentation of IRB approval. IRBs have the authority to approve, require modifications in (to secure approval), or disapprove research proposals. (40 CFR §26.1109.) It is common for IRBs to grant conditional approval subject to the investigator’s making specified modifications in the proposal; once required modifications are made, the IRB will issue formal written approval of the research. This letter of approval, along with all other corresponden ce between the in vestigators or sponsors and the reviewing IRB, must be included in the subsequent submission to EPA. (See 40 CFR § 26.1125(e) and (f).)

(2) Reviews by EPA and the HSRB. After receiving approval from an IRB, the protocol and supporting material must be submitted to EPA for review by EPA and by the HSRB. (See 40 CFR § 26.1125.)

(i) Required elements in submission to EPA. The elements to be included in a protocol submission to EPA ar e defined in the rule at 40 CFR §26.1125 and summarized in Appendix A to this guideline. Records of two kinds are required.

(A) Records describing the research proposal itself. 40 CFR sections 26.1125(a)–(d) call for a discussion of the risks and benefits of the proposed resear ch, for the consent forms as submitted to and as approved by the IRB, for information about how subjects will be recruited, and for a description of the circumstances and methods proposed for presenting information to subjects and seeking their consent. A protocol responsive to the guidance in section (c) of this gui deline will already include this range of information.

In addition, 40 CFR §26.1125(e) and (f) call for submission of all correspondence between the IRB and the investigators or sponsors (§26.1125(e)) and official notificatio n of IRB review and approval (§26.1125(f)). For purposes of §26.1125(e), “correspondence” does not include attachments transmitted by correspondence, such

18as the protocol or consent form, wh ich are otherwise required to be submitted by other provisions of the rule.

(B) Records of the IRB review. 40 CFR §26.1125 also requires submission of copies of records re levant to the proposed research which the IRB is required to maintain by 40 CFR §26.1108(a). There is some overlap between these records and those discussed above, but there are some im portant differences. 40 CFR §26.1108(a) calls for, among other records:

• Minutes of IRB meetings, in sufficient detail to show attendance; actions taken; votes for, against, and abstaining; the basis fo r requiring changes, and summarizing the discussion of controverted issues;

• A list of IRB members identified by name, earned degrees, representative capacit y, indications of experience sufficient to describe each member’s chief anticipated contributions to IRB deliberations, and any employment or other relationship between each me mber and the research institution; and

• Written procedures for the IRB in the same detail described in §26.1108(a) and §26.1108(b).

These records can only be obtaine d from the IRB. By prior arrangement, however, an IRB may submit them directly to EPA.

(ii) EPA Review. Timely EPA review of submitted proposals for research is required by the rule at 40 CFR §26.1601. EPA will first review submitted proposals for satisfaction of the requirements of 40 CFR §26.1125, and for substantiation of any claims of business confidentiality associat ed with the submission. If a submission is found to be incomplete, or to incl ude unsubstantiated claims of business confidentiality, the submitter will be prom ptly notified; the submission will not be

reviewed further until these deficiencies are corrected. EPA evaluates complete proposals in a single substantive review addressing both scientific and ethical aspects of the proposal, following the general outline in Appendix B to this guideline.

EPA will provide its completed review to the submitter. EPA’s review may call for revision of the proposal before it is reviewed by the HSRB. If EPA determines that the proposal is ready fo r HSRB review, it will be scheduled for HSRB review at the earliest opportunity. The submitter may respond to EPA’s review in writing to the HSRB docket, a nd may also comment orally at the HSRB meeting.

19(iii) HSRB Review. As provided in 40 CFR §26.1601(d), “following initial evaluation of the protocol by Agency sta ff, EPA shall submit the protocol and all supporting materials, together with the staff evaluation, to the Human Studies Review Board.” The HSRB is an advisory committee, chartered under the Federal Advisory Committee Act, charge d with advising EPA on scientific and ethical aspects of proposed and complete d research with hum an subjects. The HSRB meets several times a year in pub lic session, with opportunities for public participation. All materials they consider are kept in a public docket, except any which are subject to a substantiated claim of business confidentiality. HSRB recommendations are documented in a public report of each meeting, prepared as soon as possible after the meeting—typically within 60-90 days. As soon as it is available, EPA will provide a copy of the HSRB’s report to submitters of research proposals discussed by the HSRB. Additional information about the HSRB, including agendas and reports from past meetings and schedules of future meetings, can be found at the HSRB website: www.epa.gov/osa/hsrb

.

(e) Changes to IRB-approved research before execution. After EPA and the HSRB have reviewed and commented on proposed research, revisions to the prot ocol, consent form or other materials may be needed before th e research is executed. When IRB-approved materials are revised prior to in itiation of the research to ad dress concerns raised in EPA and/or HSRB reviews or for any other reas on, those revised materials must be resubmitted to the IRB for review and approval before the research is initiated. This requirement applies to all changes to an IRB- approved protocol, to all changes to an IRBapproved consent form, and to all changes to any other materials approved by an IRB. (40 CFR §26.1108(a)(4)). Although an IRB has the discretion to conclude that some changes do not require review by the full IRB, investigators and sponsors do not have this

discretion. EPA will treat as a breach of the regulations any planned changes to proposed research which are implemented w ithout prior approva l by the IRB.

(f) Execution of repellent studies

(1) Execution of protocol. When EPA and HSRB reviews are completed and the IRB has approved any revisions resulting from those revi ews, the research can be initiated. Subjects can be recruited and da ta can be collected as specified in the protocol. Care should be taken to ensure and document that these events occur in the proper sequence. Initiation of subject recruitment before obtaining IRB approval of the final protocol will be treated as a breach of the regulations.

(2) Quality Assurance (QA) oversight. Repellent efficacy testing, whether

conducted in the field or in the labora tory, is subject to the Good Laboratory Practices regulations at 40 CFR part 160. These rules require that each testing facility include an independent QA unit to monitor execution of each protocol and

document its conduct in accordance with the GLP regulations. (40 CFR §160.35)

20(3) Deviations from protocol. In executing even the best designed and most comprehensive protocols, unanticipated devi ations from the protocol may occur. All such deviations from the protocol should be promptly and fully reported to the IRB, and both the deviations and their im pact on the research should be discussed in the study report submitted to EPA. (40 CFR §160.185) EPA makes a distinction between planned changes to a pr otocol and deviations from a protocol. Planned changes must be treated as amen dments to the protocol, and must be approved by the IRB before they ar e implemented. (40 CFR §26.1108(a)(4))

(4) Changes in IRB-approved research after initiation. Any amendment to the protocol, consent form, or other materials approved by the IRB must be submitted to the IRB for review, and must be appr oved by the IRB before it is implemented, unless immediate implementation is requi red to avert an imminent hazard to subjects. (40 CFR §26.1108(a)(4))

(5) Reports of adverse events. Any adverse events a ffecting the subjects of a repellent efficacy test must be promptly reported to both the overseeing IRB and to EPA, including adverse events not esta blished to be related to product exposure or study participation. (40 CFR §26.1108(b)(1))

(g) Reporting of completed repellent studies

(1) Study report. In addition to the standard elements required by the GLP regulations at 40 CFR §160.185 and the append ices described below, the primary report of the study should include these elements, as applicable.

(i) Study identification: Title, identify ing study number(s), sponsor, study director, investigators, name and loca tion of the testing facility, and dates of the study.

(ii) Approved or intended la bel directions for use.

(iii) Recruiting and enrollment sta tistics, including the following:

• Demographic characteristics of th e pool from which subjects were

recruited and of the s ubject sample selected;

• Numbers of candidates contacted, interviewed, and screened; and

• Numbers of subjects enrolled, withdrawn voluntarily, withdrawn involuntarily, and completing the research.

(iv) Test arthropods: Genus, species, subs pecies or strain (if information is available) of target pests; and pest pr essure in each cage or field site. For laboratory studies, the source of test ar thropods, methods used to establish

21and maintain their disease-free stat us, rearing techniques and conditions, developmental stage, age and sex of target arthropods and methods used to sex them; preparation of arthropods befo re testing, and density per cage or

container.

(v) Test conditions: For field studies, the location and type of habitat, species distribution, climate and environmenta l conditions. For laboratory studies, a complete description of cages or containers, temper ature, relative humidity, ambient light, and air flow.

(vi) Test procedures, in cluding but not limited to:

• Preparation of subjects: Traini ng, demonstrating attractiveness, clothing and protective equipment worn, calculation of skin area and individual dose, washing, rinsing, and drying of limbs;

• Allocation of subjects to treatm ents; number of treatments per subject; blinding key, if used;

• Method of dose determination;

• Method of application of repellen t to each subject and individual dose applied;

• Time of application of repellent to each subject;

• Time of start and end of exposure of each subject.

(vii) Complete accounting for all ev ents during the testing period.

(viii) Results for each subject.

(ix) Reports of all statistical analyses

(x) Conclusions

(xi) Discussion

(2) Appendices.

(i) Complete protocol as approved by the IRB.

(ii) Consent form and any additiona l recruiting materials as approved by the

IRB.

22(iii) All amendments to the protocol, consent form, or other materials as approved by the IRB.

(iv) Reports of all deviations from the protocol and assessment of their impact on the integrity of the research.

(v) Raw data and data collection sheets.

(vi) Documentation of ethical conduc t of the research, as required by 40 CFR §26.1303, including all correspondence with IRB not previously submitted to EPA. The requirements of this se ction of the rule are summarized in Appendix C to this guideline.

(h) Retention of Records. The following record-keeping requirements apply to some or all records of research covered by this guideline.

(1) The record-keeping requirements in 40 CFR §26.1115 apply to Institutional Review Boards (IRBs) that review human research conducted under EPA’s

Human Research Rule.

(2) The record-keeping requirements of 40 CFR §169.2(j) apply to investigators who conduct pesticide research with hum ans subject to FIFRA §12(a)(2)(P).

(3) The record-keeping requirements of 40 CFR §169.2(k) apply to any person who

submits the results of research to EPA in support of a petition for a tolerance or tolerance exemption or in support of registra tion or an application for registration.

(4) The record-keeping requirements of 40 CFR §160.190 and §160.195 apply to records of any study conducted under the Good Laboratory Practices rule.

(i) Specific guidance for dose-determination studies. All subjects participating in repellency testing should be treated with th e test material at a standard dose rate, expressed by weight of repellent per unit area, mg/cm 2 of treated skin, or volumetrically

as ml/cm 2 of treated skin. Recommended methods for choosing the standard dose depend in part on the active ingredient(s ) and formulation(s) of the te st material. For testing of lotion formulations containing DEET (N,N-dieth yl-meta-toluamide) eet, a standard dose rate of 1 g per 600cm 2 (equivalent to 1.67 mg per cm 2 ) has been used in many tests of repellency. Repellents in lotion form contai ning DEET may be tested for repellency at this standard dose rate, or at a “typical consumer dose” rate determined empirically through a dose-determination study. For ot her ingredients and formulations no comparable standard dose has been iden tified, and it is recommended that a “typical consumer dose” be estimated through empirical dose-determination testing as described

below. The rationale for the dose rate act ually used in repellent testing should be documented in all study reports.

23(1) Test material. The formulated product should be used as it is or will be marketed, in the same type of container and bearing the same directions for use.

(2) Subjects. Subjects used for dose determinati on should be recruited and informed as described above, and mu st provide written informed consent for their participation. The same or very simila r eligibility criteria would apply as for repellency testing, although subjects pa rticipating only in dose-determination testing would not be exposed to any insects or to any of the risks associated with exposure to insects. The number of s ubjects used for dose determination should be justified statistically. The sample should include adults of both sexes and varied race, and should be as representa tive as possible of the repellent user population, consistent with ethica l and feasibility constraints.

(3) Measuring subject’s skin area. The surface area of subject’s limb in cm 2 can be estimated by measuring the circumference of the forearm in centimeters at the wrist and elbow, or of the leg at the ankle and knee, and in either case at one or two equally spaced intermediate poi nts; then multiplying the average circumference by the length of the limb in centimeters from the wrist to elbow or

from the ankle to the knee.

(4) Methods. Each subject should be instructed to apply the test repellent to his or her own limbs—to forearms or lower legs, or both—as they normally would apply a repellent to achieve complete c overage. Once the quantity applied has been measured and recorded, the applied repellent should be washed completely off the limb. This process should be re peated at least 3 times for each limb treated by each subject.

The applied quantity of a lotion or towe lette formulation can be determined by weighing the container or towe lette before and after use. Because less than all the spray released from a pump-spray or aeros ol container will reach the target limb,

a different method is needed to esti mate the applied quantity of a spray formulation. The target limb should be wrapped in gauze “b racelets” of known

area so that the bracelets c over only part of the skin. So long as the area of the bracelets is known, the quantity of a spray formulation applied to the skin per cm 2
can be estimated by weighing the bracelet s before and after application. Skin should be left exposed between bracelets to help subjects determine when they achieve complete coverage. Dose determ ination with spray formulations should

be conducted out-of-doors.

(5) Calculating standard dose for use in repellency trials. The mean dose applied by each subject to each limb and the grand mean across all subj ects of mean doses applied to forearms and to lower legs s hould be calculated. The specific gravity

of the test material should be used to convert the dose expre ssed by weight as mg

per cm 2 to a volumetric dose expressed as ml per cm 2 ; this calculation should be included in the study report. The volumetri c dose should be used as the standard dose for repellency testing, scaling it to the treated surface area of each subject’s

24limb or limbs. The protocol should incl ude the specific gravity of all test materials and should specify this calc ulation, and should specify doses both by weight and by volume.

(j) Specific guidance for labo ratory studies of mosquito or biting fly repellency.

(1) Species. Mosquito tests should be conducted with Aedes aegypti , an Anopheles species such as An. albimanus , or a Culex species, preferably Cx. quinquefasciatus. Stable fly tests should be conducted with Stomoxys calcitrans

. The source of test insects should be re ported, and if possible, they should be identified by subspecies or strain.

(2) Stage, age, and sex. Mosquito testing should be conducted with adult females 5- 10 days old; methods of sexing should be reported. Stable flies should be male or female adults 3-10 days old. The age range of test insects should be reported.

(3) Rearing techniques. Mosquito and biting fly la rvae should be reared under optimal conditions for the species—typically at 27° ± 3

 ° C, relative humidity 80% ± 10%, and photoperiod 16:8 hours (light:dark). Alterna tive rearing methods for mosquitoes are discussed by Gerberg et al . (1998). Use of alternative rearing techniques for biting flies and/or mosqu itoes should be fully documented and justified.

(4) Preparation of insects. Test mosquitoes should be fed 10% sucrose and no blood meal before use in a test, and star ved for 12 to 24 hours immediately before

the test. Test stable flies may be fed dry sugar cubes w ith a separate water source; they should also receive no blood meal before use in a test, and should be starved for 24 hours immediately before the test. Te st insects should be established to be free of disease; the source of insects and the methods used to ensure they are disease-free should be reported.

(5) Disposition of insects. Test insects should be used only once and destroyed immediately after the trial.

(6) Test cages. Cage size may vary, but is typically 2’ x 2’ x 2’ (8 cubic feet in volume, roughly equivalent to 232,000 cm 3 ), square or rectan gular in plan, with one or more sleeved openings for the s ubject’s arms. A mirro r on the bottom of the cage helps observers see insects landi ng on the underside of subjects’ arms. Larger cages may be used for two subjects at a time, so long as they are treated with the same repellent. Cage dimensions and design should be reported in detail.

(7) Insect density. There should be at least 200 mo squitoes in each 2’x2’x2’ cage (equivalent to one for each 1,160 cm 3 ) or at least 50 stable fl ies per cage (one for each 4,640 cm 3 .) After each exposure period all stable flies should be removed by vacuuming from each cage, and 50 fresh stable flies should be added to each cage.

25(8) Testing conditions. Ambient temperature during the test should be maintained at

21 ° ± 8°C, and relative humidity at 55- 85%. Ambient light appropriate to the preferred feeding time for the test species should be provided—e.g., more light for day feeders like Aedes aegypti , and less for night feeders like Anopheles spp. and Culex spp. When testing in subdued light , care should be taken not to compromise the investigators’ ability to observe insect activity. Investigators and subjects should avoid exhaling into the test cage; introduction of CO 2 could bias insects towards biting.

(9) Treated area size and preparation. The recommended treatment area is the subject’s forearm (wrist to elbow). This area should be washed with unscented soap, rinsed with a solution of ethanol or isopropyl al cohol in water, and dried

with a clean towel. The surface area of each test subj ect’s treated forearm (in

cm 2 ) and the volumetric dose administered to each subject (in ml) should be reported. Skin adjacent to the treated area should be covered with a light-colored

material that test insects cannot bite through. Hands should be covered with impenetrable (e.g., latex) gloves.

(10) Untreated controls. One or two untreated control subjects should verify continued landing pressure in tests of CPT. Results for these untreated subjects should not be compared to those for treat ed subjects; multiple untreated controls

should be used as a standard of co mparison only in studies of relative protection/percent repellency. To mi nimize potential interference, untreated subjects should not be treated on their other forearm. Forearms of untreated subjects should be washed, rinsed, and dried exactly like treated forearms.

(11) Positive controls. A positive control group treated at a rate of 1 ml per 600 cm 2

with 20% DEET in ethanol is recomme nded to calibrate the test system.

(12) Establishing subject attractiveness to test insects. Before the test, subjects should expose their untreated forearms to the target insects in a test cage to

establish their attractiveness. Five mos quito landings in one minute or less or 2 stable fly landings in two minutes or less are sufficient to establish attractiveness.

(13) Establishing and confirming landing pressure. Before exposing any treated subjects during each exposure period, an untreated subject’s forearm should be inserted into each cage to confirm landi ng pressure. Landing insects should be removed from the subject’s arm, by shaki ng it or by other means, before they have time to bite. The untreated subject should remove his or her forearm from the cage as soon as it has received the requi site number of landi ngs. If at any time fewer than 5 mosquitoes land on the untreat ed control forearm within one minute, all mosquitoes should be removed from a ll cages in the study and fresh insects should be added to each cage. If at any time fewer than 2 stable flies land on the

untreated control forearm w ithin two minutes, all stable flies should be removed

26from that cage and 50 fresh flies should be added. The aggressiveness of the fresh

insects should be confirmed before repellency testing continues.

(11) Exposure period. Approximately 30 minutes after treatment with the repellent, test subjects should insert th eir treated forearms into th e cage for the first time. Exposures of pre-defined duration should be repeated at regular intervals—e.g., 5- minute exposures at 30-minute intervals—until efficacy failure for each subject or the end of the study, whichever occurs first. Subjects should avoid rubbing their arms when inserting them into or re moving them from the cage and between exposure periods.

(k) Specific Guidance for field studies of mosquito or biting fly repellency.

(1) Pre-test subject preparation. Subjects in field studies of repellency should be trained in the laboratory to identify la ndings, probes, and bites of the target species and to use aspirators to collect landing insects before they have time to

bite.

(2) Choice of field testing sites. Field tests for mosquito repellency should be

conducted in at least two di stinct habitats (e.g., forest, grassland, salt marsh, wetland, beach, barns, or urban environments) where the predominant mosquito species differ. So long as the environmen ts and species distri bution differ, it is not important for test sites to be wide ly separated geographically. Repellency against black flies, stable f lies, or other biting flies t ypically occurring in only one habitat may be tested in only one habitat.

Potential sites for testing mosquito repellency should be monitored at least weekly for a month before testing is scheduled. To minimize risks to subjects, field testing should not be conducted where West Nile virus (WNV) or other mosquitovectored diseases have been detected w ithin the previous two weeks. Because biting flies are not known to vector human diseases in the USA, no comparable site monitoring is required for studi es of repellency to biting flies.

(3) Species identification. Mosquito tests should be conducted where more than one species is present. If tests are conduct ed outside the U.S., the relevance of the study for U.S. regulatory purposes should be justified in the study report. Landing insects should be aspirated or trapped before and during the test, and

labeled with the time of collection. Af ter the field study, collected insects should be identified by genus and sp ecies, and if possible, by s ubspecies or strain. The

number in each taxon collected in ea ch time period should be reported.

After identification, mosquitoes should be subjected to se rological or other analysis to determine the presence or absence of WNV or other disease organisms. The results of these analys es should be reported to subjects and included in the study report.

27(4) Untreated controls. Untreated control subjects are necessary in all study designs. To minimize the potential for inte rference, untreated c ontrols should be subjects who are not treated w ith a repellent on any limb.

When the objective is to estimate CPT, two untreated cont rol subjects are sufficient. They should expose an untreate d arm or leg briefly at regular intervals during the test to confirm continued accep table landing pressure. Results for the untreated controls should not be compared to results for treate d subjects. When the objective is to evaluate relative prot ection, more untreated control subjects are needed to provide a standard of statistical comparison.

Untreated limbs used as controls should be washed, rinsed, a nd dried exactly like

treated limbs.

(5) Positive controls. A positive control group treated at a rate of 1 ml per 600 cm 2

with 20% deet in ethanol is recomm ended to calibrate the test system.

(6) Minimum landing pressure to initiate or continue test. Landing pressure

should be measured before treatment and intermittently throughout the course of the test by untreated control subjects . Testing should not be conducted or continued unless landing pressure of the ta rget species is at least one mosquito landing within one minute, or at least one stable fly, black fly, ceratopogonid or tabanid landing within five minutes. In sects landing on untreated subjects should be collected for later identification, a nd labeled with the time of collection.

(7) Treated area size and preparation. The recommended treatment area is the subject’s forearm (wrist to elbow) or lo wer leg (ankle to knee), depending on the feeding behavior of the predominant species at the selected test site. A smaller treated area on the appropriate limb of at least 250 cm 2 is also acceptable. The

treated area should be washed with uns cented soap, rinsed with a solution of ethanol or isopropyl alcohol in water, a nd dried with a clean towel. The surface area treated for each test subject limb and the specific dose administered (ml) to each subject should be recorded. With the exception of the treated area, subjects’ heads, trunks, and limbs should all be c overed with light-colored material through

which insects cannot bite.

(7) Exposure period. Exposure of treated subjects may be intermittent—for example, for 1 or 2 minutes at 15-minute intervals, or for 5 minutes at 30-minute intervals—to reduce risks to subjects and al low them to rest between exposures.

When testing a repellent with a long period of effectivene ss, it is critical to the reliability of the data to minimize “right -censoring” of the data—i.e., the number of subjects who do not experience a failure of efficacy before the end of the test. Reliable results may be obtained for ex tended periods of protection by treating subjects up to several hours before the first field exposure, with subsequent

28exposures timed to coincide with periods of target insect activity. This approach reduces prolonged exposure of subjects to insects in the field, helps to reduce early withdrawal of subjects attributable to excessively long trials, and increases the likelihood that most or all subjects will experience a failure of efficacy.

Another acceptable design for assessing long-term repellency involves treating different subjects at different times (e .g., 2, 4, 6, 8, or 10 hours before exposure) and then exposing all subjects at the same time when target insects are active.

(8) Minimizing variation. Many factors contributing to variability in repellent field studies cannot be controlled, but mi nimizing variation when possible and designing studies with insect feeding patterns in mind can make a study design more efficient. Standardization is possi ble for such factors as time of testing, allocation to treatment, subject attractivene ss to target insects, etc. Other factors which may be managed to minimize vari ation include target ing a predominant species in its habitat, synchronizing expos ure periods with peak feeding activity of target insects, and treating limbs consis tent with the preference of the target species to feed on legs, arms, or both.

(9) Environmental conditions. The time of day at which subjects are treated and at which exposure to target insects begins a nd ends should be recorded and reported. Weather conditions during testing (inc luding temperature, relative humidity, cloud cover, precipita tion, light intensity, and wind speed) should be monitored periodically throughout the study and re ported. Testing should be not be conducted or continued if wind speed exceeds 16 kph/10 mph.

(10) Test subject pl acement and behavior. Subjects may work in pairs to assist each other to identify landings and collect insects. Because clustering of subjects may confound results, at least 3 m/10 ft should be maintained between pairs. Subjects

should avoid strenuous exercise and sweating, since these coul d affect test results.

(11) Data collection and reporting. Under the supervision of the investigator, an associate or another subject should reco rd the number and timing of each landing or bite during each exposure period for each subject. If possible, all landing

insects should be collected for identif ication and labeled with the time of

collection.

(l) Specific guidance for laboratory st udies of tick and chigger repellency. Because reliable field tests have not been developed, EPA recommends testing for repellency against ticks and chiggers in the laboratory. If field te sts are conducted, proposed label directions for re-application frequency should reflect field study results.

(1) Summary of recommended method. Each subject should place the fingers of one hand on a flat surface, with the elbow above the wrist and the forearm held at an angle of 30° or more to the surface. With a suitable instrument (such as an

29artist’s paintbrush, forceps, or a cotton swab) the investigator should place a tick or chigger, one at a time, on the subject’ s wrist, at a release point marked 3 cm below the boundary of the treated area of the forearm. The tick or chigger should be oriented gently (e.g., with paint brus h, forceps, or cotton swab) toward the treated area. After its first movement up the arm toward the margin of the treated area, each tick or chigger should be allowed 3 minutes to move across the boundary onto the treated area. A tick or ch igger that crosses at least 3 cm into the treated area (toward the elbow) is reporte d as ‘not repelled’. One that does not cross into the treated area, or that crawls into the treated area but immediately turns back or falls off, is reported as ‘repelled.’ Fresh tick s or chiggers are exposed to the treated area one at a time, at regular intervals for the duration of

the test.

While the general definition of a “crossing” is constant, the details of the method of scoring a crossing must be operati onalized in each protocol in a way appropriate to the species and life stage of ticks to be tested. A release point 3 cm distant from the treated area and scoring of a crossing when a tick moves at least 3

cm into the treated area have worked effectively in tests with nymphal

Dermicentor variabilis and Ixodes scapularis, and are provided here as examples.

(2) Species. Tick tests should be conducted us ing laboratory colonies of the tick species the label claims to repel. Common tick species in the United States include the blacklegged tick (deer tick, Ixodes scapularis ), western blacklegged tick (deer tick, Ixodes pacificus ), lone star tick (Amblyomma americanum ), American dog tick (Dermacentor variabilis ), and relapsing fever tick (softbacked tick, Ornithodoros turicata ). Chigger tests should be conducted using laboratory colonies of chiggers in the Trombiculidae family; Eutrombicula splendens; or E.

cinnabarrs . Test arthropods should be iden tified by genus and species, and if possible by subspecies or strai n. It is permissible to te st two species concurrently, alternating so that each s ubject qualifies and tests a tick or chigger from each species within each exposure period.

(3) Stage, age, and sex. When testing with blacklegged (deer tick), lone star, or softbacked ticks, either adult or nymphal life st ages are appropriate for testing. Only the adult American dog tick is recommende d, since nymphs of this species do not feed on humans. Tests with chiggers s hould use immature chiggers. The age or age range of all test ar thropods should be reported.

(4) Rearing techniques . Ticks and chiggers shoul d be reared at 25° ± 3 ° C, at high relative humidity (> 90%), and photop eriod of approximately 16:8 hours (light:dark). Any alterna tive rearing techniques shoul d be fully described and justified.

(5) Preparation of test arthropods . Ticks and chiggers should be disease-free. The

source of test animals and the methods used to ensure they are disease-free should be reported.

30(6) Disposition of test arthropods . Each tick or chigger should be used only once and should be destroyed immediately after use.

(7) Number of ticks or chiggers. Ticks or chiggers are te sted for questing behavior and for repellency one at a time, accord ing to the study design. Some studies using a single species of tic k test 5 ticks in each 30- minute exposure period; other

studies test one tick of each of two species in each 15-minute exposure period. Either pattern is acceptable.

(8) Testing conditions . Temperature should be maintain ed during the test at 20° to

25 ° C, with relative humidity at ≥ 35% and indirect ambient light 50 to 80%. The lights should be kept on.

(9) Subject preparation . Before treatment, both forearms of each subject are washed with unscented detergent and caref ully rinsed and dried. One forearm

from wrist to elbow is treated with the test material, and two lines are drawn on the subject’s wrist: a ‘boundary line’ at the edge of the treated area, and another

line—the ‘release line’—3 cm distant from the boundary line, outside the treated area toward the fingers. Similar lines ar e drawn in the same positions on the wrist of the subject’s other, untreated arm—one line at the wrist, and another 3 cm away toward the fingers.

(10) Untreated control. The untreated forearm of each treated subject is used to screen ticks or chiggers for questing behavior; only actively questing ticks or chiggers should be selected for repe llency testing. W ith an appropriate instrument, such as an artist’s paint brus h, forceps, or a cotton swab, each tick or chigger should be picked up carefully to prevent damaging its body or forelegs and placed on the release line on the wrist of the subject’s untreated arm. A tick or chigger that moves steadily from the release line across the boundary line and upward along the subject’s unt reated forearm is activel y questing, and appropriate for use in repellency testing. A tick or chigger failing this test should be immediately destroyed.

(11) Positive controls. A positive control group treated at a rate of 1 ml per 600 cm 2

with 20% deet in ethanol is recomm ended to calibrate the test system.

(12) Test procedure. With the treated forearm held upr ight above the table, one fresh tick or chigger at a time, immediately af ter demonstrating active questing on the subject’s untreated arm, is placed at th e release line on the treated arm. A “crossing” is recorded if the test organi sm crosses the boundary line at least 3 cm into the treated area within 3 minutes, and remains in the treated area for at least

one minute.

(13) Exposure period . At intervals of 15 to 30 minutes according to the study design, fresh ticks or chiggers are placed one at a time at the release point on the

31untreated arm and tested for active questing. Qualified ticks or chiggers are then placed at the release point on the treated ar m and exposed to the treated area. The cycle continues until efficacy failure or the end of the study, whichever occurs

first.

(14) Data collection and reporting. Recording of crossings and repelled test ticks or chiggers arthropods during the expos ure period should be supervised by investigators. The investigator, an associ ate, or another subject should record the number and timing of all events occurring during in each exposure period for each subject.

(m) References. The following publications were consu lted in developing this guideline.

(1) American Society for Testing and Mate rials. (2006) E 939-94 Standard Test Method of Field Testing Topical Applica tions of Compounds as Repellents for Medically Important and Pest Arthropods (Including Insects, Ticks and Mites): I Mosquitoes.

(2) American Society for Testing and Mate rials. (2003) E 1488-02 Standard Guide for Statistical procedures to use in Developing and Applying test methods.

(3) Barnard, D.R.; Ulrich, R.B.; Xue, R.; and Debboun, M. (2007) Chapter 5: Standard methods for testing mosquito repellents. In Insect Repellents: Principles,

Methods and Uses . Debboun, Frances, and Strickman, eds. CRC Press. (495 p.)

(4) Bernard, D. R., Bernier, U. R., Posey, K. H. and Xue, R-D (2002). Repellency of IR 3535, KBR 3023, para-menthane-3,8,-diol, and deet to Black Salt Marsh Mosquitoes (Diptera:Culicidea) in Everglades National Park. J. Med. Entomology39(6): 895-899

(5) Barnard, D.R. (1998) Mediation of D eet repellence in mosquitoes (Diptera: Culicidae) by species, age, and par ity. J. Med. Entomol. 35(3): 340-343.

(6) Barnard, D.R.; Posey, K.H.; Smith, D.; and Shreck, C.E. (1998) Mosquito density, biting rate and cage size effects on repellent tests. Med. Vet. Entomol.

12:39-45.

(7) Carroll, S. P. (2007) Chapter 12: Eval uation of topical insect repellents and factors that affect their performance. In Insect Repellents: Principles, Methods

and Uses . Debboun, Frances, and Strickman, eds. CRC Press. (495 p.)

(8) Frances, S.P. (1994) Response of a chigger, Eutrombicula hirsti (Acari: Trombiculidae) to repellent and toxican t compounds in the laboratory. J. Med. Entomol. 31(4): 628-630.

3233 (9) Frances, S.P.; Eikarat, N.; Sripongsai , B.; and Eamsila, C. (1993) Response of Anopheles dirus and Aedes albopictus to repellents in the laboratory. J. Am. Mos. Con. Assoc. 9(4): 474-476.

(10) Gerberg, E. J. ; Barnard, D.R.; a nd Ward, R.A. (1998) Manual for Mosquito Rearing and Experimental Techniques. AMCA Bull. 5.

(11) Govere, J.M.; and Durrheim, D. N. (2007) Chapter 8: Techniques for evaluating repellents. In Insect Repellents: Principles, Methods and Uses . Debboun, Frances, and Strickman, eds. CRC Press. (495 p.)

(12) Huntsberger, D. and Billingsley, P. (1981) Elements of Statistical Inference. 5 th
edition. Allyn and Bacon. Inc., Boston.

(13) Klun, J.A. and Debboun, M. (2000) A new module fo r quantitative evaluation of repellent efficacy using human subjects. J. Med. Entomol. 37(1): 177-181.

(14) Schofield, S.; Tepper, M.; and Gada wski, R. (2007) Fiel d evaluation against mosquitoes of regular and polymer-based deet formulations in Manitoba, Canada, with comment on methodological issues. J. Med. Entomol. 44: 457-62.

(15) Smith, C.N. (1955) Insect repellents. Quarterly Repor t, Entomological Research. Entomology Research Branch, U.S. Department of Agriculture. 8 p.

(16) Verwey, R.E. (1996) Laboratory method for testing insect repellents on human test subjects against chig gers in the laboratory. Unpublished document prepared by S.C. Johnson & Sons, Inc., Racine, WI. 3 p.

(17) WHO (2009) Guidelines for Efficacy Testing of Mosquito repellents for Human

Skin: CDS/NTD/WHOPES/2009.4.

(18) WHO (1996) Report of the WHO Informal Consultation on the Evaluation and Testing of Insecticides: CTD/WHOPES/1C/96.1.Appendix A: Checklist of El ements Required by 40 CFR §26.1125

34 40 CFR 26.1125 Prior submission of proposed human research for EPA review

Any person or institution who intends to conduct or sponsor hu man research covered by §26.1101(a) shall, after receiving approval from all appropriate IRBs, submit to EPA prior to initiating such research all information relevant to the proposed research specified by §26.1115(a), and the following addition al information, to the extent not already included:

Requirement Y/N Comments/Page Refs (1) Copies of

• all research proposals reviewed by the IRB,

• scientific evaluations, if any, that accompanied the proposals reviewed by the IRB,

• approved sample consent documents,

• progress reports submitted by investigators, and reports of injuries to subjects. (2) Minutes of IRB meetings . . . in sufficient detail to show

• attendance at the meetings;

• actions taken by the IRB;

• the vote on these actions including the number of members voting for, against, and abstaining;

• the basis for requiring changes in or disapproving research;

• a written summary of the discus sion of controverted issues and

their resolution. (3) Records of continuing review activities. (4) Copies of all correspondence between the IRB and the investigators. (5)

• A list of IRB members identified by name; earned degrees; representative capacity; indications of experience such as board certifications, licenses, etc., sufficient to describe each member’s chief anticipated contributions to IRB deliberations;

• any employment or other relationship between each member and the institution, for example, fu ll-time employee, a member of governing panel or board, stockholder, paid or unpaid consultant. (6) Written procedures for the IRB in the same detail as described in §26.1108(a) and §26.1108(b).All information relevant to the proposed research specified by § 26.1115(a)

(7) Statements of significant new findings provided to subjects, as required by §26.1116(b)(5). (1) The potential risks to human subjects (2) The measures proposed to minimize risks to the human subjects; (3) The nature and magnitude of all expected benefits of such research, and to whom they would accrue (4) Alternative means of obtaining information comparable to what would be collected through the proposed research; and§1125(a)a discussion of:

(5) The balance of risks and benefits of the proposed research. §1125(b): All information for subjects and written informed consent agreements as originally provided to t he IRB, and as approved by the IRB. §1125(c): Information about how subjects will be recruited, including any advertisements proposed to be used. §1125(d): A description of the circumstances and methods proposed for presenting information to potential human subjects for the purpose of obtaining their informed consent. §1125(e): All correspondence between the IRB and the investigators or

sponsors.The following Information, to theextent not already included:

§1125(f): Official notification to the sponsor or investigator . . . that research involving human subjects has been reviewed and approved by

an IRB.Appendix B: Framework for Science and Ethics Reviews of Proposed Human Research

35 Questions to be Addressed in EPA Protocol Review

Protocol Identification

(a) Title (b) Date (c) Principal Investigator and any sub-investigators (d) Participating Laboratories (e) Sponsor (f) Reviewing IRB

1. Societal Value of Proposed Research

(a) What is the stated purpo se of the proposed research? (b) What research question does it address? Why is this question important? Would the research fill an important gap in understanding? (c) How would the study be used by EPA? (d) Could the research question be answer ed with existing data ? If so, how? (e) Could the question be answered without ne wly exposing human subjects? If so, how?

2. Study Design

(a) What is the scientific objective of the study? If there is an explicit hypothesis, what is it? (b) Can the study as proposed achieve th at objective or test this hypothesis?

2.1 Statistical Design

(a) What is the rationale fo r the choice of sample size? (b) What negative and positive controls ar e proposed? Are proposed controls appropriate for the study design and st atistical analysis plan? (c) How is the study blinded? (d) What is the plan fo r allocating individuals to treatment or control groups? (e) Can the data be statistically analyzed? (f) What is the plan for statis tical analysis of the data? (g) Are proposed statistical methods appropr iate to answer the research question? (h) Does the proposed design have adequate statistical power to definitively answer the research question?

2.2 How and to what will human subjects be exposed?

(a) To what will subjects be exposed? (b) What is the rationale for the choi ce of test material and formulation? (c) What is the rationale for the choice of dose/exposure levels and the staging of dose

administration? (d) What duration of exposure is proposed?Appendix B: Framework for Science and Ethics Reviews of Proposed Human Research

36 2.3 Endpoints and Measures

(a) What endpoints will be m easured? Are they appropriate to the question(s) being

asked? (b) What steps are proposed to ensure meas urements are accurate and reliable? (c) What QA methods are proposed? (d) How will uncertainty be addressed? Will reported point values be accompanied by measures of uncertainty?

3. Subject Selection

3.1 Representativeness of Sample

(a) What is the population of c oncern? How was it identified? (b) From what populations will subjects be recruited? (c) Are expected participants representative of the population of c oncern? If not, why

not? (d) Can the findings from the proposed study be generalized beyond the study sample?

3.2 Equitable Selection of Subjects

(a) What are the inclusion/ex clusion criteria? Are they complete and appropriate? (b) What, if any, is the relationship betw een the investigator and the subjects? (c) If any potential subjects are from a vulne rable population, what is the justification for including them? (d) What process is proposed for recr uiting and informing potential subjects? (e) If any subjects are potentially subject to coercion or undue influence, what specific safeguards are proposed to prot ect their rights and welfare?

3.3 Remuneration of Subjects

(a) What remuneration, if any, is proposed for the subjects? (b) Is proposed remuneration so high as to be an undue inducement? (c) Is proposed remuneration so low that it will only be attrac tive to economically disadvantaged subjects? (d) How and when would subjects be paid?

4. Risks to Subjects

4.1 Risk characterization

(a) Have all appropriate prerequisite studies been performed? What do they show about

the hazards of the test materials? (b) What is the nature of the risks to subjects of the proposed research? (c) What is the probability of each risk asso ciated with the research? How was this probability estimated?Appendix B: Framework for Science and Ethics Reviews of Proposed Human Research

37 4.2 Risk minimization

(a) What specific steps are proposed to minimize risks to subjects? (b) How do proposed dose/exposure levels co mpare to established NOELs/NOAELs for

the test materials? (c) What stopping rules are proposed in the protocol? (d) How does the protocol provide for medical management of potential illness or injury to subjects? (e) How does the protocol provide for safety monitoring? (f) How does the protocol provide for post-e xposure monitoring or fo llow-up? Is it of long enough duration to discover adve rse events which might occur? (g) How and by whom will medical care for res earch-related injuries to subjects be paid

for?

5. Benefits

(a) What benefits of the proposed researc h, if any, would accrue to individual subjects? (b) What benefits to society are anticipated fr om the information likely to be gained through

the research? (c) How would societal bene fits be distributed? Who would benefit from the proposed

research? (d) What is the likelihood that each identifi ed societal benefits would be realized?

6. Risk/Benefit Balance

(a) How do the risks to subjects weigh against the anticipated benefits of the research, to subjects or to society?

7. Independent Ethics Review

(a) What Institutional Review Board (IRB) reviewed the proposed research? (b) Is this IRB independent of the inve stigators and sponsors of the research? (c) Is this IRB registered with OHRP? (d) Is this IRB accredited? If so, by whom? (e) Does this IRB hold a Federal-Wide Assurance from OHRP? (f) Are complete records of the IRB review as required by 40 CFR §26.1125 provided? (g) What standard(s) of ethical conduct would govern the work?

8. Informed Consent

(a) Will informed consent be obtained from each prospective subject? (b) Will informed consent be appropriately doc umented, consistent with the requirements of 40 CFR §26.1117? (c) Do the informed consent materials meet the requirements of 40 CFR §26.1116, including adequate characterization of the risks and di scomforts to subjects from participation in the research, the potential benefits to the subject or others, and the ri ght to withdraw from

the research?Appendix B: Framework for Science and Ethics Reviews of Proposed Human Research

38 (d) What is the literacy rate in English or other languages among the intended research subjects? (e) What measures are proposed to overc ome language differences, if any, between investigators and subjects? (f) What measures are proposed to ensure s ubject comprehension of risks and discomforts? (g) What specific procedure will be followed to inform prospective subjects and to seek and

obtain their consent? (h) What measures are proposed to ensure fully voluntary participation and to avoid coercion

or undue influence?

9. Respect for Subjects

(a) How will information about prospective and enrolled subjects be managed to ensure their privacy? (b) How will subjects be informed of their freedom to withdraw from the research at any time without penalty? (c) How will subjects who decline to participate or who withdraw from the research be dealt

with?Appendix C: Checklist of El ements Required by 40 CFR §26.1303

39 § 26.1303 Submission of Completed Human Research for EPA Review

Any person who submits to EPA data derived from human research covered by this su bpart shall provide at the time of submission information concerning the ethical conduct of such research. To t he extent available to the submitter and not previously provid ed to EPA, such informati on should include:

Requirement Y/N Comments/Page References

§1115(a)(1): Copies of

• all research proposals reviewed,

• scientific evaluations, if any, that accompany the proposals,

• approved sample consent documents,

• progress reports submitted by investi gators, and reports of injuries to subjects. §1115(a)(2): Minutes of IRB meetings which shall be in sufficient detail to show

• attendance at the meetings;

• actions taken by the IRB;

• the vote on these actions including the number of

• members voting for, against, and abstaining;

• the basis for requiring changes in or disapproving research;

• a written summary of the discussion of controverted issues and their

resolution. §1115(a)(3): Records of continuing review activities. §1115(a)(4): Copies of all correspondence between the IRB and the investigators. §1115(a)(5):

• A list of IRB members identified by name; earned degrees; representative capacity; indications of experience su ch as board certifications, licenses, etc., sufficient to describe each mem ber’s chief anticipated contributions to IRB deliberations;

• any employment or other relationship between each member and the institution, for example, full-time employee, a member of governing panel or board, stockholder, paid or unpaid consultant. §1115(a)(6): Written procedures for the IRB in the same detail as described in § 26.1108(a) and § 26.1108(b).(a) Copies of all of the records relevant to the research specified by§26.1115(a) to be prepared and maintained by an IRB

§1115(a)(7): Statements of significant new findings provided to subjects, as required by § 26.1116(b)(5). (1) The potential risks to human subjects; (2) The measures proposed to minimize risks to the human subjects; (3): The nature and magnitude of all ex pected benefits of such research, and to whom they would accrue; (4) Alternative means of obtaining information comparable to what would be collected through the proposed research; and§1125(a)A discussion of:

(5) The balance of risks and benefits of the proposed research. §1125(b): All information for subjects and written informed consent agreements as originally provided to the IRB, and as approved by the IRB. §1125(c): Information about how subjects will be recruited, including any advertisements proposed to be used. §1125(d): A description of the circumstances and methods proposed for presenting information to potential human subjects for the purpose of obtaining

their informed consent. §1125(e): All correspondence between the IRB and the investigators or sponsors.(b) Copies of all of the records relevant tothe information identified in §26.1125(a)-(f)

§1125(f): Official notification to the sponsor or investi gator, in accordance with the requirements of this subpart, that res earch involving human subjects has been reviewed and approved by an IRB. (c) Copies of sample records used to document informed consent as specified by §26.1117, but not identifying any subjects of the research (d) If any of the information lis ted in paragraphs (a) through (c) of this section is not provided, the person shall describe the e fforts made to obtain the information.