[Federal Register Volume 79, Number 98 (Wednesday, May 21, 2014)]
[Notices]
[Pages 29176-29177]
[FR Doc No: 2014-11720]

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DEPARTMENT OF ENERGY

Federal Energy Regulatory Commission

[Docket No. CP14-473-000]

Gulf South Pipeline Company, LP, Petal Gas Storage, LLC; Notice
of Application

    Take notice that on May 1, 2014, Gulf South Pipeline Company, LP
(Gulf South) and Petal Gas Storage, L.L.C. (Petal) filed in the above
referenced docket an application, pursuant to sections 7(c) and 7(b) of
the Natural Gas Act (NGA) and Part 157 of the Commission's regulations,
requesting authorization for Gulf South to consolidate into its
existing interstate natural gas operations the jurisdictional natural
gas storage facilities owned and operated by Petal. The consolidated
facilities will increase administrative efficiency; eliminate multiple
leases between Gulf South and Petal; provide greater operational
redundancies in physical facilities; and provide Gulf South more
flexibility to create additional services that the market desires
through the combination of facilities, all as more fully described in
the Application which is on file with the Commission and open to public
inspection.
    The filing may be viewed on the web at http://www.ferc.gov using
the ``eLibrary'' link. Enter the docket number excluding the last three
digits in the docket number field to access the document. For
assistance, contact FERC at<EMAIL_ADDRESS>or call

toll-free,<PHONE_NUMBER> or TTY,<PHONE_NUMBER>.
    Any questions regarding this application may be directed to J. Kyle
Stephens, Vice President of Regulatory Affairs, Boardwalk Pipeline
Partners, LP, 9 Greenway Plaza, Houston, Texas, 77046; by fax to (713)
479-1846; or by email to<EMAIL_ADDRESS>    Specifically, Gulf South seeks (i) authorization for Petal to
abandon by transfer all of its jurisdictional facilities to Gulf South;
(ii) authorization for Gulf South to acquire by inter-corporate merger
and to operate as part of its existing facilities, all of Petal's
facilities; (iii) authorization of Gulf South's request for market-
based rates for storage services to be provided utilizing the combined
Petal and Gulf South storage facilities in interstate commerce; (iv)
authorization for Petal, an existing natural gas company, to abandon
its Part 157 subpart F blanket certificate, issued in Docket No. CP95-
14-000 and its Part 284 subpart G blanket certificate, issued in Docket
No. CP93-69-000; (v) authorization for Gulf South to abandon certain
lease capacity from Petal as authorized in Docket Nos. CP13-96-000 (for
the Southeast Market Expansion Project) and CP13-532-000 (for NNS-A
Service); and (vi) any additional authorizations or waivers necessary
to complete the proposed merger of Petal and Gulf South
    Pursuant to section 157.9 of the Commission's rules, 18 CFR 157.9,
within 90 days of this Notice the Commission staff will either:
Complete its environmental assessment (EA) and place it into the
Commission's public record (eLibrary) for this proceeding; or issue a
Notice of Schedule for Environmental Review. If a Notice of Schedule
for Environmental Review is issued, it will indicate, among other
milestones, the anticipated date for the Commission staff's issuance of
the final environmental impact statement (FEIS) or EA for this
proposal. The filing of the EA in the Commission's public record for
this proceeding or the issuance of a Notice of Schedule for
Environmental Review will serve to notify federal and state agencies of
the timing for the completion of all necessary reviews, and the
subsequent need to complete all federal authorizations within 90 days
of the date of issuance of the Commission staff's FEIS or EA.
    There are two ways to become involved in the Commission's review of
this project. First, any person wishing to obtain legal status by
becoming a party to the proceedings for this project should, on or
before the comment date stated below file with the Federal Energy
Regulatory Commission, 888 First Street NE., Washington, DC 20426, a
motion to intervene in accordance with the requirements of the
Commission's Rules of Practice and Procedure (18 CFR 385.214 or
385.211) and the Regulations under the NGA (18 CFR 157.10). A person
obtaining party status will be placed on the service list maintained by
the Secretary of the Commission and will receive copies of all
documents filed by the applicant and by all other parties. A party must
submit 7 copies of filings made in the proceeding with the Commission
and must mail a copy to the applicant and to every other party. Only
parties to the proceeding can ask for court review of Commission orders
in the proceeding.
    However, a person does not have to intervene in order to have
comments considered. The second way to participate is by filing with
the Secretary of the Commission, as soon as possible, an original and
two copies of comments in support of or in opposition to this project.
The Commission will consider these comments in determining the
appropriate action to be taken, but the filing of a comment alone will
not serve to make the filer a party to the proceeding. The Commission's
rules require that persons filing comments in opposition to the project
provide copies of their protests only to the party or parties directly
involved in the protest.
    Persons who wish to comment only on the environmental review of
this project should submit an original and two copies of their comments
to the Secretary of the Commission. Environmental commentors will be
placed on the Commission's environmental mailing list, will receive
copies of the environmental documents, and will be notified of meetings
associated with the Commission's environmental review process.
Environmental commentors will not be required to serve copies of filed
documents on all other parties. However, the non-party commentors will
not receive copies of all documents filed by other parties or issued by
the Commission (except for the mailing of environmental documents
issued by the Commission) and will not have the right to seek court
review of the Commission's final order.
    The Commission strongly encourages electronic filings of comments,
protests and interventions in lieu of paper using the ``eFiling'' link
at http://www.ferc.gov. Persons unable to file electronically should
submit an original and 5 copies of the protest or intervention to the
Federal Energy Regulatory Commission, 888 First Street NE., Washington,
DC 20426.
    Comment Date: 5:00 p.m. Eastern Time on June 3, 2014.

    Dated: May 13, 2014.
Kimberly D. Bose,
Secretary.
[FR Doc. 2014-11720 Filed 5-20-14; 8:45 am]
BILLING CODE 6717-01-P


[Federal Register Volume 81, Number 247 (Friday, December 23, 2016)]
[Rules and Regulations]
[Pages 94251-94253]
[FR Doc No: 2016-31007]

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DEPARTMENT OF HEALTH AND HUMAN SERVICES

Food and Drug Administration

21 CFR Part 882

[Docket No. FDA-2016-N-4165]

Medical Devices; Neurological Devices; Classification of the
Neurovascular Mechanical Thrombectomy Device for Acute Ischemic Stroke
Treatment

AGENCY: Food and Drug Administration, HHS.

ACTION: Final order.

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SUMMARY: The Food and Drug Administration (FDA) is classifying the
neurovascular mechanical thrombectomy device for acute ischemic stroke
treatment into class II (special controls). The special controls that
will apply to the device are identified in this order and will be part
of the codified language for the neurovascular mechanical thrombectomy
device for acute ischemic stroke treatment's classification. The Agency
is classifying the device into class II (special controls) in order to
provide a reasonable assurance of safety and effectiveness of the
device.

DATES: This order is effective December 23, 2016. The classification
was applicable on September 2, 2016.

FOR FURTHER INFORMATION CONTACT: Leigh Anderson, Center for Devices and
Radiological Health, Food and Drug Administration, 10903 New Hampshire
Ave., Bldg. 66, Rm. 2656, Silver Spring, MD 20993-0002,<PHONE_NUMBER>,
<EMAIL_ADDRESS>
SUPPLEMENTARY INFORMATION:

I. Background

    In accordance with section 513(f)(1) of the Federal Food, Drug, and
Cosmetic Act (the FD&C Act) (21 U.S.C. 360c(f)(1)), devices that were
not in commercial distribution before May 28, 1976 (the date of
enactment of the Medical Device Amendments of 1976), generally referred
to as postamendments devices, are classified automatically by statute
into class III without any FDA

rulemaking process. These devices remain in class III and require
premarket approval unless and until the device is classified or
reclassified into class I or II, or FDA issues an order finding the
device to be substantially equivalent, in accordance with section
513(i) of the FD&C Act, to a predicate device that does not require
premarket approval. The Agency determines whether new devices are
substantially equivalent to predicate devices by means of premarket
notification procedures in section 510(k) of the FD&C Act (21 U.S.C.
360(k)) and part 807 (21 CFR part 807) of the regulations.
    Section 513(f)(2) of the FD&C Act, as amended by section 607 of the
Food and Drug Administration Safety and Innovation Act (Pub. L. 112-
144), provides two procedures by which a person may request FDA to
classify a device under the criteria set forth in section 513(a)(1).
Under the first procedure, the person submits a premarket notification
under section 510(k) of the FD&C Act for a device that has not
previously been classified and, within 30 days of receiving an order
classifying the device into class III under section 513(f)(1) of the
FD&C Act, the person requests a classification under section 513(f)(2).
Under the second procedure, rather than first submitting a premarket
notification under section 510(k) of the FD&C Act and then a request
for classification under the first procedure, the person determines
that there is no legally marketed device upon which to base a
determination of substantial equivalence and requests a classification
under section 513(f)(2) of the FD&C Act. If the person submits a
request to classify the device under this second procedure, FDA may
decline to undertake the classification request if FDA identifies a
legally marketed device that could provide a reasonable basis for
review of substantial equivalence with the device or if FDA determines
that the device submitted is not of ``low-moderate risk'' or that
general controls would be inadequate to control the risks and special
controls to mitigate the risks cannot be developed.
    In response to a request to classify a device under either
procedure provided by section 513(f)(2) of the FD&C Act, FDA shall
classify the device by written order within 120 days. This
classification will be the initial classification of the device.
    On October 26, 2015, Concentric Medical, Inc., submitted a request
for classification of the Trevo ProVue and XP ProVue Retrievers (Trevo
Retrievers) under section 513(f)(2) of the FD&C Act.
    In accordance with section 513(f)(2) of the FD&C Act, FDA reviewed
the request in order to classify the device under the criteria for
classification set forth in section 513(a)(1). FDA classifies devices
into class II if general controls by themselves are insufficient to
provide reasonable assurance of safety and effectiveness, but there is
sufficient information to establish special controls to provide
reasonable assurance of the safety and effectiveness of the device for
its intended use. After review of the information submitted in the
request, FDA determined that the device can be classified into class II
with the establishment of special controls. FDA believes these special
controls, in addition to general controls, will provide reasonable
assurance of the safety and effectiveness of the device.
    Therefore, on September 2, 2016, FDA issued an order to the
requestor classifying the device into class II. FDA is codifying the
classification of the device by adding 21 CFR 882.5600.
    Following the effective date of this final classification order,
any firm submitting a premarket notification (510(k)) for a
neurovascular mechanical thrombectomy device for acute ischemic stroke
treatment will need to comply with the special controls named in this
final order.
    The device is assigned the generic name neurovascular mechanical
thrombectomy device for acute ischemic stroke treatment, and it is
identified as a prescription device used in the treatment of acute
ischemic stroke to improve clinical outcomes. The device is delivered
into the neurovasculature with an endovascular approach, mechanically
removes thrombus from the body, and restores blood flow in the
neurovasculature.
    FDA has identified the following risks to health associated
specifically with this type of device, as well as the measures required
to mitigate these risks in table 1.

Table 1--Neurovascular Mechanical Thrombectomy Device for Acute Ischemic
             Stroke Treatment Risks and Mitigation Measures
------------------------------------------------------------------------
          Identified risk                    Mitigation measure
------------------------------------------------------------------------
Adverse Tissue Reaction...........  Biocompatibility Evaluation.
Infection.........................  Sterility Testing, Shelf-Life
                                     Testing, Labeling.
Tissue or Vessel Damage:
     Dissection...........  Non-clinical Performance Testing.
     Perforation..........  Clinical Performance Testing.
     Hemorrhage...........  Labeling.
Stroke Progression................  Non-clinical Performance Testing,
                                     Clinical Performance Testing,
                                     Labeling.
Emboli............................  Non-clinical Performance Testing,
                                     Clinical Performance Testing,
                                     Labeling.
------------------------------------------------------------------------

    FDA believes that the special controls, in combination with the
general controls, address these risks to health and provide reasonable
assurance of the safety and effectiveness.
    Neurovascular mechanical thrombectomy device for acute ischemic
stroke treatment devices are not safe for use except under the
supervision of a practitioner licensed by law to direct the use of the
device. As such, the device is a prescription device and must satisfy
prescription labeling requirements (see 21 CFR 801.109 Prescription
devices).
    Section 510(m) of the FD&C Act provides that FDA may exempt a class
II device from the premarket notification requirements under section
510(k) if FDA determines that premarket notification is not necessary
to provide reasonable assurance of the safety and effectiveness of the
device. For this type of device, FDA has determined that premarket
notification is necessary to provide reasonable assurance of the safety
and effectiveness of the device. Therefore, this device type is not
exempt from premarket notification requirements. Persons who intend to
market this type of device must submit to FDA a premarket notification,
prior to marketing the device, which contains information about the
neurovascular mechanical thrombectomy device for acute ischemic stroke
treatment they intend to market.

II. Analysis of Environmental Impact

    The Agency has determined under 21 CFR 25.34(b) that this action is
of a type that does not individually or cumulatively have a significant
effect on the human environment. Therefore, neither an environmental
assessment nor an environmental impact statement is required.

III. Paperwork Reduction Act of 1995

    This final order establishes special controls that refer to
previously approved collections of information found in other FDA
regulations. These collections of information are subject to review by
the Office of Management and Budget (OMB) under the Paperwork Reduction
Act of 1995 (44 U.S.C. 3501-3520). The collections of information in
part 807, subpart E, regarding premarket notification submissions, have
been approved under OMB control number 0910-0120, and the collections
of information in 21 CFR part 801, regarding labeling, have been
approved under OMB control number 0910-0485.

List of Subjects in 21 CFR Part 882

    Medical devices, Neurological devices.

    Therefore, under the Federal Food, Drug, and Cosmetic Act and under
authority delegated to the Commissioner of Food and Drugs, 21 CFR part
882 is amended as follows:

PART 882--NEUROLOGICAL DEVICES

0
1. The authority citation for part 882 continues to read as follows:

    Authority: 21 U.S.C. 351, 360, 360c, 360e, 360j, 360l, 371.

0
2. Add Sec.  882.5600 to subpart F to read as follows:

Sec.  882.5600   Neurovascular mechanical thrombectomy device for acute
ischemic stroke treatment.

    (a) Identification. A neurovascular mechanical thrombectomy device
for acute ischemic stroke treatment is a prescription device used in
the treatment of acute ischemic stroke to improve clinical outcomes.
The device is delivered into the neurovasculature with an endovascular
approach, mechanically removes thrombus from the body, and restores
blood flow in the neurovasculature.
    (b) Classification. Class II (special controls). The special
controls for this device are:
    (1) The patient contacting components of the device must be
demonstrated to be biocompatible.
    (2) Non-clinical performance testing must demonstrate that the
device performs as intended under anticipated conditions of use,
including:
    (i) Mechanical testing to demonstrate the device can withstand
anticipated tensile, torsional, and compressive forces.
    (ii) Mechanical testing to evaluate the radial forces exerted by
the device.
    (iii) Non-clinical testing to verify the dimensions of the device.
    (iv) Non-clinical testing must demonstrate the device can be
delivered to the target location in the neurovasculature and retrieve
simulated thrombus under simulated use conditions.
    (v) Non-clinical testing must demonstrate the device is radiopaque
and can be visualized.
    (vi) Non-clinical testing must evaluate the coating integrity and
particulates under simulated use conditions.
    (vii) Animal testing must evaluate the safety of the device,
including damage to the vessels or tissue under anticipated use
conditions.
    (3) Performance data must support the sterility and pyrogenicity of
the patient contacting components of the device.
    (4) Performance data must support the shelf-life of the device by
demonstrating continued sterility, package integrity, and device
functionality over the specified shelf-life.
    (5) Clinical performance testing of the device must demonstrate the
device performs as intended for use in the treatment of acute ischemic
stroke and must capture any adverse events associated with the device
and procedure.
    (6) The labeling must include:
    (i) Information on the specific patient population for which the
device is intended for use in the treatment of acute ischemic stroke,
including but not limited to, specifying time from symptom onset,
vessels or location of the neurovasculature that can be accessed for
treatment, and limitations on core infarct size.
    (ii) Detailed instructions on proper device preparation and use for
thrombus retrieval from the neurovasculature.
    (iii) A summary of the clinical testing results, including a
detailed summary of the device- and procedure-related complications and
adverse events.
    (iv) A shelf life.

    Dated: December 19, 2016.
Leslie Kux,
Associate Commissioner for Policy.
[FR Doc. 2016-31007 Filed 12-22-16; 8:45 am]
BILLING CODE 4164-01-P


[Federal Register Volume 76, Number 26 (Tuesday, February 8, 2011)]
[Rules and Regulations]
[Page 6694]
[FR Doc No: 2011-2688]

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DEPARTMENT OF HOMELAND SECURITY

Coast Guard

33 CFR Part 117

[Docket Number USCG-2011-0029]

Drawbridge Operation Regulation; Upper Mississippi River, Keokuk,
IA

AGENCY: Coast Guard, DHS.

ACTION: Notice of temporary deviation from regulations.

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SUMMARY: The Commander, Eighth Coast Guard District, has issued a
temporary deviation from the regulation governing the operation of the
Keokuk Drawbridge across the Upper Mississippi River, mile 364.0, at
Keokuk, Iowa. The deviation is necessary to allow the bridge owner time
to perform the needed maintenance and repairs to the bridge that is
essential to the continued safe operation of the drawbridge. This
deviation allows the bridge to remain in the closed-to-navigation
position for thirty days.

DATES: This deviation is effective from 12:01 a.m., January 30, 2011
until 9 a.m., February 28, 2011.

ADDRESSES: Documents indicated in this preamble as being available in
the docket are part of docket USCG-2011-0029 and are available online
by going to http://www.regulations.gov, inserting USCG-2011-0029 in the
``Keyword'' box and then clicking ``Search''. They are also available
for inspection or copying at the Docket Management Facility (M-30),
U.S. Department of Transportation, West Building Ground Floor, Room
W12-140, 1200 New Jersey Avenue, SE., Washington, DC 20590, between 9
a.m. and 5 p.m., Monday through Friday, except Federal holidays.

FOR FURTHER INFORMATION CONTACT: If you have questions on this rule,
call or email Eric A. Washburn, Bridge Administrator, Western Rivers,
Coast Guard; telephone<PHONE_NUMBER>, e-mail<EMAIL_ADDRESS>
If you have questions on viewing the docket, call Renee V. Wright,
Program Manager, Docket Operations, telephone<PHONE_NUMBER>.

SUPPLEMENTARY INFORMATION: The City of Keokuk, Iowa requested a
temporary deviation for the Keokuk Drawbridge, across the Upper
Mississippi River, mile 364.0, at Keokuk, Iowa to remain in the closed-
to-navigation position in order to facilitate needed bridge maintenance
and repairs. The Keokuk Drawbridge currently operates in accordance
with 33 CFR 117.5, which states the general requirement that
drawbridges shall open promptly and fully for the passage of vessels
when a request to open is given in accordance with the subpart. This
deviation allows the bridge to remain in the closed-to-navigation
position from 12:01 a.m., January 30, 2011 until 9 a.m., February 28,
2011.
    There are no alternate routes for vessels transiting this section
of the Upper Mississippi River.
    Winter conditions on the Upper Mississippi River coupled with the
closure of U.S. Army Corps of Engineer's Lock 20, mile 343.2, Lock 21,
mile 324.9, and Lock 22, mile 301.2 from January 30, 2011 to February
28, 2011 will preclude any significant navigation demands for the
drawspan to open.
    The Keokuk Drawbridge, in the closed-to-navigation position,
provides a vertical clearance of 25.0 feet above normal pool.
Navigation on the waterway consists primarily of commercial tows and
recreational watercraft. This temporary deviation has been coordinated
with waterway users. No objections were received.
    In accordance with 33 CFR 117.35(e), the drawbridge must return to
its regular operating schedule immediately at the end of the designated
time period. This deviation from the operating regulations is
authorized under 33 CFR 117.35.

    Dated: January 24, 2011.
Eric A. Washburn,
Bridge Administrator.
[FR Doc. 2011-2688 Filed 2-7-11; 8:45 am]
BILLING CODE 9110-04-P


Hello-

I have read through the various responses and do not feel it necessary to revise my previously submitted answers to the charge questions.

Have a good weekend!

Joel


you could look for specific responses. And we've developed a preliminary proposal to do that. Or as Professor Van Der Kraak indicated, you could look for more general responses. And some work he's done in his laboratory using things like differential display, are useful techniques that we could ask the question, is anything change in that axis and work from that position. So that's another idea you might think about.

But we couldn't agree more. I don't think at this point in time we can key in on any one specific mode of action. Absolutely.

DR. ROBERTS: Dr. Coats. Oh, Dr. Solomon, did you want to respond also?

DR. SOLOMON: Just to follow up. And, Fumio, thank you.

The effects in the Sprague Dawley rat were reproducible and consistent at different times, different labs, et cetera. Once the mechanism was understood in the fact that it was specific to Sprague Dawleys that, I think, was very helpful. I was at the SAP here in Crystal City a few years ago to listen to that discussion, so it was very interesting to see it being used in a risk assessment context.

But we haven't yet, at least in our own hands, been able to get a 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 197 reproducible robust response that we can use to track back to a potential mechanism. Given that, obviously, we would love to do it.

DR. ROBERTS: Dr. Coats and then Dr. Isom.

DR. COATS: In the field studies, I guess it was particularly in the South African one, there are quite a few other triazine or metabolites note there. I was curious about the water levels that are reported. Are they filtered samples or unfiltered? And are there sediment values, and do you think those would be important or not?

DR. DU PREEZ: If you're referring to if they're filtered or not,

you're referring to the weekly sampling of the water?

DR. COATS: The water, yes.

DR. DU PREEZ: No. Well, I'm not  I wasn't involved in those analyses. But to the best of my knowledge, they were not filtered.

DR. SOLOMON: The analyses were done by Piet Johnson from Rensburg at the Potchefstroom University using GC mass spec. And he used SPE cartridges and liquid liquid extraction using both methods to confirm. And then he used unfiltered water. But it was filtered through the SPE cartridges which were then eluted. So anything that was trapped on a solid, would have been extracted. And then the liquid liquid were extracted from the direct water samples, 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 198 no filtration.

Sediments samples were looked at and no pesticides were detected in the sediment samples. Given the KD binding constance of atrazine which is relatively water soluble, that's consistent with what we would expect, that nothing was present in the sediments.

DR. COATS: Thanks.

DR. ROBERTS: Dr. Isom.

DR. ISOM: Dr. Giesy, I'd like to follow up on your comment you just made a few moments ago about aromatase. The EPA is proposing that that will be the second tier, that is, if we can pass the first tier, to start to look at that. And you made the comment that perhaps  I'll paraphrase you that perhaps we're looking in the wrong direction.

Yet if you look at the literature Sanderson's group has shown in human cells, tumor lines, that it atrazine induces aromatase. And I think at higher concentrations perhaps even inhibits so that you can get that inverted U shaped dose response curve.

I'd like to have you follow up on that comment then and any other studies that you have done, could you describe those with that enzyme?

DR. GIESY: Absolutely. Well, yeah, I'm an author on those 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 199 papers; so I do know about them. I think it's important maybe to explain a couple things about how those studies were done and why they were done and I'll do that.

But let me answer your question very specifically. I think what I meant was, if we look at aromatase, if there are open issues there, I think we need to take a fundamentally different approach than we've taken previously. Not to dismiss it entirely. But if we do go ahead with that, and I do agree with EPA that it shouldn't be first tier. But if we go ahead with it, I think we have to do it in a way that we look in specific tissues. And we can do that by QRTPCR.

In our laboratory, we've now developed a molecular beacons to do that. So we can do the quantitative PCR. I think it would be important if we follow that mechanism up that we do it through the developmental stages where we know there are changes in aromatase expression and we do it in a tissue specific way.

First of all, my comments were not to just look in a gross way because I don't think we'll see. So I think it is stage dependent and it's tissue specific. So if we do go that way, that's what we need to do.

But at the same time, based on the literature we do know, it would be appropriate at the same time to look at other plausible 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 200 mechanisms. And one of the other plausible mechanisms is through the hypothalamic pituitary gonadal axis. I think we've got enough evidence as Professor Matsumura point out to not completely dismiss that.

Why do I say that? Because, in fact, I think some of the effects that we do observe are more consistent relative to what can be caused through that mechanism of action relative to testosterone depression for instance.

Now, beyond that, the studies that Thomas did when he was in my laboratory were designed specifically to try to understand why we observing what we considered to be anomalous results. And that is we knew that atrazine didn't bind, at least in our hands, to the estrogen receptor. But in some cell lines, we did see what looked like estrogenic responses. The question was why.

Now the first experiment I had Thomas do was just to dose them with perpronobutoxide because I though maybe what we were looking at were metabolites that were being formed, hydroxy metabolites which, in fact, might be estrogenic. Subsequently, we tested all those. They basically weren't estrogenic. But in doing that, we thought we were knocking out the MFO activity that might form the metabolites; when, in fact, what we were doing most likely was 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 201 knocking out the aromatase.

So we did those studies in part to understand mechanistically why we were observing this effect in vitro. They were done at pretty high concentrations. They were done at 30 micromolar, which is about 6 parts per million in the tissue. And they were done  and the result, then, of that, too, was  and we looked at message. We looked at expression, and we look at the activity. Depending on which one you looked at, the range of response that we got was somewhere between two to four fold. Not a huge response.

So I don't dismiss it entirely because I think we haven't investigated at these time specific and tissue specific responses. So if we do go ahead, that's where we need to look. But I really think based on the literature that's out there, we shouldn't dismiss the other potential pathway at the same time. That was my point. Not that you shouldn't consider it at all. But how you do it if you do it and not forget about this other pathway which I think is also consistent with the results that were observed.

DR. ROBERTS: Dr. Kendall, I think that before your panel continues with the next aspect of your public comments, my panel needs a break.

DR. KENDALL: Mr. Chairman, our panel yields to your panel. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 202 Let's take a break.

DR. ROBERTS: Let's take a 15 minute break.

( Break; session resumed at 3: 45 p. m.)

DR. ROBERTS: Before the next round of presentations, Dr.

Green had a question as a follow up from our discussion right before the break.

DR. KENDALL: Very good.

DR. ROBERTS: Dr. Green.

DR. GREEN: Yes. I just have a quick question for clarification to Dr. Du Preez. You mentioned that some of the answers to the questions I posed earlier were available in a document that was submitted. Can you clarify which document that was so I can go back and make sure we all have the same details?

DR. DU PREEZ: The documents that we've submitted to the Panel, that would be SA01B and SA01C.

DR. GREEN: And in those documents, there are the details regarding water quality analysis on the ponds, frequency

DR. DU PREEZ: Everything is in there.

DR. GREEN:  changing and rainfall and that sort of thing.

Okay.

DR. DU PREEZ: Including the raw data. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 203 DR. GREEN: Thank you.

DR. ROBERTS: Let's then proceed with the next round of presentations. The Panel will have opportunity to ask more questions after those.

DR. KENDALL: Thank you, Mr. Chairman. As we earlier indicated, we would lead off with the core presentation by Dr. Van Der Kraak. And we, as a panel, are very impressed with your panel.

You came very well prepared. And we do accept the criticism and welcome the comments and have gained a great deal of future insight related to how we would proceed with our research.

I think the panel can relate to particularly the last couple of years, spent a lot of effort and we've amassed now a great deal of information and a lot of manuscripts are stacking up that are going out for review as we speak. So this discussion, interaction is welcomed.

And from our core presentation, our panel is continuing, as I speak, to evolve data. And I wanted each of them to have a brief opportunity to  I didn't realize we would have as many questions as we did. But we welcome them. But I wanted each panel member to have a chance to briefly address this SAP to give you an opportunity to further understand what their contribution was to our overall 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 204 effort. And I emphasize our efforts have been collective. And our publication, we have discussions on our publications and they are collective.

And I also wanted to make a comment about Dr. Bob Silken.

We probably ought to make him an honorary panel member. He's made himself so available. He sits in on all panel meetings now,

conference calls and so on. And he's very gracious with his contribution, although he drives us crazy sometimes related to all of his questions. But I think it's made our science better.

But anyway, I'd like to begin. Dr. Van Der Kraak gave our core presentation. Dr. Giesy, I'd like for him to comment as he feels appropriate. And the Panel is welcome to ask any questions of our scientists as we proceed. And then we will close our comments today after this period by offering you some responses to the White Paper questions from our panel that can be shared with you. So Dr. Giesy.

DR. GIESY: First of all, I'd just like to say I think this is an excellent panel. I'm very impressed by how well prepared everyone is, but you're all experts. And, two, to reinforce that I think the EPA White Paper was an excellent document. They have a really tough task to do, to balance all this and try to find a way forward. So I thought they did an excellent job. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 205 We've been involved in this for a couple of years now. I guess what I've learned from that is that we started from not much and have made a lot of progress. But it's certainly a work in progress.

Certainly things aren't perfect and can be developed much more in the future.

My main interest in really in the mechanism of action, what is the plausible mechanism for how atrazine may cause effects so we can put into context what are the right endpoints to measure, what we refer to in risk assessment as measurement endpoints, what would be the most sensitive and relevant endpoint to measure that then could go into an assessment endpoint; which is ultimately what the EPA has to deal with. And so I feel strongly that we do need to know what the critical mode of action is. And that's my interest. So anything relative to where we are, where we go relative to that is something I'm interested in.

So we've set up some studies. When we started, we wanted to work with some native species. The protocols aren't all completely worked out for that. One of the grad students in my lab, Katie Cody,

did a lot of work just to be able to figure out how to do a study with green frogs. Having done that, if the EPA asks me should we use green frogs, I'd say no because there are some real issues with time to 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 206 metamorphosis for green frogs.

But I think we have learned a lot as a panel. And I think as a scientific community, things are unfolding in this whole endocrine disruptor area.

With that, you know, I think this panel can have a huge impact on the future, where we go, where EPA goes, where the science goes.

And I think that would just be great. But I don't have any other specific comments.

We do have one ongoing field study. What Dr. Kendall wanted us to do was get you up to speed on anything that's been done since the reports that we've provided to you. From my laboratory, there isn't anything else really. But we do have an ongoing field study. We have students in the field right now that will continue with all the warts and imperfections of trying to do field work.

But other than that, you have everything that we've done at this time. And if you have any further questions, I'd be glad to try to field them.

DR. ROBERTS: Any question? Dr. Kelley.

DR. KELLEY: So I'd like to comment and I think this also applies to Dr. Hayes about the use of this laryngeal dilator muscle cross sectional areas and assay. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 207 So we studies the development of the larynx in Xenopus. And there are two features that I think are important for your use of it as an assay for masculinization. First of all, let me point out, it's a very good assay for masculinization as you show yourselves. It's a very androgen sensitive organ. And it's certainly extremely sexually dimorphic in adulthood.

So the cross sectional area that you guys measure represents two things, the size of muscle fibers and the number of muscle fibers.

And we actually never measured cross sectional area because it confounds the two. We looked at number of muscle fibers. And we also showed that the ability of androgen to cause growth of the larynx, both hypertrophy and hyperplasia, was dependent on prior exposure to thyroid hormone. So in an assay system where your animals are taking a long time to metamorphose  all right. So let's say normal is two weeks. These guys are taking a month and a half or longer, we've also shown that the animals are exposed to androgen during that period. So it's possible that variability in time to metamorphosis can contributed substantively to variability in whether you see a sexually differentiated laryngeal cross sectional area.

Now, that all washes out by the time the animal is three months old. So you don't have to worry about that any more. So I make a 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 208 plea for you guys to stop looking at laryngeal cross sectional area at Stage 66. I don't think it's very appropriate time. And just give it a couple months more.

We divided post metamorphic development into six stages that are very well characterized by laryngeal weight. You can standardize all of your animals no matter how long it's taking them by body weight and laryngeal weight to those stages. And it should be possible to come up with some well characterized, low variability assay for whether atrazine has an effect on masculinization by using that assay at a slightly later time period.

Anyway, that's my input on the laryngeal bioassay which has, in fact, been variable. But I think it's been variable for reasons of rearing variability in terms of length to metamorphosis in the studies that pretty much every has done. And that makes concordance between the studies very difficult.

DR. KENDALL: Good points.

DR. GIESY: Excellent point.

DR. ROBERTS: Any other questions for Dr. Giesy? Okay.

Let's move on.

DR. KENDALL: I did want to make sure of one thing. Dr.

Kelley, you mentioned two references, two papers on the hormonal 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 209 measurements. And we need to make sure we get those for Dr. Giesy.

DR. KELLEY: I have the papers. I'd be happy to give them to you.

DR. KENDALL: Very good.

DR. CARR: I really don't have any new data to present. And I don't have any additional specific comments that weren't addressed in the previous session.

I did have a general comment. I think there was discussion earlier this morning regarding the subjectivity of some of the gonadal assessment. And you can only come to that conclusion when you look at all the different terminology that's used to assess intersex,

hermaphrodite, et cetera.

But one thing to remember in our study and most, if not all, of the other studies, is that these are analyses and the treatments are conducted with no knowledge of what the treatments actually are.

These are blind tests in essence. And so I feel very confident about our data in terms of when we see something that is unusual and this is an intersex, it's very different from what we would expect to see in a normal male or female. So I think that reduces some of the subjectivity at least in Xenopus.

We have finished an experiment in Rana pipiens that the things 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 210 were a little bit different there. We don't have data to share on that yet.

But that's really all I had to say. And I'll be happy to answer any additional questions.

DR. ROBERTS: Any questions for Dr. Carr? Great. Thanks.

Dr. Smith, did you want to proffer something?

DR. SMITH: We have continued the work in Iowa, the field study in Iowa, primarily, the laboratory component of it. From the histology standpoint, we have found so far one testicular, one animal with testicular oocyte which turns out to be about .6 percent of the total number of animals that we have analyzed.

We have also taken the plasma for testosterone analysis and gone over the period of collection because, as I said earlier, the representative time representing Period 1, 2, and 3 which would be late spring, early summer, later summer and early fall. And the indication there is that there is time dependent change in plasma testosterone concentration. So time becomes a variable and a factor that has to be taken into consideration. However, over the period,

there is no significant difference for plasma testosterone when you compare the adult animals during the specific period.

The other observation is that during that period, there is 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 211 significant difference between the adult and the juvenile plasma testosterone concentration using a 60 gram cut off point as the difference between the adult versus the juvenile for that comparison.

We are presently in the process of utilizing the steriological approach that we use for the South African Xenopus laevis adult testicular histological analysis to evaluate the fractional volume sperm, spermatocytes, spermatogonia and what we consider as other cells, which includes blood vessels, connective tissue. And that data will be made available pretty soon as to the contribution from that standpoint.

And that's where we are presently. I'll entertain any questions from you.

DR. ROBERTS: Questions for Dr. Smith? Great. Thanks for the update. Dr. Du Preez.

DR. DU PREEZ: Last night I prepared a quick PowerPoint presentation, so I'm going to walk you through a couple of slides.

This a picture that I've quickly inserted here to give you an image. This is one of the larger ponds that we've used. This specific pond was referred to as Site E6.

We did four studies in South Africa. SA01A was the evaluation of the sites where we characterized the different sites, the 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 212 populations, the mark and recapture studies. And based on this study,

we identified five experimental sites and three reference sites.

Phase B, SA01B, was the study of a one field use season. We on a weekly basis at first and later bi weekly collected water samples that were analyzed. Elemental scans were performed on water samples as well as on sediment samples. And then SA01C, where we've collected the specimens. The blood samples were collected,

shipped over to Dr. Giesy's lab where they've conducted the hormone aromatase analyses. Gonads were shipped over to Dr. Smith's lab.

And he just mentioned the histology.

Then SA01D is the study on the microcosm that I'm going to expand a little bit more on. And then just for interest, I've been busy with the first study at this stage where we're going to look at the reproductive cycle of Xenopus. Our period of months, we're going to quantify the nuptial pads, the cloacal folds, the hormone levels, and so forth.

If we now focus on the microcosm study, this formed part of a thesis of Alaric Uester. We had a microcosm. We had 12 ponds, and this was the layout. Three reference ponds, three ponds with one microgram per liter, three with 10, and three with 25.

These were initially earthworm pits that we referred to. We 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 213 emptied them, lined them with a membrane, filled them with water,

add some microphytes, leave them for six months to stabilize, and then started the experiment.

Individual microcosms were covered with that frame with hail netting. And that would be to keep predators out, primarily dragonfly, because the dragonfly larvae is the one animal that you don't want in a microcosm, and also birds.

Just a couple of tadpoles there to show that the tadpoles schooled in my opinion in a natural fashion. And at Stage 66 animal,

they are in the water. The water quality from measured concentrations and from a visual inspection was very good.

And there is a set of microcosms. What I want to point out in this slide is, from a logistical point, we had a problem that not all the ponds were exposed to the same amount of sunlight. You can see that these first four ponds are shaded here by a tree. Then there are a couple of ponds in the middle that received more sunlight. And then on the other side, again, two ponds that were semi shaded.

The water temperature was much lower than you would expect in a natural pond. And this had an effect on the development. And we, indeed, experienced a delayed development. But that's, in my opinion, not a concern in this study. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 214 This is the actual recorded atrazine levels. At one instance, in one of the reference ponds, we did detect 0.1 microgram per liter atrazine. Again, I was not worried about this because two weeks later it was gone. I think it was either an error or contamination. But to make sure, I went back and I split the reference pond samples in two.

I compared the specimens collected from that specific Pond No. 3 with the other reference samples, and there was no difference.

Then on cross morphology, that would be a normal male, the female. And this is the type of deformities that we've observed. And the only deformity that we found was discontinued testes.

Now, I'm actually a parasitologist working on parasites of amphibians. And for over 15 years or more, I've literally opened up thousands of frogs. And this is what you see from time to time, a single testis. Usually, when there's a single testis, it would be larger.

You do find discontinued testis, but I've never come across a true hermaphrodite in frogs that I've collected. And I've worked in both corn growing areas where atrazine would be applied and areas more pristine reserves and so forth.

Based on the gross morphology, we've observed a 4 percent in the reference, 1.3 in the 1 microgram per liter, .6 in the 10, and 3.7 in the 25 microgram per liter. So, again, no statistical difference here. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 215 And this is up to the point where we've reported in the study SA01D. In the recent weeks, we've conducted the histology. And I'm quickly going to report on our findings there.

This is my slave, Alaric Uester. We selected 54 specimens per concentration, randomly selected. That adds up to 216 of the 600 specimens selected for the gross morphology. Six specimens were lost in the preparation for the histology. 120 to 200 sections per specimen, four to six slides per specimen. That all adds up to more than a thousand slides. And 31,000 histological sections that were examined individually. So that's why I'm not very popular with my students at this stage.

What we've observed and what I want to point, testicular oocyte would appear and disappear in seven slides sectioned at 6 micrometer.

So what I wanted to stress with this is it's not good to check every 20th section. You have to check every one if you want to detect the testicular oocytes.

A testis with a testicular oocyte there, a cross section,

longitudinally section, and the ovary. And you can clearly the lumen in the middle.

This is what the histology revealed. No less than 56 percent of the animals from the reference ponds had testicular oocytes. Again, 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 216 56 in the 1 microgram, 58 in 10, and 38 in the 25.

Just this past week, we did another study where we looked at grow outs. Now the rate columns represent Stage 66 and the blue,

10 month grow outs. And there is a reduction in the number of oocytes. Now I must point out that this blue set, the 10 month grow outs is only 10 specimens per concentration.

Then we calculated the number of specimens with testicular oocytes where we found the oocytes in the single testis or both testes and no statistical significant difference there.

If we now look at the mean number of oocytes per specimen with testicular oocytes, we find that there is an average of about 10 oocytes per specimen. But there is a significant reduction if we look at the 10 month old grow outs. And then if we split up the sample and divide the number of oocytes in categories, zero, 1, 2 to 10, 11 to 12 and so on, 51 to 60, we find the following. And, again, not the significant reduction in the number of oocytes.

The maximum number of oocytes in the Stage 66 samples was in this 25 microgram per liter. And that was 58 oocytes in a Stage 66 Xenopus. The maximum for the 10 month grow out was 5. Only one specimen had 5, three had 2, and the rest had a single oocyte.

So from this, tadpoles developed slow as a result of cold water. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 217 I've already addressed that. Ponds were partially shaded, covered with hail netting and did not have a shallower part as you would find in a natural pond. Often these Xenopus in a natural pond would school in the shallower water.

I've already addressed this point of the atrazine that was detected in this one reference pond. Again, no reason for concern there.

No gonadal abnormalities were observed in the females. No hermaphrodites were observed. Males showed a low percentage of deformities at a cross morphology level. A high percentage of frogs had one or more testicular oocytes at all concentrations, but there was no dose response. And then a significant reduction in the number of testicular oocytes as metamorphs grow older.

And from the South African studies, for me the take home message would that atrazine does not appear to affect the gonadal development of Xenopus laevis at environmentally relevant concentrations. The second point would be that, in my opinion, one would expect that if atrazine had a negative or adverse effect on Xenopus laevis, it would reflect in the population dynamics after four decades of intensive use of atrazine. And in previous decades, it's been documented that in South Africa they used atrazine by the tons. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 218 I mean they really applied it.

Now days, they use more conservative, recommended amounts.

But surely, if there was something to be worried about, you would see it in the Xenopus populations out there.

And then in my opinion, field studies and microcosm studies does have its limitations as greatly pointed out by the EPA's White Paper. But I don't think we should underestimate the value of field or microcosm studies. Because in the end, the question is what's happening out there.

Thank you.

DR. ROBERTS: All right. Any questions from panel members.

Dr. Green.

DR. GREEN: I think you could probably anticipate this question from me. How cold was the water? And you didn't mention anywhere in this document the conditions of the water quality, pH conductivity. If someone were to try and reproduce this experiment,

it would be very important to have that information if you're going to reproduce this microcosm in a lab.

DR. DU PREEZ: That's correct. That data is available. What we've handed out is a handout to summarize this presentation. That's not the full report. In the full report, we have all the other data. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 219 DR. GREEN: But in my opinion, I think the water quality and the parameters that define this microcosm are very important,

especially to a laboratory Xenopus person. So what was the temperature on this?

DR. DU PREEZ: Too cold to swim in. We did not measure water temperature on an hourly basis for example. We measured in this study at 10 o'clock in the morning when we took the water sample to check for atrazine. And that was in the low teens, around 10 to 14,

which is lower than you find in the natural population.

DR. ROBERTS: Any other questions. Yes, Dr. Kelley.

DR. KELLEY: If you wouldn't mind my asking you a question about the previous field study. I'm looking at the mass of the frogs on page 59 in the study where you collected them from the three reference ponds and the five experimental sites. And I just want to make sure I'm reading this right.

So I think the goal was to get 20 of each sex from each pond.

But it looks to me like that was difficult to achieve at the experimental sites. Was there another reason for that that you could think of?

DR. DU PREEZ: I was really upset when I wanted to collect the specimens, especially from Site E1. Because with this excessive 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 220 rains, some catfish were washed into that pond and literally wiped out the Xenopus population in that pond to such an extent that we had difficulty to collect the target number of specimens.

DR. KELLEY: So your feeling that the difficulty in collecting at this sites where you have full representation in your reference sites from the cow pastures, but a sparser representation for the experimental site was due to predation.

DR. DU PREEZ: That's my opinion.

DR. KELLEY: Thank you.

DR. ROBERTS: Any other questions? If not, let's move on to Dr. Gross.

DR. GROSS: My comments will also be relatively brief. We have been coordinating now for the past year and a half the studies looking at another one of the mini field studies, looking at the cane toad in sugar cane agricultural areas in South Florida. Those studies are currently continuing and ongoing, so we have little to add at this point that's new that the Panel has not already seen the documents provided.

I think several things can be noted, though, from the materials that have been provided. First, I think as indicated in EPA's review which we fully agree with, these experiments were simply preliminary 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 221 examinations, mostly fact finding, range finding, site identification,

species identifications kind of studies. And, therefore, it's very difficult from these studies if not impossible to draw any conclusions relative to causation of single chemicals or even mixtures of chemicals or other factors.

None the less, the studies that we've seen do demonstrated some effects. We think that these effects, though, are vastly different in this species compared to what else has been reported for Xenopus or the various ranid species that had been examined because we're looking at a toad species, the cane toad, which has a bitters organ and rudimentary ovarian structure, which basically, I think, allows a differential kind of response. We believe these responses currently to be mostly in the adult or sub adult rather than during the metamorphic phases that have been described previously for the other species.

None the less as I indicated, we are continuing with these studies. Actually, we've been continuing since February, have little data yet to add because these studies are ongoing. I think the comments of the panel today have been mostly addressed as we designed this second phase of study, the second tier; and, hopefully,

we will have better answers in the next year.

But I'll be happy to entertain any questions you might have 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 222 relative to the studies we've conducted on cane toads.

DR. ROBERTS: Any questions for Dr. Gross? All right. Dr.

Solomon.

DR. SOLOMON: I'm sort of wrapping up this with a few brief comments. And my role in the panel, apart from helping formulate the reports, was to bring the concept of risk assessment to the data that we were developing. And as we heard earlier from Dr. Bradbury and others, risk assessment involves a component of integration of exposure and effects data. And as you probably very well know, there is an excellent data base for atrazine concentrations from the ecosystem through the efforts of the U. S. Geological Survey and other labs. We have very large data sets in the hundreds of thousands of data points. Maybe not hundreds of thousands, but certainly tens of thousands of data points. So there is a good data base to go out there to work on a measured concentrations to compare to effect concentrations.

We also have good modeling data that allows us to estimate exposures in areas where we don't have good measured values and helps to get a tighter definition of temporal variation that we don't get from sampling once every week or two weeks.

I've been somewhat frustrated in this because the other side of 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 223 the equation, the effect side, has not materialized to the point where I can use this in the risk assessment process. And I think Dr. Van Der Kraak pointed out in his slide that we need robust and consistent concentration responses that we can feed into the risk assessment process to determine probablistically or deterministically what types of responses we might expect to see in the environment. So that's been my frustration. And, hopefully, it's a frustration on the Panel as well. I certainly heard that when I read the White Paper as well, that there was insufficient data at this point to do a risk assessment.

Thank you very much.

DR. ROBERTS: Any questions for Dr. Solomon? Yes, Dr. Van Der Kraak.

DR. VAN DER KRAAK: They overlooked me.

Just to comment in terms of some ongoing studies that are occurring in my laboratory. Some of the experiments are being done in relation to some of the questions that Dr. Kloas addressed in that we've been looking at more of the responses of Xenopus to atrazine in terms of effects on steroidogenesis. And so we're looking and doing some mechanistic studies, trying to tease apart places in the pathway that are responsive and develop the methodology in order to do in vivo exposures and to take gonadal tissue outside of the animal and do 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 224 ex vivo incubations to test for specific lesions in the steroidogenic pathway. So those studies are just ongoing.

DR. ROBERTS: Any questions for Dr. Van Der Kraak. Dr.

Kendall, I think there's another phase of your public comments.

DR. KENDALL: Yes. Thank you, Mr. Chairman. We were encouraged to as a panel, and we did, respond to the White Paper in terms of the questions being posed to the SAP and would offer, Mr.

Chairman, that we discuss that or summarize it or provide this for the record. But each member of the SAP has been given our panel's response and literature backup as to some of the scientific issues we see. I would yield to how you recommend that we proceed.

DR. ROBERTS: I think that if you could summarize it for us,

your opinions and responses on these, that would probably be the most useful. Let me be sure that I've got it.

DR. KENDALL: This document is entitled, " The atrazine Endocrine Ecological Risk Assessment Panel's Response to The United States Environmental Protection Agency's Questions in the ` White Paper on Potential Developmental Effects of atrazine on amphibians.'"

DR. ROBERTS: All right.

DR. KENDALL: The date is June 17, 2003. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 225 ATTY2: Got it. Thanks. If you can maybe just proceed and summarize it for each question, that would be very useful.

DR. KENDALL: The members of the SAP, of course, will receive your charge and the questions probably Thursday morning.

Mr. Chairman, I really don't want to read each question, not unless you want me to. But we thought we'd at least project them on the board. And I wanted to just very quickly touch on some of the responses that we had as a panel.

Initially, we as a panel would like to compliment the Agency on their effort in bringing this White Paper together. Again, this is an emerging and one in which there's not a lot of standardization both in the science to do the research as well as the terminology. And would encourage the SAP to engage, not just the science, but the terminology so we call all begin to speak the same language.

Never the less, in terms of Question 1, in particular 1. a., " Does the SAP have any comments and recommendations..." In general, our panel supports the Agency's evaluation of the existing body of data and generally agrees with the conclusions; although there were two point from our perspective that needed to be considered.

We as a panel felt that the Agency focused more on the limitations of the data versus what the data could offer if it were 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 226 looked at in a robust, comprehensive manner. For instance, the limitations of low concentrations of atrazine in the reference site,

relatively great inherent variability in hormone concentrations, and other issues such as the time to metamorphosis.

Again, we accept the criticisms. We welcome the opportunity to address these criticism in the future. But let us not lose sight of the forrest for looking too hard at the individual trees.

The second point that we think merits attention is the Agency's statistical analysis of the data. Whenever the Agency reanalyzed our data, I want to emphasize for any Panel member, we provided all raw data for all studies. So from that perspective, one could take this and analyze this as one deemed appropriate. Never the less, the Agency's analysis of our data, if they found a statistically significant difference that our panel did not find, it was not always because EPA's analysis in terms of how they approached it in many cases EPA's analysis pooled animals together over sites, tanks, replicates rather than approach it as we did in preserving the structure in the data and including the animal site, tank, and replicate in the analysis.

And Dr. Silken was adamant as to how we approached these analyses in the context of statistical background. And we would off that it just be considered as one looks at all of these data together. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 227 In terms the b. part of Question No. 1, from a statistical perspective, the Agency's overall characterizations of the currently available studies tend to treat differences that are found to be statistically significant by one method analysis as true differences that are biologically significant and beyond question but tends to treat results, in terms of no statistically differences, as questionable.

So as we approached it, and I heard the comments today as my colleagues did. We approached it with the best science and the best statistics that we could apply. We are not seeing robust, repeatable statistical differences. Never the less, we would welcome the SAP to look carefully at this as well as assist the Agency in looking at their statistical approaches.

In 1. c., a number of studies have been done that address the effects of atrazine on development of anurans and also on the occurrence of testicular oocytes have been published in the literature.

Others that should have been included include Allran and Karasov 2000 and 2001, and Brown Sullivan and Spence of 2003. And we go into more detail on that particular area.

In Question No. 2, we agree that field studies have limitations as discussed by EPA. While these limitations are acknowledged, field studies we believe are extremely useful in a weight of evidence 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 228 approach and the results are still relevant.

This is particularly important for emerging areas of science where we need to make decisions. Data are coming in. And we believe that a comprehensive field and laboratory integration are critical.

The studies conducted in the field we believe were designed to test the presumption of risk. That is, they were designed such that, if there were no differences locations or correlations between exposure to atrazine and responses, it could be concluded that atrazine did not cause these effects under relevant environmental exposures. And we would welcome further deliberation by the SAP on this particular approach.

Never the less, Dr. Du Preez comments and I think his elegant summary of some of the works going on in South Africa, we challenge the concept that if atrazine had caused robust, sustained,

comprehensive population level effects on native Xenopus laevis in South Africa as one may have suspected from preliminary studies,

then it would have been reflected in disturbances of population structure, particularly after 40 years of application in that area.

Moving on to Question 3, in terms of 3. a., we believe that laboratory studies provide a plausible basis to establish a hypothesis 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 229 concerning the potential for atrazine to cause developmental effects provided that sample size is adequate the small incidence of abnormalities that have been reported in some studies, and these have been relatively small in some studies, and provided that the exposures overlap with critical windows of gonadal differentiation, and provided that sampling design takes into account the variability of timing in gonadal differentiation in different anuran species.

Many of these issues were raised today. I think this SAP panel is highly alert to these concerns and I think will address this nicely and assist the Agency with their recommendations. We believe that the overall body of data clearly indicates that the response of Xenopus laevis to atrazine varies under the conditions described in the available studies. And there are a lot of reasons for this. And, of course, this SAP will address many of those reasons.

Although the degree to which differences in experimental design and husbandry influence the contradictory findings remains a matter of debate, the fact that a relationship between atrazine exposure and development of gonadal abnormalities is not consistently found raises the question of the ecological significant and the relevance of observed effects of atrazine on gonadal differentiation. We think that's at the essence of the questions that 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 230 you will be addressing, Mr. Chairman.

In Subsection b. of that question, to date, the laboratory and microcosm studies have followed logical assumptions inherent in the scientific method. And we believe that given the inconsistent responses observed, it is not possible at this time to predict how Xenopus laevis will respond to atrazine under yet a different set of environmental conditions.

Although it is relatively clear that atrazine does not dramatically affect thyroid function and does not influence estrogen receptor activity at environmentally relevant concentrations, the ability of atrazine to influence gonadal differentiation is an as of yet unelucidated pathway cannot be predicted for available studies. And we are still searching, as Dr. Giesy and other members of our panel related to you, for these underlying mechanisms of action for potential effects.

Give the lack of a repeatable effect and the absolute lack of evidence for a cellular mechanism underlying the reported effects of atrazine on gonadal differentiation, it is not possible at this time to predict the dose response relationship or the rank order potency of atrazine metabolites relative to the parent compound.

In Question No. 4, we concur with the conclusions reached by 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 231 the EPA that there is no evidence to conclude that atrazine causes induction of aromatase in frogs, at least at this time.

In Subsection b. of that question, there is not evidence to suggest that analytical issues are a major factor contributing to variability in plasma sex steroid hormone levels or aromatase activity measures in frogs. I might add that, again, I emphasize, our projects were implemented on GLP like performance standards under SOPs where standards of performance were measured. The Eco Risk QA unit went to every laboratory, check and recheck process. So we did take into account analytical and measurement performance.

At least for the research associated with the Eco Risk panel here, our quality measures are in place. And we believe they are within acceptable bounds. And we do believe that perhaps variations in analytical results, particularly in the area of sex steroids, may have some biological underpinning moreso than analytical underpinning.

In Subsection c., we believe that in anurans, sex differentiation is sensitive to sex steroids during critical periods of development.

And I think this SAP will help elucidate for the Agency how one might approach a better measure of that for repeatable type data acquisition. There is little evidence to suggest that anurans are particularly sensitive to estrogens in terms of the induction of 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 232 testicular oocytes following sex differentiation. And our panel has discussed this issue. So this area, we would welcome the SAP to fully explore and offer some perspectives to the Agency on the relevance in either the reproductive or ecological relevance of these testicular oocytes.

Compounds acting as estrogens or anti androgens would be predicted to cause feminization of males. There are no data available to suggest that atrazine functions as an estrogen receptor agonist through binding to the estrogen receptor or induction of estrogen dependent responses. Similarly, there is no evidence that atrazine binds to the androgen receptor and functions as an anti androgen.

There is some real experts on this SAP related to this particular subject area. This is what we believe to date as the state of the science. We welcome your critical review.

Moving to the next question, just a few more minutes, Mr.

Chairman. I'm trying to roll through this.

Question 5, Subpart a., our understanding of spermatogenesis in anurans lags far behind that for mammals, especially for amphibians.

It is not known if accelerated growth precedes histological differentiation of gonads which would influence the developmental 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 233 rate at early stages as well as gonadal morphology. It appears that sex determination exhibits considerable plasticity and the ability of gonadal differentiation to respond to environmental factors may be highly adaptive. Therefore, it may be resulting in some of the differences in data that we are seeing currently.

Gonads in Rana curtipes initially differentiate into ovaries, and later, in the prospective males, the ovaries degenerate and transform into testes. This represents a semi differentiating type of gonad.

Thus, interpretation of background rates of ovotestes and/ or testicular oocytes occurrences in amphibian species, as well as interpretation in the context of environmental exposure and risk assessment, requires significant experimental evaluation under controlled laboratory conditions in addition to the evaluation of populations in the natural habitats.

And so we believe this SAP can contribute much thought and idea in the proposed future laboratory work necessary with atrazine and frogs. But let us not discount the importance, relevance, and contribution our field work can contribute to this whole subject area.

In subpart b. of this question, there is no information currently available that explicitly tests whether the presence of a few testicular oocytes would result in any impairment of reproduction. Therefore, 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 234 one would wonder is this an aberration or not.

The presence of robust populations of frogs, and this is debatable, Dr. Skelly, and we respect your opinion and welcome your seat on the SAP in contributing to the population understanding of this subject area. But there were presence of robust populations of frogs where there is a relatively great incidence of testicular oocytes further argues against such an adverse effect of this condition. This is point we want to make.

Recently, the research accomplished by our research team in male amphibians does not suggest that the presence of testicular oocytes and ovotestes results in the reproductive impairment via reduced fertility.

In Subsection c. of this question, the major point here  we addressed other points. But the laryngeal development in frogs, as Dr. Kelley has mentioned, is a sexually dimorphic process; and the formation of a larynx capable of a male calling behavior is androgen dependent. Under normal conditions the laryngeal dilator muscle of the male Xenopus laevis is larger than that of females. It's been hypothesized that atrazine could decrease plasma concentrations of testosterone in Xenopus laevis by up regulating the expression of aromatase, the enzyme that converts testosterone to estradiol. Yet 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 235 this remains to be proven.

And, again, this whole area, I think, would welcome further input from the SAP. It is probably inappropriate, we believe at least at this point with the laryngeal dilator muscle, to be using this effect as an assessment endpoint, perhaps a measurement endpoint. But this point may be moot, since at least at this point in time, we do not seem to have reproducible, sensitive measures of potential estrogenic or anti androgenic effects of atrazine.

In Question No. 6, our Eco Risk panel supports EPA's conclusion that the data currently available from both laboratory and field studies involving a wide range of amphibian species does not support the hypothesis that atrazine causes development effects in amphibians. That is yet to be proven. Given the low background incidence of gonadal effects reported in most studies, particular attention should be devoted to sufficient sample size so that the statistical power of the study is sufficient to properly test the null hypothesis. This has already been described by Dr. Gibbs.

The use of concentrations greater than those expected in the environment would allow evaluation of a threshold for gonad specific effects to be determined. Therefore, levels of exposure greater than 25 ppb would be necessary. The study should also consider the 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 236 possibility that effects could be caused by metabolites and/ or environmental break down products of atrazine and related triazines.

Hazard and risk assessments for pesticides and other substances have always been based on the principle of dose response and/ or concentration response. We believe there is neither a framework nor a general precedent for risk assessment based on U shaped, dose, or concentration responses. We will welcome SAP deliberation on this subject.

In Question 8, while some aspects of the study plan proposed by the U. S. EPA are reasonable; again, we compliment highly EPA's White Paper and their effort to engage the best science possible on this whole area of atrazine exposure in amphibians. It may be unwise to base the decision tree solely on histological effects in the gonad when there is currently no reason to believe that this the most sensitive endocrine response. And there has been no plausible mechanism of action that has been suggested by studies done to date.

Dr. Kloas's eloquently argued on other enzyme approaches and other endpoints that may be more elegantly sensitive to such effects is being considered by the SAP.

In the b. part of that question, it appears that the major set of endpoints is covered under the present approach that EPA has offered 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 237 except some of the ones I just mentioned. Work so far has revealed that the degree of gonadal differentiation at completion of metamorphosis is highly variable within species and especially between species. In fact, there is no a priori reason to suspect that gonadal differentiation would be timed with completion of metamorphosis. So it is unclear why the Agency is using this same sampling time, end of metamorphosis, to gauge the degree of sexual differentiation.

And I'm sure the SAP will deliberate and offer the approaches in particularly the lab component of these future proposed studies that will address these concerns that we have.

In Subsection c., it is extremely important to properly design for possible tank and site effects. It is also important to include possible tank or site effects in the analyses of the study data.

Not only is it important to design studies with multiple tanks or sites per treatment level, but it also important to include multiple tanks or sites at the control level. Even two or three tanks or sites at the control level may be insufficient to capture the true tank to tank variability. And this has already been discussed by Dr. Silken previously at the table today.

Based on initial power analysis performed by our Eco Risk 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 238 panel prior to conducting its laboratory studies, a minimum of eight tanks per treatment are necessary to account for inter tank variation.

In terms of the last few comments in Subsection e., within ranid species there are differences in the expression of androgen hormone dependent secondary sexual characteristics. This will have to be take into account. There are also differences at the hormonal and potential differences at the developmental level that are not clearly understood in ranids which would point to caution in assuming that the developmental processes and the mechanism of gonadal differentiation can be appropriately tested in Xenopus laevis alone which argues that Xenopus laevis may in fact be an initial good model species to test. But we should not count other native frog species that may be differing in their sensitivity, gonadal differentiation, et cetera.

Many ranids, for example Rana catesbeiana, are difficult if not impossible to breed under laboratory conditions, meaning that eggs would have to be collected from natural ponds with unknown exposure histories. Rana pipiens can easily be breed in the lab. So we have differences of how these various species will reproduce in the lab and/ or have to be collected from the environment.

In Subsection g. of the last question, we believe and we hope 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 239 that you will talk about this. But a toxicokinetic study of the uptake,

organ distribution, and depuration of atrazine in Xenopus laevis, rana species, as well as other frogs will be useful in determining whether extrapolation is possible between frogs. In other words, we offer caution that to take a non native laboratory model and try to extrapolate this to robustly to our native frog populations.

Thank you, Mr. Chairman. That concludes all we have to say in a summary nature of our document.

DR. ROBERTS: Thank you, Dr. Kendall. Reminding the panel that our opportunity to respond to these questions will come later, are there any questions to Dr. Kendall or other members of this panel regarding their rationale or basis for their responses to these questions. Dr. Kelley.

DR. KELLEY: I wanted to make sure I understand your summary. As I understand it, you felt that there was no robust and reproducible effect of atrazine in terms of development of intersex.

And yet in the Carr, et al., study, you say, and I quote, " Exposure to either estradiol or 25 micrograms atrazine per liter increased the incidence of intersex animal based on an assessment of gonadal morphology."

So do you mean that that was not a robust effect? I mean, there 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 240 were 11 replicates of that particular treatment group.

DR. CARR: Again, we found 4 percent intersex in that particular treatment. In terms of the percentage of animals that were affected, I wouldn't consider that a necessarily robust response.

Neither was the qualitative assessment of the intersex. Again, we had initially used the term hermaphrodites to refer to these gonadal abnormalities. But they weren't true hermaphrodites in the sense that there were testes and eggs sticking out. In that sense, they were fairly subtle.

And, again, when we looked at the histological level, they were identifiable as male or female at the histo level. But there were differences in the shape and structure of the gonad that popped out at the gross level.

DR. KELLEY: But it was statistically significant.

DR. CARR: Absolutely.

DR. KELLEY: It was reliable within your groups.

DR. CARR: Right.

DR. KELLEY: So you would conclude that this particular dose of atrazine did have an effect even if you felt that it was a high dose and it was a small effect.

DR. CARR: Absolutely. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 241 DR. KELLEY: But still you would stand behind the effect.

DR. CARR: Yes.

DR. KELLEY: Okay. Thank you.

DR. KENDALL: That's a good question. And, again, in the presentation what we emphasized, we went through enormous planning on the statistical power of that experiment. And had not that statistical power been so strong, we probably would not have detected that effect. But we stand by it, although it was small and we do not believe robust.

DR. VAN DER KRAAK: I think there's an additional point to add to that. There's another component to your question, though.

And the question was: Whether the response was reproducible across multiple studies. And in fact

DR. KELLEY: Oh, yeah, within your study. That's all I asked.

You did 11 replicates. I wanted to make sure it was reproducible within your study.

DR. VAN DER KRAAK: Correct.

DR. KELLEY: Right. Good.

DR. VAN DER KRAAK: The question, the additional point was whether it was reproducible across studies.

DR. KELLEY: 25 micrograms at that dose. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 242 DR. CARR: Right. And

DR. KELLEY: I really want to know about your study, whether you stood behind that conclusion.

DR. CARR: Right. I do. And the thing that might add some clarity is that those abnormalities were distributed across the replicates. They weren't from one tank. That's the point I wanted to make.

DR. KELLEY: Great. Thank you.

DR. KENDALL: That's a fair question.

DR. ROBERTS: Any other questions? Dr. Green.

DR. GREEN: One last question. I was wondering how the panel felt about the possibility that in order to get a handle on the significance or the number of frogs in the wild population in different species that live with ovotestes. How does the panel feel about the possibility that field studies would involve going out and capturing healthy frogs from healthy frog populations and literally having to kill hundreds of them, maybe thousands depending what the statistician tells us, just to verify that it is or isn't a problem in the absence or presence of atrazine?

DR. KENDALL: We may ought to wait to let the SAP answer that question. I think that's a great question. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 243 DR. KELLEY: They're not our frogs. They're SA's frogs.

DR. GREEN: I assume there would be some in the United States. You can see I find it's a bit troubling. Do you think the data right now supports, for the edification of the panel, that we consider having to go out and capture healthy frogs to determine how significant this is in a wild population?

DR. KENDALL: I think initially what we believe is going to take a lot of frogs to have the power to detect the effects because we don't believe the effects are that robust. That's a very good point.

Never the less, if one is going to get the correct kinds of data for future purposes to regulate this chemical relative to frogs, we will need to take the lab work to the field to see if, in fact, these effects are occurring and that they have any reproductive and/ or ecologic consequence.

DR. GIESY: That's an excellent question and one, certainly,

within my group we consider all the time. We have ongoing field studies that we're doing where we are collecting a lot of frogs. And,

of course, we do that under our animal use permits. But it's something we don't do lightly. And I don't think we should do lightly.

And I would encourage the Panel to consider that in whatever design you come up with for experiments. We would not want to do that 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 244 wantonly.

DR. DU PREEZ: One can also look at museum specimens.

There are literally thousands of Xenopus sitting in bottles throughout the worlds. But it's difficult to persuade the curators of those collections that you want to open up some specimens. They are sometimes very difficult. But that is a possible option. And now we know why frogs are declining.

DR. ROBERTS: Other questions from the Panel? Sorry. Dr.

Skelly.

DR. SKELLY: Perhaps I should have asked this before. It dawned on me, thinking about this, that one of the questions I wanted to ask your group was whether you have looked or whether you plan to look for any evolved tolerance to atrazine.

DR. SOLOMON: Perhaps I can answer on behalf of the group.

We certainly thought about this. Two aspects. One is has there been a genetic bottleneck that these frogs have gone through. And, in fact,

in frozen at minus 80 degrees celsius we have blood waiting for DNA analysis if anybody's interested in it. We'd welcome contributions from the Panel and the audience as well. Not contributions financially, but suggestions as to how we do that in case you get me wrong. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 245 The other issue is  I mean there have certainly been some suggestions of resistance. And I think the White Paper mentioned and we know that there's resistance in plants. Plants have a mechanism of action, receptor mechanism, that is susceptible to selection, change in the protein sequence. Of course, plants are under strong selective pressure, so they have evolved being selected for this resistance.

For this to have happened in frogs, I suspect we'd have to have a fairly strong response that either causes death or change in reproduction. And for my part, I haven't seen that yet. In my own experience with resistance, I think you need one of those or both of them to actually have selection take place.

DR. SKELLY: Just as a follow up. I guess that's my point is that there are two ways to document a demographic effect. One of them is to watch a population for a really long time. The other ways is, if you find an evolved response, that implies a past demographic effect.

DR. SOLOMON: Yes.

DR. KENDALL: That's a good point.

DR. ROBERTS: Any other questions? Or more response from Dr. Giesy.

DR. GIESY: The issue of resistance, I think, is a very 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 246 interesting one. And one that I'd like to hear some responses from the Panel. I personally have some ideas from a molecular biological techniques how you could approach that. I think it is a question you can pose. I think we do have tools that we can look at that. And I think it's probably worth looking at.

DR. ROBERTS: Any other respondents? Dr. Kendall.

DR. KENDALL: I just want to say, Mr. Chairman and distinguished members of the SAP, thank you for the opportunity.

We're very grateful to have had this time frame to discuss these issues with you.

I also appreciate EPA for setting up the opportunity to have such a scientific discussion and to give us the opportunity to look towards the future. We also appreciate Eco Risk for facilitating our efforts between our universities. And we appreciate our sponsor for giving us the support necessary to engage what we believe will be some important discussion in the next few days. And we welcome hearing from you. And thank you for your criticism, you input, and particularly your patience this afternoon.

DR. ROBERTS: On behalf of the Panel, I'd like to express our appreciation for you and your colleagues willingness to come here and discuss your research in very open fashion with the Panel and 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 247 answer all their questions regarding it. I think that's been very helpful in us understanding these issues.

DR. KENDALL: Thank you.

DR. ROBERTS: All right. Well, that concludes this public comment. There are plenty more to come. But at this point, I want to assess, do a stamina check on the Panel. The next public commentor,

we have two public comments from the registrant. They will take a total of two hours, but they could be divided. Potentially, we could do one of them. Then next one will take about an hour.

Okay. I can see by the looks on your faces. That answers my question. I just thought I would bring it up, but I got a pretty clear non verbal response on where we stand today.

In view of that, let us go ahead and adjourn the meeting today.

We will reconvene at 8: 30 tomorrow morning. We will continue with public comments, the first of which will be two public commentors from the registrant, Sygenta. And then we will proceed with other public commentors beyond that.

Paul, do you have any announcements before we close for today?

MR. LEWIS: Nothing to add, Dr. Roberts. I'm looking forward to continuing our discussion tomorrow morning. Thank you. 248 1 DR. ROBERTS: Today's session is closed. We'll reconvene 2 tomorrow morning at 8: 30.

3 [ Meeting convened at 4: 35 p. m.]

4  oo0oo 1 2 3 4 5 6 7 8 9 10 249 CERTIFICATE OF STENOTYPE REPORTER I, Jane F. Hoffman, Stenotype Reporter, do hereby certify that the foregoing proceedings were reported by me in stenotypy,

transcribed under my direction and are a verbatim record of the proceedings had.

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JANE F. HOFFMAN 1 2 3 4 5 6 7 8  9 10  250 ** I N V O I C E**** **** I N V O I C E****

JANE F. HOFFMAN TODAY'S DATE: 6/ 26/ 03 DATE TAKEN: 6/ 17/ 03 CASE NAME: FIFRA SAP/ Potential Developmental Effects of Atrazine on Amphibians TOTAL: PAGES: 363 LOCATION OF DEPO: Crystal City, VA DELIVERY: 7 day SPECIAL INSTRUCTIONS: Tapes to be included