ESTROGENS and ESTROGENIC EXPOSURE

The following document was prepared for a nutrition course I was taking. My project was given orally so some of the text is in point form. The text was originally prepared in Tex and may contain some remnants of Tex formatting. Any factual errors contained in this document are ones that I have made interpreting the available literature. Scientists don't always agree and all divergent opinions may not have been available. Many ideas are not included here due to lack of space (I was limited to about 10 pages). This is currently a topic of considerable scientific interest. Hopefully, this document will stimulate some of you to investigate some areas more thoroughly. The document was prepared over several months in the spring of 1994. - Ron Lyons
Please email comments, corrections, or criticisms to Ron Lyons. I still find this topic very interesting.

1. INTRODUCTION

While the literature is extensive and our awareness of the problem is growing in leaps and bounds the total ramifications of estrogens in the environment are unknown. Perhaps the reason it hasn't had a higher profile in the news media is because we have been saturated with bad news about the environment. So why should we regard this problem any differently? Just as the first picture of the Earth from space heightened our awareness that the Earth was not an infinite source or sink, and economic factors have forced a recognition of the global nature of the economy, so the problem of environmental estrogens may force us to acknowledge the global nature of the environment and the impact every industry, every activity and every person has on it.

In 1939 Buxton and Engle in talking about the clinical use of estrogens warned that estrogens are `` not innocuous substances but act on various organs and functions of the body not intimately related to the gonads...... The observed effects in women and the abundance of data on progressive effects in animals should raise a reasonable doubt in the mind of the clinician that these substances may be given with impunity''(quote from Edelman (1986)). For example, Grossman (1984) pointed out that the sex hormones regulate and were regulated by the immune system and the degree of their interactions is far more complex than ever realized. The variety of estrogenic compounds and their widespread distribution in the environment prompted Field et al (1990) to point out that we now live in a ``sea of estrogens''.

Perhaps the first alarm based on real life experience was sounded around 1970 when reproductive problems in avain populations were related to DDT. While useage stopped shortly thereafter in North America, useage still contiues in other areas of the world. DDT was found to have estrogenic properties as well as some of its metabolites. Since its introduction in the early 1940's, many other chemicals have been introduced into the environment. While many problems in wildlife populations are related to loss/degradation of habitat, a number of reproductive and behavior problems in shellfish, fish, bird, turtle and mammal populations are gaining widespread attention. (Surprisingly, no mention was made about the worldwide decline in the frog and toad populations that made the news last year. Perhaps this was an oversight in the articles I read.) Increasingly, attention is being focussed on the prevalence of estrogenic compounds in the environment and hence the food chain. Recently (Sharpe & Skakkebaek, 1993), environmental estrogens were suggested as a possible cause of falling sperm counts and rising levels of male reproductive tract problems being reported worldwide. Estrogens in the environment have been the subject of several conferences, the most recent in January of this year (Raloff 1994).

The problems related to the study of estrogen in the environment and its impact on the life forms in that environment are difficult and require input from many areas and in some cases many years of detailed work. Advances in science and technology may force the entire effort to be set aside if the avenues of exploration or data-taking have not been broad enough. A coordinated cross-disciplinary approach will be necessary.

2. DEFINITIONS

estrogen - generic term for estrus producing steroid compounds
-- -- female sex hormones.
-- -- formed in the ovary, possibly the adrenal cortex, the testes and the fetoplacental unit.
-- -- various functions in both sexes.

estrogenic - producing estrus; having properties of or similar to an estrogen
- Hertz's definition (quoted by Soto et al 1991)
-- -- primary effect of an estrogen is stimulation of mitotic activity in female gential tract
-- -- a substance which can directly elicit this response is an estrogen
-- -- a substance which cannot elicit this response is not an estrogen
-- -- (question: are there any substances that do in animals but not humans?)

3. ENDOGENOUS ESTROGENS

estradiol - most potent naturally occuring ovarian and placental estrogen in humans. The most active of the two isomers is estradiol-17&beta.

estradiol ethinyl - orally effective semi-synthetic derivatie of estradiol.

estradiol fatty acid esters - fully potent metabolite of estradiol (Hochberg et al 1990)

estriol - reduction product of estradiol and estrone

estrone - oxidation product of estradiol

4. ESTABLISHING ESTROGENICITY

Normal operation of estrogen in the body.

All classes of biologically active steroids are lipids. Their major metabolites, the conjugates are highly polar and usually charged. This renders the steroid water soluble so that it can be excreted easily. The fatty acid esters are somewhat different in that they are lipophilic. There is an indication that the estradiol metabolism pathway shifts in aging women from estrone which is less active and can be cleared easily to LE2 the lipoidal derivative, an active fully potent metabolite.

In general, the recognition of an estrogenic substance is accidental -- most are discovered after they are in the environment. Some test procedures used to establish or characterize estrogenic substances are:

  • induce estrus in ovariectomized female rats
  • measure growth rates in culture of estrogen sensitive cancer cells
    -- -- test for cell proliferation in human breast tumor MCF7 line
    -- -- (so far these satisfy Hertz's definition (Soto et al 1991)
  • search for active compound
    -- -- contaminant - use different preparations
    -- -- life force metabolite - generally won't work in culture
    -- -- in vivo activity may differ from in vitro due to body's processes
  • investigate environmental impact with model ecosystem (Metcalf 1980)
    -- -- use radiolabelled isotopes to follow metabolic pathways
    -- -- indications of bioaccumulation, biodegradation and degradative pathways
    -- -- can also test combinations for synergistic/antagonistic effects

    Several authors indicated the need for new procedures so that estrogenic compounds could be identified. McLachlan (1993) called for a ``functional toxicology'' so that chemicals could be defined more on their biological activity in a number of functional tests rather than their chemical properties. This seems reasonable as the body's steroid receptors seem to response favorably to a wide range of chemical structures. Because of this, it is difficult to use only a chemical's structure to determine its ability to mimic estrogen. In addition, numerous investigators in this field have questioned the validity of extrapolating the responses of animals exposed to estrogenic substances to human beings. Shellenberger & Sheehan (1978) indicated that, in oral contraceptive tests in rats of progesterone combined with estrogen, the progesterone level was too low to be physiologically active and the estrogen level high enough to be hyperactive. As a result only the estrogen was tested, not the combination.

    Appearance/behaviour aspects that can vary between estrogenic substances:

    Points to consider before ruling on estrogenic classification:

  • a) may be toxic in large doses, estrogenic in sub-toxic amounts
  • b) toxic effects may modify the estrogenic ones or the estrogenic response
  • c) substance being tested may have contaminants which are antagonists
  • d) substance may have contaminants which dilute effect of pure substance
    -- -- e.g. one of contaminants in DDT is estrogenic
  • e) may not break down, thereby escaping normal control mechanisms in vivo
  • f) may not bind, therby bypassing normal regulation methods in vivo
  • g) may bind to receptors other than those used by estrogen in vivo
  • h) some estrogenic effects may due to periperal effects in vivo
    -- -- e.g certain PCB's alter rat liver metabolism of estrogens
    -- -- e.g. changes concentrations of one or more naturally occuring steroids
  • i) action may be affected by other hormones in vivo
  • j) indicators should be sampled frequently enough and long enough to define impact
    -- -- e.g. Nafoxidine lower than estradiol initially but extended action
  • k) the effects observed may be species dependent

    The questions people are asking and the methods people are using to conduct their investigations have changed considerably with our knowledge and technology.

    5. DIETHYLSTILBESTROL

    - best documented story of progression of knowledge
    - leaves a lot to be desired, probably impossible to complete
    - good example of how not to do research and clinical studies
    - following largely based on Edelman (1986)

    history of DES development
    - story began in 1930's as search for synthetic (cheap) yet potent estrogen
    - 1933 E.C. Dodds reported first artificially synthesized estrogen
    -- -- stilbenes - a hydrocarbon series from ethylene
    -- -- diethystilbestrol (DES) was the most potent of the stilbestrols
    -- -- lacked basic ring structure of steroids so non steroidal
    -- -- compared well with estrogen in biologic properties
    -- -- orally active, inexpensive unlike natural estrogens
    - 1941 FDA approves for various gynecological conditions (not pregnancy)
    - early 1940's George and Olive Smith
    -- -- successful use in the prevention of spontaneous abortion, premature labor
    -- -- 1946 useage regimen throughout pregnancy (32nd week)
    - 1947 FDA approval for high risk patients in pregnancy
    -- -- approved under Food Drug and Cosmetics Act of 1938
    -- -- only need to show that it was safe when used in pregnancy (not effective)
    - 1950's clinical studies questioning effectiveness in pregnancy
    - 1956 through 1971 - place in practice, not a research issue
    - 1960's thalidomide catastrophy
    -- -- concerns about neonatal exposures and possible fetal abnormalities
    - 1970's safety and use questionned extensively
    - 1971 - report linking DES in utero to rare form of vaginal cancer
    -- -- awareness that there might be long term impact
    - 1971 - FDA withdrew approval for use in pregnant women
    - 1974 - National Cancer Institute study
    -- -- incidence of genital tract abnormalities and cancer among DES exposed
    - 1978 - Surgeon General's warning of hazards in pregnancy
    - 1970's and 80's - attention to adverse affects in utero
    - 1980's correct rationale for use of estrogens to support pregnancy (maybe)
    - 1985 still approved for a number of problems including
    -- -- some estrogen replacement therapy
    -- -- advanced breast cancer or prostate cancer
    -- -- used as a morning after pill
    - current status?

    other uses and relevant work
    - 1928 - prenatal exposure to estrogens feminized male rats
    - 1930's - estrogens can cause breast cancer in mice ==>> carcinogens
    - 1940 - prenatal exposure (estrogen, DES) congenital malformations in some animals
    - 1943 - used as a fattening agent in chickens (increased subcutaneous fat)
    - 1948 - approved for weight gain and feed utilization in cattle and sheep
    -- -- for cattle to increase weight (increased lean meat)
    -- -- approved on a ``no residue'' basis since carcinogen
    -- -- (Delaney Clause of Food Drug and Cosmetics Act of 1938)
    - 1959 - banned in chickens by FDA
    - 1962 - Food Drug and Cosmetics law amended to require safety and effectiveness
    -- -- does not appear to have been enforced with DES (no effectiveness studies)
    - 1979 - banned in cattle by FDA

    studies were done
    - most of the studies are flawed in significant ways
    -- -- several of the blind studies had lost their group designations (long term)
    -- -- critical information is missing
    -- -- reasons for exposure in first place
    -- -- if used perhaps because there were problems in fetus to begin with
    - DES was manufactured by about 270 companies under many different names
    - other hormonal products were available during the time period
    - exposure estimates from 1947-1971 range from 500,000 to 4 million
    -- -- 97 million births ==>> as high as 3% exposure rate
    - reconstruction of what went on 20 to 30 years or more before is difficult
    -- -- most of the studies are after the fact
    -- -- sales records that could give geographical information are not available
    - many of the conclusions are not firm but these strongest ones
    -- -- genital abnormalities consistent with dose response relationship
    -- -- higher rate unfavorable pregnancies perhaps tied in with abnormalities
    -- -- no beneficial effects reported
    -- -- more problems found in females (also studied more)
    -- -- reproductive/genital problems in animals (varies with species, strains)
    - too soon to look for second generational effects (if any)

    one would like to think that some good came out of this
    - awareness that long term consequences need to be evaluated
    -- -- perhaps some erring on the side of caution
    -- -- but similar studies have not been conducted for other estrogens in use
    -- -- eg clomiphene citrate (non-steroidal estrogen widely used, little studied)
    - modifications to FDA approval methods
    - evolution in testing / evaluation methods
    - 1971 - establishment of Registry on Hormonal Transplacental Carcinogenisis
    -- -- voluntary submission of cancer data
    -- -- major database but may be biased by voluntary aspect
    -- -- changing names/ different diagnoses with practionner
    -- -- some chance (albeit poor) of investigating long term impact
    -- -- baseline data
    - importance now
    -- -- study of hormonal system
    -- -- research on impact of estrogens on animals)
    - importance of ethical issues
    -- -- how to do studies on people

    6. SOME SOURCES OF EXPOSURE TO ENVIRONMENTAL ESTROGENS

    As indicated previously substances with estrogenic properties are widespread in the environment. Most of the chemical substances or byproducts produced in our daily lives have not been tested. Their operation as agonists or antagonists to the body's own steroid chemistry is unknown. Their operation in combination is also unknown. It is also apparent that most estrogenic substances are determined accidentally.

    A number of substances that have been examined have been called ``weakly estrogenic''. Clark & Peck (1979) point out that estriol, normally called a ``weak'' estrogen, is, under continuous or repeated exposure, fully potent. The weakness stems from its inability to remain combined with the nuclear receptor long enough to elicit a uterine growth response. It is quickly cleared. Repeating the exposure will elicit the growth response. They prefer the term ``short-acting'' to ``weak''. It remains to be seen whether all so-called ``weak estrogens'' are weak for the same reasons. This terminology does not appear to have been adopted and the use of the term ``weak'' may lead to false impressions.

    1) urine and feces
    Metabolites differ depending on the method of excretion. In solution, the higher the concentration, the more stable it is. In soil, the compounds adhere to the soil. The sensitivity of soil microbes is unclear but there is some evidence that plants can take them up and pass them on. Plant tyrosinase can deactivate estrone. Beet roots, potato juice and mushroom extract can inactivate and certain bacteria can degrade estrone. In arid areas, they do not degrade as fast. Natural estrogens appear quite stable in water. In model ecosystems, they concentrate in certain aquatic food chains (Knight 1980).

    Adult cows excrete 30 mg of estrogen/day, cycling heifers 2.2 mg/day. Hens have 1.6 mg of estradiol per gm of dry excrement. Estrone which is less potent than estradiol is found in human pregnancy urine, male human urine, mare pregnancy urine and stallion urine as well as palm kernal oil.

    2) phytoestrogens
    Over the millenia, many plants have developed chemical pathways that produce compounds not directly related to their growth or reproduction. While many of these secondary metabolites are toxic, some of them are estrogenic and have been used in native medicines for centuries.

    In 1926, the presence of estrogenic substances in plants was demonstrated by inducing estrus in animals. The first isolation of an estrogen from plants was reported in 1933. This initial work was not followed up until the 1950's when people began to recognize that animal infertility could result if too much estrogen rich plant material was consumed. Some people were looking at estrogen rich plant material as an inexpensive way to fatten livestock Farnsworth et al (1975).

    The phytoestrogens, coumestrol and genistein, interact with the estradiol receptor. Genistein has been the suggested cause of an outbreak of infertility in sheep grazing on clover in Australia (Rall & McLachlan 1980). Infertility has also been reported in cattle, mice and quail (Hughes 1988).

    Farnsworth et al (1975) listed a number of plants that they felt might have value as new antifertility agents. Hughes (1988) indicates that over 300 plants in 16 different families have been found to contain more than 20 estrogenic substances of 4 chemically distinct classes (steroid estrogens including estradiol and estrone, isoflavonoids, coumestans, and resorcylic acid lactones).

    3) Fusarium mold (Pathre et al 1980)
    Fusarium mold is the agent responsible for many of the wilts, blights and rots found in commercial agriculture. It can cause serious economic losses in major cereal crops like corn, wheat, barley, oats, sorghum, sesame, hay and animal rations. The estrogenically active compounds are the zearalenones, hormones that regulate the production of the sexual stage. Under the right conditions, concentrations can increase in storage. Corn is most often affected and if severe enough possibly present in corn oil too. Zearalenones can cause mycotoxicoses in animals and are sometimes found with other toxins, complicating the symptomology.

    The estrogenic effect varies greatly with species being particularly severe in swine - estrogenic effects in females, feminizing effect in males. It can stop ovulation and produce infertility. It affects the fertility of ganders and male turkeys while having no effect on laying hens and broilers. It is used to promote growth in cattle and sheep (Zeranol as an ear implantation (Knight 1980)). It is used to produce medication that alleviates post menopausal distress.

    4) medical uses
    -- oral contraceptive
    -- hormonal pregnancy tests (now banned in U.S. but used elsewhere)
    -- estrogen replacement therapies
    -- treatment of osteoporosis
    -- treatment of threatened abortions
    -- treatment of various functional ovarian disorders
    -- treatment of prostate cancer

    5) pesticides/insecticides
    -- DDT, one of the chlorinated hydrocarbons was found to be weakly estrogenic through the form o,p' DDT a 15-20% contaminant. It is concentrated by food chains. The metabolite DDE is weakly estrogenic. It is found in body fat and breast milk (Rall & McLachlan 1980). DDT analogues compete with estradiol for receptor sites (Eroschenko & Palmiter, 1980).

    -- Methoxychlor is a biodegradable substitute for DDT used as an insecticide for animal parasites home and garden with breakdown products that are similar to DES and estradiol. Around 1980, it was estimated that about 10 million pounds were used annually (Metcalf 1980) Sporadic occurrence of uterine enlargement in laboratory rats was traced to dusting with insecticidal powder whose active ingredient was technical grade Methoxychlor (Hertz 1980).

    -- The pesticide Kepone (Metcalf 1980) is used as a stomach poison bait for cockroaches and ants, as well as thrips on bananas. The active ingredient, chlordecone, exhibits weak estrogenic activity. Kepone was discharged into the air and water at its manufacturing plant in Virginia. In one case about 100,000 pounds spilled into James River which flows into Chesapeake Bay. Mirex, a pesticide used to control fire ants in the south-east U.S, breaks down to Kepone in the environment (Eroschenko & Palmiter 1980). In large dose Kepone is toxic, it is a neurotoxin and is carcinogenic. In 1975 846,000 lb was produced.

    -- Kepone (Eroschenko & Palmiter 1980) causes rapid developmental changes in immature quail similar to estradiol-17&beta. It causes reduced fertility, reduced egg integrity over long periods and inhibits ovulation. It causes testicular atrophy in males. There is a dose dependent stimulation of reproductive tract growth in mice. It is highly persistant and accumulates in aquatic food chains (Metcalf 1980).

    5) PCB's
    Found in sealants, additives to paints and plastics, PCB's are weakly estrogenic. They have been found in breast milk are are reported to reach the fetus (Rall & McLachlan 1980).

    6) Polycyclic aromatic hydrocarbons (PAH)
    PAHs arise from fossil fuels. A hydroxylated form is weakly estrogenic (Rall & M$c$Lachlan 1980).

    7) Alkylphenols (Soto et al 1991)
    Alkyphenol compounds are used as antioxidants in the plastics industry. They may leach from the PVC used in food processing and packaging as well as PVC pipe carrying water. They are used in industrial detergents and have been reported in sewage sludge. They have been found to bioaccumulate in fish. Nonylphenol was identified as an estrogenic substance after the manufacturer of polystyrene test tubes modified his formula to improve their integrity under centrifuge operations.

    8) Breast Milk and Cow's Milk

    9) Water Supply
    While it is clear that estrogenic compounds are dissolved in the water resources it is not clear to me how much is removed in water purefication. Certainly estrogen loads in un- and under- developed countries will be at least as large as the animal population.

    10) Air
    Many estrogens are volatile. The worldwide distribution of DDT by the atmosphere is well known. In addition, aerial sprayings and release of toxic fumes contribute to the air component.

    7. ENDOGENOUS ESTROGENS AND DIET

    Work discussed by Adlercreutz (1990) indicates that diet can have an impact on the level of plasma sex hormones. Diet has been suggested as the main single determinant in the origin of hormone-dependent cancers. High fibre diets increased elimination of estrogens apparently because of the ability of the fibre to bind the sex hormones. It is also associated with higher levels of the Sex Hormone Binding Globulin (SHBG). Fat decreased estrogen elimination. A high ratio of protein to carbohydrate in the diet decreased the plasma levels of SHBG. Proteins inhibit the formation of SHBG in the liver. The highest ratios were found in the breast cancer patients and the lowest in the vegetarians. (Breast cancer is an estrogen sensitive cancer.) Adlercreutz notes that the pattern of the hormones of the breast cancer group was associated with a Western type diet.

    Adlercruetz concludes that the sex hormone metabolism is influenced by all dietary components and that the wrong diet may influence hormone dependent cancers in the promotional stage.

    Estradiol is deactivated primarily along two metabolic pathways with very different biological activity. Michnovicz & Bradlow (1990) discuss the possibilities of using diet to change the preferential pathway.

    8. SOME FOOD FOR THOUGHT / SPECULATION

    1) The alarm over DDT occurred in the 70's. Pesticides, estrogenous growth hormones, and estrogen treatments had been around for at least 30 years so the estrogen load in the environment was increasing rapidly. Does the ``feminist revolution/movement'' have a component due to environmental contamination?

    2) Exogenous estrogens disrupt the hormone balance. Studies have should they lead to a feminizing effect in males. Is the violence in our society and the increasing tension world wide a response to higher estrogen loads.

    3) Is the anectdotal correlation between vegetarianism and reduced aggression due partly to the estrogenic properties of plants?

    4) In breeding plants more resistant to insects and molds, are we breeding plants that have a higher estrogen content? After all the plant needs to control or reduce grazing and population control is a long term desire. Insects have developed ways of using or inactivating plant toxins. Have people done this too? (Adlercreutz (1990) points out some of the benefits of phytoestrogen isoflavinoids.)

    5) Is plant estrogenicity changing? Could intensive agriculture change it? How would we know since our techniques have improved so much in since the first discovery? The time base is very short.

    6) People noticed the ``spring flush'' in cattle. In eating foods from all over the world as we do are we subjecting ourselves to higher ``spring flush'' loads? Is the idea of eating with the seasons a way to reduce (smooth out/purge) the estrogenic load? Some estrogenic compounds take a long time to clear out of the system. Could seasonal eating promote this?

    7) Are sprouted foods more estrogenic (``spring flushy'')?

    8) Will chemicals that affect our reproductive capacity be nature's ultimate way of controlling human population growth? It's subtle, it isn't necessarily fast but if it is just gradual who's going to be around to notice something is wrong and how long will it take to realize it. By them it may be too late to actually do anything.

    9a. References - Journal Articles

    Adlercreutz, H. 1990 ``Diet, Breast Cancer and Sex Hormone Metabolism'', Annals New York Academy of Sciences 595, pg. 281-290.

    Colborn, T., vom Saal, F.S., Soto, A.M. 1993 ``Developmental Effects of Endocrine-Disrupting Chemicals in Wildlife and Humans'', Environmental Health Perspectives, 101, pg 378-384.

    Davis, D.L, Bradlow, H.L., Wolff, M., Woodruff, T., Hoel, D.G., Anton-Culver, H. 1993 ``Medical Hypothesis: Xenoestrogens as Preventable Causes of Breast Cancer'', Environmental Health Perspectives, 101, pg 386-387.

    Eroschenko, V.P., Palmiter, R.D. 1980 ``Estrogenicity of Kepone in Birds and Mammals'', in Estrogens in the Environment, ed. J. McLachlan (Elsevier; North Holland), pg 305-325.

    Farnsworth, N.R., Bingel, A.S., Cordell, G.A., Crane, F.A., Fong, H.H.S., 1975 ``Potential Value of Plants as Sources of New Antifertility Agents II'', Journal of Pharmaceutical Sciences, 64, 717.

    Field, B., Selub, M., Hughes, C.L. 1990 ``Reproductive Effects of Environmental Agents'', Seminars in Reproductive Endocrinology, 8, pg. 44-53.

    Hertz, R. 1980 ``Personal Experiences Emphasizing the Environmental Impact of Estrogens'', in Estrogens in the Environment, ed. J. McLachlan (Elsevier; North Holland), pg 347-352.

    Hochberg, R.B., Pahuja, S.L, Larner, J.M. Zielinski, J.E. 1990 ``Estradiol-fatty acid esters: endogenous long-lived estrogens'', Annals New York Academy of Sciences, 595, pg 74-92.
    W1 NE788 v595

    Hughes, C.L. 1988 ``Phytochemical Mimicry of Reproductive Hormones and Modulation of Herbivore Fertility by Phytoestrogens'', Environmental Health Perspectives, 78, pg 171-175.

    Knight, W.M. 1980 ``Estrogens Administered to Food-Producing Animals: Environmental Considerations'', in Estrogens in the Environment, ed. J. McLachlan (Elsevier; North Holland), pg 391-401.

    McLachlan, J.A. 1993 ``Functional Toxicology: A New Approach to Detect Biologically Active Xenobiotics'', Environmental Health Perspectives, 101, pg 386-387.

    Metcalf, R.L. 1980, ``Model Ecosystems for Environmental Studies of Estrogens'', in Estrogens in the Environment, ed. J. McLachlan (Elsevier; North Holland), pg 203-211.

    Michnovicz, J.J., Bradlow, H.L. 1990 ``Dietary and Pharmacological Control of Estradiol Metabolism in Humans'', Annals New York Academy of Sciences, 595, pg 291-299.

    Pathre, S.V., Mirocha, C.J. 1980 ``Mycotoxins as Estrogens'', in Estrogens in the Environment, ed. J. McLachlan (Elsevier; North Holland), pg 265-278.

    Raloff, J. 1994 ``The Gender Benders'', Science News, 145 pg 24-27.

    Raloff, J. 1994 ``That Feminine Touch'', Science News, 145 pg 56-58.

    Rall, D.P., M$c$Lachlan, J.A. 1980 ``Potential for Exposure to Estrogens in the Environment'', in Estrogens in the Environment, ed. J. McLachlan (Elsevier; North Holland), pg 199-202.

    Sharpe, R.M., Skakkebaek, N.E. 1993 ``Are oestrogens involved in falling sperm counts and disorders of the male reproductive tract'', The Lancet, 341, pg 1392-1395.

    Shellenberger, T.E., Sheehan, D.M. 1978, ``Estrogens, Estrogen Receptors, and Biological Responses in Experimental Animals'', Frontiers of Hormone Research, 5, pg 203-219.
    ( Estrogen Therapy: The Benefits and Risks), ed. Lauritzen, C., van Keep, P. (Karger; Basel, Switz)

    Soto, A.M., Justicia, H., Wray, J.W., Sonnenschein, C. 1991 ``p-Nonyl-Phenol: An Estorgenic Xenobiotic Released from ``Modified'' Polystrene'', Environmental Health Perspectives, 92, pg 167-193.

    9b. REFERENCES - BOOKS

    (catalogue numbers refer to UCSD Biomedical library) ``Estrogens in the Environment'', ed. J.A. M$^c$Lachlan, Elsevier: North Holland, 1980. WP 522.S989 1979E BML

    ``Female Sex Steroids -- Receptors and Function'', Clark, J.H., Peck, E.J., Springer-Verlag: Berlin, 1979. WP 520.C593F 1979 BML

    ``Diethylstilbestrol -- New Perspectives'', Edelman, D.A., MTP Press Ltd:Great Britain, 1986. WP 522 E213D 1986 BML

    Desired but previously checked out by someone else:

    ``Estrogens in the Environment II: Influences on Development'', ed. J.A. McLachlan, Elsevier: New York, 1985.

    ``Chemically induced alterations in sexual and functional development: the wildlife human connection'', ed. T. Colborn, C. Clements, Princeton Scientific Publishing: Princeton, NJ., 1992.

    Ron Lyons (volunteer 1990-1999)
    Chula Vista Nature Center, 1000 Gunpowder Point Drive, Chula Vista, CA 91910-1201