“Help, help, I’m being slowly poisoned!’’ is a distress cry we hear often from more and more distraught people. Triazine, DDT, mercury, lead, asbestos, stilbesterol, nitrosamines, vinyl chloride-strange sounding names infrequently used. Are they harmful? Should I use products containing these substances? Is my workplace safe? Will this practice hurt me later? Decision, decisions, and questionable answers come from the experts. Some of these experts have to be chemists-chemists who can develop the required analytical procedures that will demonstrate the presence of the allegedly toxic substances in a biological specimen. Unless this can be done, the allegations that a substance has been absorbed and is causing harm cannot be objectively substantiated and the case cannot be proved. This challenge to analytical chemists is substantial because all too often, the quantity of offending agent that may be involved is quite small, and its concentration in the biological specimen may be in the microgram or nanogram range. And there are separation problems. Because these problems are not easily solved, many a case is made on the basis of the observations of the reaction of experimental animals or man to the chemical in question. This has to suffice in some instances because of the analytical deficit, but it would be preferable to have the analytical data so that we would be more certain of the conclusions drawn from the animal experiments. The converse must also be considered. Analysts have developed many sensitive methods for the detection of chemical hazards. They have applied these techniques to many specimens and have determined that the particular agent, DDT or, more recently, mercury, is present in detectable quantities. Does this now mean that any time we can put a realistic number down as an analytical result, people are being poisoned? The small 212A
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amounts now detectable because of improved analytical methods are not necessarily indicators of dangerous exposure and potential hazard. Interpretation of these numbers in terms of human experience is essential, and this becomes increasingly difficult when long-term chronic exposures to small amounts of potential hazards are involved.
Vinyl Chloride A fitting summary of this problem is the clarifying statement of a recent letter to the Occupational Health and Safety Administration by the American Chemical Society on its position on the agency’s “no detectable level” workplace standard proposed for vinyl chloride. The statement reads: “In the absence of knowing the tolerable limit for workers exposed to vinyl chloride, the standard should be set as low as can be detected by a currently accepted reliable analytical
4 7 , NO. 2, FEBRUARY 1975
procedure: namely, at 1 p p m . At the same time, work should continue t o define a threshold value so that, when it is defined, the standard m a y be suitably revised. Since development of t h e means t o attain the recommended level m a y take time, we recommend a reasonable period for users of vinyl chloride t o come into compliance. ’’ All of these quoted sentences must be used to indicate the present status of the problem, and to cite any one out of context is to distort and confuse the intent of the statement and is grievously unfair. Medications Our environmental exposure to chemical hazards is not limited to the food we eat or the air and water that surround us at home, at play, or at work. Other chemicals are in common use in our everyday activities. Our desire for immediate relief of headache,
Irving Sunshine Cuyahoga County Coroner’sOffice Cleveland, Ohio 44106
nagging backache, and the common cold, let alone the more serious ailments to which we occasionally succumb, results in a plethora of medications which are used with minimal concern for their toxic potential. These medicaments are not toxic when taken as directed or prescribed; in fact, they may be beneficial under these circumstances and necessary to ensure our health. But when directions for their use are disregarded, when those for whom the medication was prescribed allow others to use it, and when polypharmacology ensues because a patient or a host of others unbeknown to the prescribing physician take the prescribed medication, then real harm can result. One may add to this group who misuse medications, those individuals who knowingly or unknowingly take too much of any medication and those who abuse drugs for whatever reason. Now you have a real problem in which analytical chemistry can play a lifesaving role. Although medications are most frequently involved in accidental or suicidal poisoning, many household products and other chemical agents found in the home are also involved.
Dosage Problems Recently, those chemists whose talents have been applied to providing analytical data on whethef or not a person absorbed any potentially harmful agents have had another challenge. When a physician examines a patient subsequent to a previously
prescribed direction for the use of a medication and finds that the anticipated response has not been achieved, he has many questions to resolve. Among these are: Was the medication taken as prescribed? Was the dosage sufficient to achieve the therapeutic concentration in the patient’s blood? Are there other substances present which may interfere with the efficacy of a given agent even if it is present in the usual therapeutic concentration?
Analysts’ Role With these increasing concerns about the safety and well-being of the entire community has arisen an increasing interest by analysts as to what their role can and should be and how they can help. Those who presently fill this role are the environmentalist, the industrial hygienist, and the forensic and clinical toxicologist. The following presentation proposes to familiarize the reader with the current status of the writer’s field of interest, its problems, some of the remedies for these, and what the future holds. Analytical toxicologists are still a rare breed, although their numbers are growing. This growth lags behind the increasing demand for service and research, Hence, this open invitation to colleagues to swell our ranks so that they too can enjoy the satisfaction of applying their scientific talents to helping fill the health needs of their community while they concomitantly satisfy their scientific curiosity by developing novel and ingenious methods for analysis of biological specimens.
h e king’s f a s f e n is aeab. N o t b e c a u s e h e inabuerzfenfly m i s j u b g e a one of h i s s a m p l e s ana so p o i 6 b y h i s enRon of j u a g m e n f , b u t rzafhen b e c a u s e h i s Linean bescenbanfs pnef e n m o n e elegant ana s o p h i s t i c a f e a m e a n s fo a c h i e v e the s a m e goat w i f h o u f j e o p a n a i z i n g f h e i n own well b e i n g .
Wizardry The king’s taster is dead. Not because he inadvertently misjudged one of his samples and so paid by his error of judgment, but rather because his linear descendants prefer more elegant and sophisticated means to achieve the same goal without jeopardizing their own well-being. They resort to techniques which are overtly similar to that which their forebearers, the wizards, practiced. Much of the activity in today’s toxicology laboratory strikes the onlookers as wizardrya pass of the hand, a puff of smoke, and a crystal ball in which one sees the answer. Surely this describes atomic absorption spectrophotometry with its graphite furnace and recorder printout. Gas chromatography might also fit this description, except you don’t see the puff of smoke-the detector does. The colored handkerchiefs that came from nowhere surely are no less attractive than a chromatogram of many drugs spread over a thin-layer plate after it has been exposed to spray reagents for functional group analysis, Examining the entrails of an animal as an augury of the future? Did you ever analyze stomach contents? Enough of this imagery. Where are we today? Remarkably farther ahead then just 10 years ago. The rate of development in analytical toxicology has been geometric and has been aided and abetted by real and significant contributions of many outside the field. Without this help, many of the answers commonly available today would not be provided. Examine the picture in detail and see for yourself. Inorganics-classically lead, mercurials, arsenicals, bromides, and cyanides-were the stock in trade. Wet chemistry techniques, tedious and costly, were involved. Today atomic absorption spectrophotometry, with the graphite furnace or flameless electrode, allows for precise and sensitive results. AA procedures are rapid, relatively simple, and require remarkably 213 A
uch of t h e a c t i v i t y in tobay’s toxicology l a b o n a t o n y stnikes t h e onlookens as wizanarzy-a p a s s of t h e h a n b . a puff Of smoke, ana a c n y s t a l b a l l in which one sees t h e answerz.
small samples. For instance, it is now feasible to screen all children in an area known to be particularly susceptible to lead poisoning effectively, economically, and with a minimum of trauma to the kiddies because a pin prick of blood suffices for the sample. This type of testing program can do much to ensure that the youngsters tested are not injured as a consequence of their potential chronic exposure to lead. In contrast, the furor about mercury in fish resulted from the data provided by sensitive flameless atomic absorption methodology. But this surely would not have been so intense a concern if the analytical results were better understood. Today the list of detectable substances easily processed by atomic absorption is significant and growing. Complementing these developments in atomic absorption are the advances in analytical polarography. Differential pulse voltammetry and anodic stripping voltammetry are providing answers in the parts-per-billion range for many substances and are rapidly being accepted and used. Ion selective electrodes also play a significant role in inorganic analysis. Many anions are easily detected by these electrodes, frequently with little or no sample preparation. Gas chromatography is being used, not only for the analysis of cations, but now for bromides and cyanides as well. Organic Analysis Interesting as these developments are, they contribute to solving only a small percentage of the problems, because much of the toxicologist’s work concerns itself with the detection of organic compounds. Herein lie the major challenges. They have been met with extraordinary successes. Many analyses considered beyond any hope of solution seven years ago are now commonplace. Morphine in blood, polycyclic tranquilizers, phenobarbital and diphenylhydantoin, and a thousand urine samples a day for multiple analysis for many common drugs are but a few of these achievements. These were attained by adapting many innovative techniques. One of these, immunoassay, has grown by 214A
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leaps and bounds. Many applications have been developed using radioimmunoassay, hemagglutination inhibition, or homogeneous enzyme multiplied immunoassays. Originally developed for morphine, these techniques are now available for barbiturates, amphetamines, methadone, cocaine, diphenylhydantoin, digitoxin, and digoxin. Others will be developed depending on the demand. Truly a whole new era of analytical methodology is beginning with these few immunoassays. T o stress this latest approach is not to minimize the continued and ongoing contributions of many other techniques. Gas chromatography surely is noteworthy. Merely with a flame ionization detector and several differently packed columns, the number of detectable toxic agents is significant. When this is added to the electron capture detector and the nitrogen detector, the sensitivity and specificity of gas chromatography are significantly increased. To top all this, there is a surge of investigators developing GC/ mass spectrometry techniques which further enhance both specificity and sensitivity. As if all this were not enough, automated head space analyses of small samples are also a reality, not only for ethanol, but also for many of the industrial volatile hazards. Without this approach, data on vinyl chloride wouid be very difficult to obtain. Where would many laboratories be without the relatively simple, inexpensive, thin-layer chromatographic techniques? For small labs and for large ones plagued with a large daily work load, there still is no substitute for thin-layer chromatography. In contrast, high-pressure liquid chromatography is just beginning to be used in a few developmental iaboratories. The initial results indicate that this technique has much to recommend it in terms of speed, sensitivity, and ability to resolve mixtures. General acceptance and routine use of this approach are probably several years away but undoubtedly will come. Except for atomic absorption there has been no mention yet of spectro47, NO. 2, FEBRUARY 1975
photometry. Ultraviolet and fluorescence procedures are in common use and, in many instances, are the techniques of choice for many laboratories. These procedures are capable of producing reliable and valuable data economically. Many a laboratory depends on ultraviolet spectrophotometry to determine if a sedative drug is a contributing factor to coma of undetermined origin. This relatively simpleto-perform procedure has been lifesaving many times over. Another heavy demand on the laboratory is for a drug whose presence in urine can be used as a gauge of heroin abuse. Morphine, a metabolite of heroin, can be detected economically, rapidly, and reliably by fluorometry. We could make many points for this test as the most desirable for a primary screening procedure for morphine or heroin. Speaking of fluorescence, many substances can be detected on thin-layer chromatograms as reliably and with similar sensitivity to other procedures if fluorescent indicators are used. Here, as in gas chromatography, derivative formation is a powerful tool to increase sensitivity and specificity. This technique is frequently used to develop fluorophores on the chromatograms by using selected spray reagents which produce fluorescent derivatives in situ. Since this paper is not intended to be a comprehensive review of analytical toxicology, but rather an introduction to this field for our scientific colleagues, there is little need to labor the points already made with more detail. I t should be apparent that the ongoing problems one faces in this field are those relative to separation of the suspected offending agent from the biological matrix in which it is found and then to its identification by reliable, rapid, and economic means, while a t the same time maintaining sensitivity, precision, and accuracy. True, this may be the goal of all analysts, but the point being made is that among other things, the toxicologist is first and foremost an analyst. Many of the achievements cited or inferred above were developed by analysts whose primary responsibilities were in areas other than toxicology. For whatever the reasons they were developed, toxicologists either applied these directly or modified them to suit a particular need. Previously, most of those working in toxicology came to it through forensic laboratories and governmental agencies. With the changing times and needs, it’s interesting to note that in a recent list of “situations wanted,” 40% of those listed indicated they had skills in toxicology. Whence came these? How qualified are they? What is the extent of the accumulated expe-
oes t h i s n o w mean f h a f any time we can p u t a n e a l i s f i c n u m b e n bown a s an ana1;yticaL rzesulf, people ape b e i n g p o i s o n e b ?
rience of these individuals? The number of publications related to toxicology is one measure which may reflect some of this. There is no question of their variety and scope, as well as the abundance, relevance, and competence. Like the trade winds, these developments soothed the perspiring brows of those in toxicology laboratories who have toiled long and assiduously to maintain the requested daily services. These contributions and many others are needed. But it’s true that it’s time to legitimatize this development. Look about and note that most of those now interested in entering toxicology laboratories are doing so on their own. They acquire the techniques and the toxicological background on a self-informed, self-taught basis. This can be and often is productive. But obviously it is time to develop programs for those in the field so that they have someone else with whom they can discuss their problems and the solutions and so that they can benefit from others’ experience. That’s education and a sign of intelligence. It’s past due. Not only are continuing education programs on toxicology needed for related scientists, but it is essential that properly trained individuals be encouraged to enter this challenging field. The present in-
sufficient supply of trained toxicologists requires augmentation, and as yet training programs are generally unavailable. One can be misled too easily by formidable lists of toxicology training programs. Upon inspection, it is readily apparent that analytical toxicology and service-oriented programs are not any significant percentage of the cited programs. The need for formal training in analytical toxicology exists and must be met. Basic analytical chemistry curricula should be expanded to provide options in toxicology. These would not be difficult to implement and, once included in a program, would serve hopefully to invite some of the more adventuresome to participate in more advanced training. Predoctoral and postdoctoral programs should be encouraged in existing centers of excellence and in new ones that should be developed. Then and only then will the community have the assurance that their needs in toxicology will be satisfied in a reliable fashion. Aided by associates in related fields and assured of a continuing supply of welltrained novitiates, the analytical toxicologist will be able to continue to help protect the health and welfare of the people in whose community he serves.
Irving Sunshine is chief toxicologist for Cuyahoga County Coroner’s Laboratory and professor in toxicology in the School of Medicine at Case Western Reserve University. Dr. Sunshine earned his BS (chemistry), MA, and PhD degrees from New York University in 1937,1941, and 1950, respectively. After teaching experience in analytical chemistry, he worked in analytical research and analytical control of USP and CP chemicals in New Jersey until he became a chemist-toxicologist for the City of Kingston Laboratory, N.J. He left this position to take on his duties at Cuyahoga County in 1951. Dr. Sunshine has over 75 publications in various medical, clinical, and analytical journals. He is a member of ACS and Sigma Xi and has played active roles in the American Association of Clinical Chemists, American Academy of Forensic Sciences, and other organizations involved with poisons, alcohols, and drugs. Currently, he is an instructor in toxicology workshops, which are held in major cities in the U.S., and is editor-inchief of “Biosciences,” CRC Press. In 1973 Dr. Sunshine received the Ames Award from the American Association of Clinical Chemists.
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