1. DO NOT BE DECEIVED!
AllDS AND CANCER ARE CURABLE!
FRIENDSPAY ATTENTION TO THE FOLLOWING:
If you are deceived into believing that there is NO cure for Aids o r
Cancer and are suffering from or have loved ones who are suffering
from these dreaded dis-eases. the following may be o f extreme help in
reducing or even arresting the furtherance of yours or their suffering.
Check out the following extracts.
These are n o t Irepeat NOT from any unrecognized sources or journals
but from highly-respected individuals and institutions!
The Government, in association with the FDA. AMA and even the press.
is being NEGLIGENTt o the welfare of our fellow human beings.
DO NOT FALL PREY TO THEIR NEGLIGENCE
AND THE LACK OF RECOGNITIONOF THEIR OWN DATA!
Rise up from the doldrums o f apathy and unbelief!
Our ignorance and lack o f care t o the laws of the Mother and
our personal insensitivity(ies) has caused these problems.
Demand utilization NOW o f these scientifically, time-tested.
safe. non-toxic. harmless and life-saving compounds.
Demand this from your Government officials. health and welfare
institutions. doctors, colleges o f research and the press NOW!
CHECK OUT THE EXTRACTS BELOW!!!
The following dtzitlon Is from BLOOQ The Jourrul of the A m e r i a n Sodety o f Hematology, .
Vol. 78. No. 7, October 1, 1991:
lnoctivalon of Human lmmunodefulency Vlms Type Iby Ozone In Vhro
By Keith H. Wells, Joseph Latino, lerrie Gavalchin, and Bernard 1. Poiesz
A device was designed to deliver a constant source of given concentration of ozone to fluids containing human immuno-
deficiency wrus type 1 (HN-1). Ozone was found to Inacttvate HIV-1 vlrons in a dose-dependent manner. Greater than
11 log inactlvat~onwas achieved wthin 2 hours at a concentration of 1,200 ppm ozone. Similar concentrations of ozone had
min~maleffect on factor Vlll activ~tyin both plasma and immunoaffinity-purifiedpreparationsof factor Vlll treated for the
5ame time period. The data indicate that the antiviral effects of ozone include viral particledisruption. reverse transcriptase
~nac~vation,andor a perturbationof the ability of the vlrus to bind to 16receptor on target cells. Ozone treatment offers
p m b e a a means to lnacttvate human retroviruses In human body fluids and blood product pmparatlons.
Q 1901 by The American Sociefy of Hematology
llw~following dtetfon Is from the mpected d e n t l f i c journal
SCIENCE,Vol. 209. August 22, 1980--om deven yecm ago:
Ozone klecttvely lnhibtts Growth of Human Cancer Celk
Abrtract: The growth of human cancer cdls from lung, breast. and uterine tumon was sdectivdy Inhibitedin a dose-
dependent manner by ozone at 0.3 to 0.8 part per million of ozone in ambient air during 8 days of culture. Human lung
d~ploidfibro-blastssewed as non-cancerouscontrol cells. The presence of ozone at 0.3 to 0.5 part per millioc ~nhib~tedcancer
cell g r m h 40 and 60 percent. respectively. The non-canceroa lung cells were unaffectd at these levels. Exposure to ozone
at 0.8 pert per rnllllon inhlbhed cancer cell growth more than 90 percent and control cell growth less than 50 percent.
Evidendy, the mechanisms for defense against ozone damage are imparted in human cancer cells.
The tollowing bmklet showlng the cause of cancer as cellular anoxia (oxygen deprlvatlon) and toxlcRy
was wrltten by Dr. Otto Warburg. the ONLY man In the history of sclence and medldne to be given the
Nobel Prize in Medicine hvlce and nominated a thlrd tlme tor hle dlocoverlee In health:
The PrimeCause and Prevention of Cancer with two prefaces on prevention- Revised lecture at the meeting
of the NobelLaureates on June 20, 1966 at Lindau. Lake Constance, Germany by Otto Warburg. Director, Max
Planck Institutefor Cell Physiology. Berlin-Dahlmer; English Edition by Dean Burk, National Cancer Institute,
Bethesda, Maryland. USA 1967.
Ifyou desire more information, call or write:
P.0. BOX 1360 'PRIEST RIVER. IDAHO 83856 -

2. Warbure, Blass and Koch: Men With a Message
Dr G A Fre~bott
lnternatronal Assoc~atlonfor Oxygen Therap)
P 0 Box 1360
Pnest Rrver, ID 83856 USA
'Today's Scientists have substituted mathematics for experiments, and they wander off through equa-
tion after equation, and eventually builda structure which has no relation to reality." '
"The scientists from Franklin to Morse were clear thinkers and did not produce erroneous theories.
The scientists of today think deeply instead of clearly. One must be sane to think clearly, but one can
think deeply and be quite insane."
'Nobody today can say that one does not know what cancer and its prime cause is. On the contrary,
there is no disease whose prime cause is better known, so that today ignorance is no longer an excuse
that one cannot do more about prevention. That the prevention of cancer will come there is no doubt,
for man wishes to survive. But how long prevention will be avoided depends on how long the proph-
ets of agnosticism will succeed in inhibiting the application of scientific mowledge in the cancer field.
In the meantime, millionsof men must die of cancer unnecesarily." .
'A11 truth passes through three stages: First, it is ridiculed. Second, it is violently opposed. Third, it is
accepted as self-evident."
These quotes are the frank opinions of the Fsther of Alternating Cur-rent, a two-time Nobel Laureate, and a
nineteenth century German philosopher.
Warburg, Blass and Koch were men whose truths are beconling evident in today's world of science. These
were pioneersand scientists of the highest magnitude. This abstract uncovers, postmortem, the discoveries of
these men and their contributionsto future scientific studns.
Otto Warburg won his first Nobel Prize in 1931for the oxygen-transferringenzyme of cell respiratlonand hrs
second in 1944for his discoveryof the hydrogen transrerrir~genzyme.
Regarding his discoveries he IS quoted as follows:
"Even for cancer, there is only one primary cause. Summarized In a few words, the prime cause of cancer
is the replacementof the respirationof oxygen in normal body cells by a fermentation of sugar."
"Because no cancer cell exists the respiration of which is intact, it cannot be disputed that cancer could be
prevented if the respiratlonof the body cells would be kept intact."
Dr. F. M. Eugene Blass, an oxidation specialist and enginerldesigner of the PennsylvaniaSteelcokeovens,
clinically verified Warburg's foundational work. Returning to the United States in 1925, cured of his cancer
and armed with the knowledge of the Institut fur Sauerstoff-Heilven-ahren, Blass adamantly represented the
'The recognizable resultr of an insuffkient oxidation erther because of a lack of minerals or oxygen or because of the pres-
ence of foreign matter In the bloodstream are the symptoms wh~chhear the lmpostng nomenclature of modern 'dis-ease.'
The drfferent knds of parasrtes, whlch are the 'germs' commonly blamed for the creabon of these various symptoms, find
food anc! lodglng In the d~seasedsoil which accumulates rn the body, but logically they are not the cause of disease. A
clean body will not tolerate such habrtatron and normal vrtal flulds constrtute the best Insuranceagainst sickness." '
"OXIDATION is the source of Life, its lack causes impaired health or disease, its cessation death."

3. William F. KO&, M.D., Ph.D., a well-pl~b~shedpathologist and medical school professor, presented his co-
practitioners with 'a study of the phenomena of the free radical, the double bond, and its alpha-placed hydro-
gen atom in the pathogenesis and correction of neoplastic, viral and bacterial diseases." This professor and
teacher was truly the granddaddy of ALL oxidative therapies, now referred carte blanche to "oxygen thera-
pies". It is absolutely a fact that all REAL"oxygen therapies 'are oxidative in nature and should litterally "flood
the body with oxygen". But not all "oxidattherapies are "OXYGEN therapies". The following quotes illustrate
further:
"Oxidationhasse
..v e d DOSI~IO~Sof aontml in it* nrvcess in line with our ~twtulate.The first isthe potency
of the K G (Functional Carbonyl Group) which must start the process by dehydrogenating the fuel. When this car-
bonyl group is not free, as when the hydrogen it removes from the fuel is not taken away by some electron acceptor
system, then oxidation is blocked. And for this oxygen is essential as the ultimate electron acceptor in aerobic organ-
isms. So lack of oxygen has two steps in blockingoxidation or hindering it.""
"Our postulate provides for the polymerization of the carcinogenictoxin as it developsto the cancer producing stage,
and this provislon is based upon the chemical and clinical circumstances that stare one straight in the face. Atrophy
precedes neopiasia. If one answersthat the neoplasia is a reactionto the atrophy stimulusas hay fever is to the pol-
len stimulus, one must still offer a mechanismfor the reaction. The simplest mechanismthat could be involved is that
the toxin producesboth changes, and this medianism we have already explained as due to block in energy production
and transfer. Recovery from the state caused by the carcinogenicagent, be it virus or chemical, is therefore a satis-
factory support to the contention, since the same agency accomplishes the correctiaras of all states, atrophy, pre-
growth toxic state, cachexia, and the tumifactions.""
"After the pathogen, be it a virus, carcinogen, or some allergen has made the pathogenic integration,
the need for oxygen in the diseased cells is all the more imperative, and removal of all sources of the
pathogenicamines is the prime consideration. And not until a goad dispersion of the tissue colloids is
had, and a goad oxygen supply is present in the diseased cells should the reagent be given. f i r if it
does not have a molecule of bxvuen at hand tv wmblne with the f m a d b l formedbv each dehvdrv-
gemtion, them will be no curativea ~ ~ o nof oxidation, and the muent is uiven in vain. This also
applies to the free radicals produced by the use of the reducing agent" (Possibly the MOSTImportantstate-
mentever pronouncedby Dr. Koch regardingthe PROPER use of his treabnent(s).)
"The best proof of the correctness or practicabilityof any postulate in medicine is doubtless the curative value of its
app~ication."'~
REFERENCES:
1) Nikola Tesla, Radio Power Will Revolutionize t h e World, Modem Mechanix and Inven
tions, July, 1934, p.2.
2) Ibid.
3) Otto Warburg, The Prime Cause and Prevention of Cancer, 1969, p.16. (Translation by
Dean Burk, National Cancer Institute)
4) Arthur Schopenhauer, International Tesla Society Journal of Power and Resonance,
Vo1.5, No.4, 1990, p.40.
5) Otto Warburg, The Prime Cause and Prevention of Cancer, 1966, p.6.
6) Ibid.
7) F.M. Eugene Blass, Oxygen-Therapy-Blass: Its Development into a Complete Uniform
Treatment of Disease, Oxidation News, Vol. 1, No.1.
8) F.M. Eugene Blass, Oxygen Therapy: ItsFoundation, Aim and Results, 1927, p.1.
9) W.F. Koch, The Sunrival Factor in Nmplastic and Viral Diseases, 1961, 1967, p.257.
lo) Ibid. p.24.
11) W.F. Koch, Survival Factor in Neoplastic and Viral Diseases, 1961, 1967, p.257.
12) W.F. Koch, Neoplastic and Viral Parisitism, ItsBasic Chemistry and Its Clinical Rever-
sal (An Introduction to Free Radical Therapy), 1967, pp.4142.
13) W.F. Koch, Survival Factor in Neoplastic a n d Viral Diseases, 1961, 1967, p.25

4. The Prime Cause
.l'fe
at
tde
at %+t;
h d '8 Of
and Prevention of Cancer
a9 t4c
l q e c *obcl.f by *arburgcell
OQSta .?ore
*c., "res for
Gcr 00
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ht2s y3n~-lns'1tute
j " a ~ ~ C J ~ 1 , v e ~ ~ r > r ,
' 1966
<
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? 2
As cn~pharizcd, it is the first prrcor~ditionof the proposed treatment
that all growing body cells be saturated with oxygen. It is a second pre-
condition that exogenous carcinogens be kept away, a t least during the treat-
ment. All carcinogens impair rrspir;?tion tlirectly o r indirectly hy deranging
capillary circulation, a statenlent thar is proved hy the fact that n o cancer
cell exists, the recpiratior~of which ir trot impaired. Of course, respiration
canrlot be repair[-d if it ic irrrpnircd .it rhc came rime by carcinogcris.
:..
:. :: ::
'1'0 prf*verlrc.anc.rr it i rll(~rc.fc,r-cI>rc)r~c,~c~clfirrt to krep tile rpc.cd o f tlrc
I,lootl <tre;lrrr (I Irigh that t l ~ rc.rlr,u t>lood ti11 rontait~ssufficierrt oxygen:
c(%c.o~icl,t o l<cc)l)igl~the ~ . I ~ I I L ~ * I I ~ I ~ ~ I I ~ ~ ~ Iof l~c~rio~It)l>iriill tlic l~loocl;tl~irclt o
add alfa),r t o tlrc food, c c n oC hc.iltlry pc%ol>lr,tlic active grorlpc o f the
rcspiratol-- cn/yllrcr: anel ro i~~c.rc,nctlic clorc of rlrre groups, if a pre-
cariccrorrs tatc:') lias alrcad, dcclopctl. l i ar tllc .inre time cxogcr~uur
carciriogcri arc cxclrrcicd rigoroucly, rlicn r~rudrof t l ~ rc n d o g c ~ i o u ~cincer
niay bc prcvcrrtccf toclay.
'llicrc propocals arc in 110 -.I- uropiat~.O n tlie rorltrary, they niay be
rcalizc-d h y everybody. c.cr..l~er;c,at all?. I~our.Unlike the prevention of
nialiy otlier direaces tlie prc-crrtio~~of cariccr rrcluirer no government help,
and riot much tiioney.
1. :. .._I_ .. ,.
.l'lrcrr arc. pritrrc ant1 cc~c~orrd.rr~xrrcnf dicc..~rr. f'c~r csariiple, t l ~ v
prime cause of tlic plague is the plagrrc bacillus, but rccontlary causes of rhc
plaguc are filth, rat, and the fleas th.it tranrfer tlie plague bacillus from
rats to rrlan. By a prime cause o l .I dicare 1 meall one that is found in
t'i.'CYJ c.trc. of tlic dirarc,.
Cancer, ahovc 211 otlirr diccacr. 11.1, i~~urrtlrcscc.ondar!. cartscs. Alriiot
anything can cauc. caticer. But. ~ ~ I , I Ilor carleer, thcrc i orily onc primc
e.arrre. S U I I I I ~ I ; ~ ~ ~ / I ~ I ~iri a feu. u.orci. .rIrc pririlc cinc of a n c c r is thr
r~p~.icerti~rrt01 tlrc. resl>ir;itioti 01 i r t trc>rrirSil lu)cly cr!ls hy a
fernletitation of ru!:ar. All ~~c,rmalhotlv cells meet their cncrgy needs hy
rcqpiration of oxyget~,wl~crcacat1c.c.r ccllr meet tlirir rncrgv ncccls in great
lwrt l y fcrtiic~~tatic~rr.All tic~rni~ill ~ ~ c l ~cclls are tl)us oI>l;gatc arrobrr,
,hercar all c.1lrrc.r cells .ire parti;ll .it~aerobcr. 1:roni tlic randpoint c,f
tllc physics and dlrtnirtry of Iifc thic ditfrrcricr hrtween riorn~al;ir~dcancer
cells ir 50 grc.it tliat one can scarrrly pictrtrc .I greater cliflcrc.t~cr.Oxygen
gas, the donor o f energy iri plant5 arid animals is dctllror~cdin the caiiccr
cells and replaced l,y at? energy yirlding reaction of the lowest li,ing forms,
namely, a fcrn~ent~itic~no f glucore.
Eitglish Edition by Dean Burk
National Canccl- Ttitirr~tc,Rcthc~d;l.?.I;lryland, [ISA

5. 111 all).
cac.. c l u r ; ~ ~ ~tlic cAricrI- cl<.c-loliriic.~itr l ~ cos!,g~ri-i-c*~irario~inl,,l< i.ills,
frrriirrltaticin nppr.lr5. .i11t1 rhc hislily ditTcrcnriatcd cclls a r c tran<for~iicdt o
fcr111~lirili~;i~iaCrol>ec.vliich l i i i v ~Ioqt nll tllcir I~ociyfunctions alld rrtain
onlv rlir IIO.r~<clcx.;propcrr!. oi grc~vrll.'l'liur, when rccpiratior~dival~pcars,
lit.<* clor c I i < ~ p p c ~ i r .>III l i t , v , c ~ I ~ I ; I , , ~o f lifr tii;~ppc.~rr,anii w l ~ n tri-111aitis
. ~ r r< r o v i ~ , ~111,ldlinc.rli.ir clt..~ro!. tlic 1 1 1 1 ~ 1 ~ .in i,llich the! grow.
bIa11y cxpcrtt .igr-cc. 1h;lt orir c17ulJ prccnc about 80°.0 of a11 cancers In
man, if one could ;iway rhc knon-n c.ircinogcnc from the nornlal body
cells. 'l.lic prc,entron of cancer ~iiigl,rillvolvr 1 1 0 gre.lt cxpcnses, a11decpecially
would require littlc frlrtlicr rcseard~t o I>ring about canccr Prcvcntion in
u p to 80 pcrcent.
r, :: ':
Most cspcl-tc ngrcc thnt 11cat.l~80'l 11 of cnncerc could be prevcr~ted.if all
contacl with the Itnonn cuc~gcnc~tl<cnrc.inosrr~rc o ~ ~ l t lheavoitlcd. But h o w c a n
the rc%riininin,,:20" (1, t h r c l i d ~ ~ ~ c r i o r l r,r r, c ~ ~ i l l c * t lIloritancous cancers, hc pre-
acntril?
Ilcc:luc n o cnlicer cell ~ s i ~ t r .tlli. r('~~>ir,1liol1of ~vllichis intact I). it cannot
hc tlicputcd tllnt canccr could b~ pl-c,.cnr<~cl;f ~ h rrespiration of the body
cells would hc kept intact.
$4 + C ..-,
W h y does it happen that iri spite of all [hie so little is done towards the
prevention of cancer? T h e answer has always bccn that o n e does not know
what cancer o r thc prilnc causc of cancer is, a n d that one canllot prevent
roriicthi11~that is not known.
Hut nobody today cnn say that onc does not kliow w h a t cancer alid its
prilnc causc is. O n the contrary, there is n o disease whose prime cause is
better known, so that t o d a y ignorance is no loliger a n excuse that one cannot
do ~ n o r ca b o r ~ tpreventioli. 'l'hat tlic p r c r e ~ i t i o no f cancer will come thcre is
n o do~rbt,for m a n wiclies t o si~rvivc.Rut h o w long prevention will be
avoided d e p e ~ i d son h o w long the prophets of agnosticism will succeed in
inhibiting the application of scientific knowledge in the cancer field. In the
1ne.ir~tin1c.niillions of men ~irustdie o f cancer un~lrcessarily.
Literature to Preface of Second Edition: a 3 4
WILI.STAETTER,WIEI,ANDand EUI.FR,1.ccturcs (>ti cni-ynlcs at thc CcrlrcIiary 01
tlie Gesellslhalr Dn~tsclirrNaturforschcr. Rcricl~teder -I)cul.;cl~c~i( : l ~ c r ~ l i ~ c l l ~ ~ i
(;esellrchaft, 55, 35S3? 1922. The 3 lectures of the 3 famous chcmisrs show that
it1 the year 1922 the action of all enzy~iicsn.is ctill a mystery. N o .ic~icgroup
ol any enzyme was known then.
OTTOWARBURC,Biocheni. Zeitschrift, 152, 479, 1924.
O r l o WARRURC,1ie.ivy Mrt;iIs as pros~licri~Groups of vll/yriicr. <:1.1rvo~Io11
I'rers, Oxford, 1949.
OTTO WARBURG,Wasser~tnffiibcrtrag~ntleFcrmcnte, Verlag W'erncr Singer,
Berlin, 1948.
I)FAN~ I J R K ,1941. On the specifirit)- of glycoly5ir in rnaliKn.lrit livrr rulrior~a'.
ccirr~paredwith honi~logousadult or Rrosing liver tissues. in Sympo~ic~~iiof
Respirator Enzymes. Univ. of Wi5consili Press. pp. 235-245. 1942.
I ) F A N RIIRK.1956. SCICIICC,123, 514. 19Sh Y'oods, hl. YX'., S~ir~dfttr~I.K. K..
Ilurk, U., arid Itarle. V. R. J. Nariorr.il (:arrccr lrlsritutr 23, IOi!,-- 1049. Ii)5').
1 ) r . q ~BURK,1964. Burk. I).. Woocis, hl. arid flunter, 1. On tlic SiFriific.iricc of
(;lucolvtis for Cancer Grtrvth, wi~liSpcci.11 Rcfercr~ccre, hifrrri~I<.it 1lcp.1~
Icmas. Journ. National Cnnccr l~irtitutc38. 839-865. 1967.
0. WARRURGund F'. KVBOWITZ,Hioch. %. 189, 242, 1927; 11. (;III I , I ~ I . A T Tuntl
G. CAMTRON,J. Expcr. Med. 97, 52.5, 1953.
(3. WARBURG,17. Musbad~erKolloq~riurn,April 1966. VcrlaF:SprirrKvr, I lciclrl-
l>i,r~,1966. '
0. V('ARRURG,K. GAU-FIIN,A. W. <;FI%I I R, 1). KAYSIR allel ';. ~ . ( > K IN7,
Klitritrhc Wochrnsrhrift 43. 2%9, 196.5.
0. WARBURG,Oxygen, The C:reator of 1)ilfcrc1itiario1l,l)ir~chcnitc.ilI-~irrgriic.
Ac.idcmic Pres5. New Ynrk, 1766.
0.WARBURG,New Methods of Cell Physic>logy,Gcorg Thicn~e,S~utrfi;lrt;nlid
Intcr$cience Publisher$, New York, 19112.

6. .;; Ozone Selectively Inhibits Growth of Human Cancer Cells
. .
Abstract. 7'111~~rutt*tIr~!~IIII~PI~I:Ictrttt,eri~rll.*,lh)t~~l t t t r ~ ~ .hrt*ti.~,~I~II/ttIt,ritrt* rrth11w.s
HYJS side(.rit.(~/yit~/tihitt,tlit1 tr dtj.~t'-tlt./)t*111/1*~11ttrct~tttt-r.h!. o:t,trt* rrr 0.)Io 0.81ttrr1prr
million t~'o:otrt~itr ~r)t~hit~trtuir drtrirtg A tht)..t ~~I't~111tttri~.//I~III~I~I/II~I~tli/~lt)it/)ihrt~.
blusrs srrt-rdt r . ~ttorri.trtrc,cJrtJrrsc.t~tnrr~lt~1~1l.s.7'hc /It c,.ttdt~tr t!/'o:tjnt~ trr 0.1to u..4ptrt
prr milliott i t ~ h i h i ~ t ~ t lt.trtrt,c3r tat,/I protr.tlr 40 cttrtl 61)/~t~rt~c~ttt,rt*.r/~c~r+~it~c~l,y,7'111,nt)tl.
cuni.rrort.1 11111~r.rI/. ~t&t;rt*ttn~~fli*~.tt*tl111 tl~c.~taII*W/.. t,x/~t).rrrt~111tj:t)trt6 tit 0.8pcrrt
prr mill;t~n~trlribitt~tlt,trt~t.t,rt.i*ll grtttcv11ttrtjrt. IIIIIIIMJl)tvtvt3trrt r t ~ t li.ontr1j1t*t*/Igrcjtt,t/r
Irss thun SO pc.rt.mr. Et~it/~~t~rly,~ h ctt~c~r.htrttitnrs~ i j rtlt;fi.trst*rtgtrin.tr t1:onte dtin~trgv
urr inlpuirrd in 11rrtt111t1rrtnt~c~i~v1l.v.
The effects of ozone on hum;tn health
have been o focus of pl~hlicconcern and
scientific investignlion for more than two
decades (14). Considerable atlention has
been devoted to itshessing its cellular ef-
fects (3) because il is thc major citnslitlt-
en1 of the ground.leve1 oxid;rnts in pi$-
luted air, Much hit3 hecn leurncd ahtrt~t
the efTecls of ozone on normal tissues,
but little is known itbttut its action on
cancer cells. We have conducted experi-
ments in which cctnclnuous exposrlre to
ozone at 0.3 pprn (hI electively inhibited
the growth ofh~~rnirncilncer cells 40 per-
cent in 8 di~yh.
Conrrolled levels of ozone (0.3 l o 0.8
ppm) were contint~ot~slygeneratedby 111-
trrviolel irridii~tiunol' filtcred itmhicnt
air. The ozon;rtcd itir, contitining S per-
cent curhon dioxide, W~IS introduced illit
constitnt flow rille of 4.0 IitcrJmin into ;In
environmcnti~lck;lmhcr in itn incuhntor
mitintitineJ ;I! 37°C (t:ig. 11, 'l'hc u?one
Icvels were u:tytbd ditily with tt spcWrb
photometric i t r o l ) ~.i~n;~lyrer.I:or con!-
piirison, nonc;tnccroirs humicn lung dip-
loid fibroh1;tsts (7) were oultitrcd in tho
chitrnhcr itlong with thi* cancer cells. The
cancer cells werc fl.orn itlveulur ( I ~ ~ n g l
tidcnr~c;rrcinorn:tuby). bre;trt'adcni~c;~rci-
nomus ( 9 ) .uterine cnrcinos;rrcom;~s.and
endornetri;tl citrcinom;~h (10). All thc
cells were grown in 60-mm petri dishes
Fi&.I.Schemu~icdi;rgmm (not
to scdC) of the rytcm rlaedfor
culturing humun.cells in ctP+
natcd umbicnt ..rir. Fillered
unbicnt air wus mixcd with
cubon dioxide (5 percent)and
introduced inlo n duul chum-
ber incubator lNutional 335 1).
in 10 rnl of medium and were placed in
the ch;trnher at the surne time. Contrd
cclls wcra inctrbirred in an ui{joiningcom-
purtmcnt reccit in^ fillered urnbient air
contilininl: 5 percent c;trhlm dioxide (4.0
litcr~rnin~.Three petri dishes for each cell
type rr t.rc removed from each of the two
c.omp;rrtmcnlsewry 48 hours for 8 days,
and thc nurnher of cells per plate werc
co~~nted.All of the canccr cells showed
marked dose-dependent grow.th inhibi-
tion in wont. iit 0.1itnd 0.5 pprn (Fig. 2).
'I'hcrc t w s no yrowlh inhiYtion of the
nons;ttrccrolls Itrng cells at these alone
level, and they were murphalogically
identi~itllo the corresponrlinp conlrol
cells. At 0.8 pprn. lhc growth of the non.
ci~nccrvuscells Wits inhihrled SO percent.
httt it11 fullr types of'cilncer cells we:% in-
hihilt4 n1or.c thirn Wl percent.
Al'lcr hti11ycul1111.cdthri~trgh(14 pas-
hoy~.the noncirlrccrotis cells e~hibited
nrt*;ln~r;thlc growth inhibition rtnd mor-
pholtyicill ~ h i t n g ~ stvecuolation) ' in
irrimc ill 0.5 ppln, sttggctiny th;rt.oginp
increases the sensitivity of norrnnl lung
cells 10 oxonc (Fig. 3), In cultured 'hu-
nlitt1 dipl1)id fihrohl;hrls, morpholopicnl
ch;tngcs :tnJ :Igrad11;llJt.creitse in rate of
growth h:tw hcen ;~ttribtrttdto a buildup
of ccllul;~~d;tm:tgc with eitch st~ccessive
division (11. 12). Ozone may accelerate
processes similar l o those nJurully
Half was conducted through u 1 I
ul~braledozunc ~eneritor((;r c
con$i.ctin~of a quaflz PYM
tube irrudiatcd with ultravtolet B
light and then into u herme~i. J -Aully senled (?O hy 20 by 20
em) glass rnd ntrinlcsr stcel
. cnvironmcntul chiunhcr (C')
containing u ~;txhe~edw i e s s
d m . Output ofoxcMc from the pnemIMvaried lens thitn Ipercent per duy. t h e ozone content
of the vented icir ('I frvm Ihc chamher ~ 8 1 smeuuurt-dJail) with r spccttophi~tomctricuzctne
malyzet.(Dirbibi IO3.AH). Malignant rnd normill h~~rn;tncclls werc incubated in chumkr E
satumttd with water vupctr. Comspnndi~cells reruing it ct~ntrolrwere incubated in the 4.
joining comprrtment (('1, alsa utuntcd v ~ t hwu~crvrtprlr.
a SCIENCE. VOL. 209. 22 AUGUST I'M) Wth WU7.(~11o.II(??.WZISN~clH) Cup)nyhl b IW AAAS

7. causin~cellular damage and may de- Evidently, cancer cells are Ics ilhle to
crease the arowth rate of fhr aging compcnsnte for the oxidative tr~lrdcno f
fibroblast colony. However, in'ozone ut otonc than normal cclls. 7'hc mitrtied
0.5 ppm, dl o f the human cancer cdls sensitivity orcancer cells to ozone rilises
(which do not age) had growth rates questions about the possible mcchs-
several times lower than that of the nisms o f oxidative inhihitwn o f their
wed, noncancerous cells (Fig. 2). growth. Virtually every m;t.ior com-
Fig. 2. Inhibition by ozone of roo
growth of malignant and non-
malignant cclls in culture oa
day 8. Each of the cell types
were grown in 10 ml of DJI-
&GO'S modified EMe's mini- 80
mum esscntial. medium con-
toipirg 10 percent calf serum. f
In a typical experiment, 12
dishes pcr cell lina (urudly
three or four cell lines were 80
t e e r e n w e
loaded into the environmental
chamber with an equal number
of control dishes in the adjoin- ging compartment (Fis. I). The
initid population was 3 x 18
cells,per dish. Every 48 hours g
three dishes for each cell type
were removed from both com-
pnmcnts and the cells were W
tested for viability with 0.4 )
percent trypan blue and
cwnred w~rha hemocytom-
etcr. Each data point rtpre-
wnlr the number d expcri- 0.3 0.5
mcntJ cells divided by the
0.8
number of correspondinpcon-
Ozone (ppm)
11-01 cells per dish multipliedby 100(thepercentage of ccotrol growth) md is plottedagainst the
measurrdlevelof,ozone in the environmentalchamber. The percentage ofgrowth inhihition is
calculated by subtractin8the percentage of grtiw~hfrom la).The dataore from ccll collntingon
duy.6 of incubation. There is a nearly linear rcla!ion between inhibition of the growth of the
cancer cells and increasingozone levels. The noncancerouscell line IMH-W) (A)bcpim to dis-
play measurable growth inhibition only when oz,onc levels exceeded$5 ppm. a level that pro-
dufcd approximately 60 percent inhihition in it11 of the cancer cell lines tested IA, ;llvcolar
adenocarcinoma(A-W9); 0,breast adenocarcinnma (MCF-7);0.uterinecarcinosarcomn110);
O, endometrialadenocarcinoma(lo)].Then was some srowtb inhibitioninnoncancerouscdls
aged through 14 passnges (0).The mcnn populations of the cells serving as controls were as
fdlp;vs (per dish on day 8): IMR-90, 34;8 x lq: A-549. 36.5 x IF; MCF-7. 57.0 x los; en-
Jomctrial adenwarcinoma, 64.2 x loS;myomelrialcarcinosarcoma. 121.1 x 10".
. .. .
! ,
Fip.3. Photomicrographs( x 100) showing growth inhibt~ionand morphologicalchanges in lung
dvcular adcnocarcinomacells after IIdrylr dinc~~hutioninozone et 0.5ppm. (A) Control A-549
cellh, (U) Ozone-treated A-W9 c 1 5 ahirwing viic1101e formation, a typical morphulogicd
change urscrcialed with uowth inhibition.
932
poncnt o f normal cells has been found to
be ~ffectedby elevated ozone levels ~ 7 ) .
However. glt~tathione in its reduced
form (GSH) has been credited with pro-
viding the first line of defense against the
peroxides and free rddicals generaled in
all cells by ozone and oxygen (1. 13-15),
It deactivates peroxides and rddicals by %
donating one hydrogen atom to the rcac- '
tive species. Loss of a GSH hydrogen
loxid;~tion) rcsulrs in formation of oxi-
dized glutathione (GS-SG). The cellular
respiratory system is responsible for re-
ducing,GS-SG to GSH.The GSH-linked
respiratory system in normal and cancer
cells. before and after exposure to
ozone, must be examined to learn wheth-
er ;If ~ ~ n c ~ i o n dimpairment of this system
is associated with the marked sensitivity
of cancer cells to the oxidant.
These findings lead us to believe that
o7one-alone, in combination with radi-
utic~ntheri~py(16),or in chemotherapy
utilizing electrophilic compounds (17)-
may have therapeutic value for patients
wit% certain fi?rms of lung cancer.
FILI>LRICK S W ~ E T
MING-SI~IAN KA.
SONG-CHIAU D. LEE
Dr.purtnrr,nt oJOhstc~lric~crrrd
C;?.n~c*ologp,Wt~shingronUnil,rrsity
Sr.lioi)l of Mt*d(c.itir.
St. Lrtitis. Missorrrt'63llO
WILL L.HAGAR
City of St. I.rruis Air Pt)llution
Control, St. Liwis 6303
WILEENE. S W ~ E T
Air Qrrcrlity S~rrion,E~ISI-Wesr
(;(I rot,(ty Coordirrurin~Council,
SI. Lortis 63102
R d a m n d Ndta
I.D. I..Dunwarth, C. E.Cross. J. R.Oilteapic,
C. G. Ploppcr, in Ozone Chrn~isrryand Tet.h-
noltrxr. J. S. Murph and J. R, On. EB.
(FranklinInslitu!c. ~hiradclphin.1973).chsp.2.
2. H. E. Slokinlcr and O. Coffin, inAlr P~rlkrtion.
A. C. Stem. W. (Academic Rers, New YocC.
Iwnl. vcd. I.p f .446-546.
3. H. D. Kerr rl ul.. Am. Mrv. Nrspir. Dis. Il!
761 lt97.5).
4. J. n.Hackney. W. S. finn. C. D. L w . S. K.
Krm7.. Orccnhcr~.R. D.Backley. E.E. Ped-
crsen, Arch. E n ~ . ~ r r ~ n .Hrrrlrh 30. 385 (1973).
5. 8. D. C;~)Ids~ein,h i , .t,nvircm. Iiculfh 2, In
11977).
b. Ncwmsl httrnvn ruhjerts loleraled breathin80.3
ppm ozimr in air 2 hours p r day for Iweek a
0.25 ppm ozone 2 hours per day for 3 weeks (I).
The two gri~~pseny~gdinli&hrexercised u r k
thposurc. Allhqh bolh poups developd
chest disccnnfmand moderately docreasedrrs.
rMarry functionduriwexposure, their removal
romthe oxidativeenvironmenlresultedia rapid
disapwarmcc or the symptom.. nK mean
doserer onrccurvesfrom this ~tudyshow r no.
d*eetrhf'-emcct lcvcl at 0.25 to 0.50 ppm. A
rimilar rrudy LO ftund that humanwbjrclr 101-
erared exposure lo 0.5 ppm ozone for up to 6
hnurb. Pulmonary functlun wur .Ilccled and
chent diaromkrldevelopedat this level. with no
rignihcw~ diUerences abrervtd &we-
amokera Yd nonsmokcn.
7. Thcbr cells {IMR-90)were obtained from
Human Adtq Cdl Reposilwy and plated r,
hnursnher r h ~piw T h i s ccU r y p w u c h u r -
tcrizcdby W, b.~/rho)s;D.G. Murphy. V:J.
Cri!lohlu, L.H. Toji. A. E. tircenc, and St A.
htyht (Sc.irnr,r 196,61(l1971),
SCIENCE. VOL. 209

8. Inactivation of Human l m n ~ u n o d e f i c i e n c yVirus Tyve 1 by Ozone I n Vitro
By Keith H. Wells, Joseph Latino, Jerr~eGavalchin. and Bernard J. Poiesz
A device was designed to deliver a constant source 01ulrrll
concentrations of ozone to fluids containing human immuno-
deficiency virus type 1(HIV-1).Ozone was found to inactivate
HIV.1 virions in a dore-dependent manner. Greater than 11
log inactivation was achieved within 2 hours at a concentra-
tion of 1,200ppm ozone. Similar concentrations of ozone had
minimal effect on factor Vlll activlty in both plasma and
immunoaffinity-purified preparations of factor Vlll treated
HUMAN immunodeficiency virus type 1 (HIV-I). the
etiologic agent of acquired immunodeficiency syn-
drome (AIDS), is a lentivirus that cornpleles its replicative
cyclc by budding through the host ccll membrane. acquir~ng
host-derived and v~rus-encodedcomponents in thc process.
Subsequent rounds of replication requlre an Intact lipid
envelope containing the viralk encoded envelope proteins
necessary for receplor binding.' 11has been suggested that
perturbation of the HIV-I envelope may be a suitable
approach to inactivating HIV-1.: Compounds that fluidize
membranes by removing cholesterol (AL721)-'or by com-
plex~ngmembrane cholesterol (umpho~ericinB methyl
ester)'can inhibit HIV-1 replication in vitm. Although their
precise mode of action has yet to be defined. these
compounds may reduce HIV-1 infectivity by perturbing the
envelope of HIV-1. We therefore investigated the activity
of another membrane active agent, ozone, on HlVLl
infectivity in vitro.
Ozone. the triatomic allotrope of oxygen, is an extremely
potent oxidant that has been shown to possess broad
spectrum antimicrobial activity."' It has been widely used in
sewage treatment. in water purification. and in medi~ine."'~
In particular. ozone has been shown to be effective against a ,
number of enveloped and nonenveloped viral species. with '
enveloped viruses being more susceptible to ozonc inactiva-
tion than those lacking lipid envelopes.'."" We report on
the in virm inactivation of HIV-I in cell culture media and
dclibcrately infected factor VI I1 preparations at concenlra-
tions of ozone that are not toxic to target cells, while
maintaining the biologic activrty of factor V111. We also
investigated the mechanism by which Ozone mediates its-
antiviral effect.
- 1 - , . -:
- -
for the same time period. The data indicate that the antiviral
effects of ozone include viral yrticle disruption, reverse
transcriptase inactivation. andlor a perturbation of the abil-
ity of the virus to bind to its receptor on target cells. Ozone
treatment offers promise as s merm to inactivate human
retroviruses in human body fluids and blood product prepara-
trons.
r 1991 by The American Society of Hematology.
Ozollc has previously been shown to possess potcnt
antiviral activit~.cspcciall$ whcn used against liprd-
cnvclopcd viruscs. For this rcason wc inveslrgatcd Ihc usc
01 iIzonc ;I% a polrntral i~nt~-MlVI ~pcnt.Wc used a hollou
lihcr J c l ~ v c nsybtcnl that mdxrrnlzcs the >url'acc arcit
a,ailablc for ozone to interact with the fluid rnal'crial ol'
interest. The syslem also allows for the precise regulation of
concentrations of ozone to be delivered into the hollow
fiber cartridge. Afferent as well as efferent concentrations
of ozonc can also be monitored to determine ir-saturating
levels of ozone are achieved in the treated material. With
regard to laboraton. safety concerns, this closcd system has
proven to be safe and Icak-proof, thereby reducing possible
exposure of laboratory personnel to ozone and human
retroviruses to an absolute minimum.
We first examined the ability of ozone to inactivate
cell-free HIV-1 in ?ell-free CM. These results indicate that
ozone has potcnt anti-HIV-1 activity. Preliminary experi-
men15 using an escalating dose regimen indicared that a
1.200 ppm dose of ozone achieved a 2 2 ly ~nactivationof
virus. Thc data s h o ~that neither incubation of the virus
preparation at room temperature and at atmospheric condi-
tions for the duration of the experiment nor mechanical
sh&r created b~ the virus through the syslcrn
inactivate the virus to any great extent. However. therc was
a signiticant inactivation of the virus due to exposurc ol'
virus-containing CM to :h~.slrcam of carrier nitrogen.
Exposure of the virus to purc nitrogen for 6 hours results in
an almost 85% inaclivntion of virus.
From rhr Deponm& of~?dtcrnrand ~ ~ r v b ~ o l o g v~lmmu;ldlo&
SUNY Health Science Ccwer. S,vmcw. W, rhu Drpunmrn~of
Hcmatolog?:Oncul~.Thr Brwklyn Hosprral, Nm York hY. utui
rhe Blolopcal Techn~calSrrvrc~s.Mcrck Phannlrcruncal. Munufucrur-
-mg Dtvrsron. Wrsr Pornr. PA

17. 136 AN INTRODUCTION TO FREE RADICAL THERAPY

18. TERMINATION OF THE MALIGNANT PHASE 137