Treatment of Lowland Frogs From the Spawn Stage with Homeopathically Prepared Thyroxin (10 -30 )

Abstract

The influence of a highly diluted agitated, i.e. homeopathically prepared thyroxin solution (10(-30), final concentration in the basin water 10(-35) parts by weight after the first application) on metamorphosis in lowland Rana temporaria from the spawn stage on was studied. The treatment with homeopathically prepared thyroxin solution (10(-30)) starts at the frogspawn stage. It represents a tool to learn more about the previously standardized amphibian model, where the thyroxin solution was applied from the two-legged stage on only. Lowland frogs were pretreated by immersing spawn in an aqueous molecular thyroxin dilution (10(-8) parts by weight). In later stages of development (2 to 4 legged), this has been found to speed up metamorphosis by around 15%. In accordance with the homeopathic idea of detoxication or cure, hyperstimulated animals (spawn or, in subsequence, larvae) were treated either with thyroxin that had been highly diluted and agitated in successive steps, i.e. homeopathically prepared (10(-30)), or analogously prepared blank solution (water). Development was monitored by documenting the number of animals that had entered the four-legged stage. It has been found that animals treated with the test solution metamorphosed more slowly than the control animals, i.e. the effect of the homeopathically prepared thyroxin was opposed to the usual effect of molecular thyroxin. The number of test animals that reached the 4-legged stage at defined points in time was slightly smaller in the group treated with homeopathically prepared thyroxin at some, but not at all points in time, compared to control. The results in this study sustain the previous multi researcher findings that highly diluted homeopathically prepared thyroxin is able to slow down metamorphosis of Rana temporaria.

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Brief Research Report
TheScientificWorldJOURNAL, (2007) 7, xxx–xxx
TSW Holistic Health & Medicine
ISSN 1537-744X; DOI 10.1100/tsw.2007.220
*Corresponding author.
©2007 with author.
Published by TheScientificWorld: www.thescientificworld.com
1
Treatment of Lowland Frogs From the Spawn
Stage with Homeopathically Prepared Thyroxin
(10
-30
)
Helmut Graunke
1
, P. Christian Endler
1*
, Waltraud Scherer-Pongratz
1
, Michael Frass
1
and Harald Lothaller
2
1
Interuniversity College for Health and Development, Graz, Castle of Seggau, Austria;
2
University of Graz,
Austria
E-mail: college@inter-uni.net
Received June 1, 2007; Revised July 27, 2007; Accepted July 27, 2007; Published
The influence of a highly diluted agitated, i.e. homeopathically prepared thyroxin solution
(10
-30
, final concentration in the basin water 10
-35
parts by weight after the first
application) on metamorphosis in lowland Rana temporaria from the spawn stage on was
studied. The treatment with homeopathically prepared thyroxin solution (10
-30
) starts at
the frogspawn stage. It represents a tool to learn more about the previously standardized
amphibian model, where the thyroxin solution was applied from the two- legged stage on
only. Lowland frogs were pretreated by immersing spawn in an aqueous molecular
thyroxin dilution (10
-8
parts by weight). In later stages of development (2 to 4 legged), this
has been found to speed up metamorphosis by around 15%. In accordance with the
homeopathic idea of detoxication or cure, hyperstimulated animals (spawn or, in
subsequence, larvae) were treated either with thyroxin that had been highly diluted and
agitated in successive steps, i.e. homeopathically prepared (10
-30
), or analogously
prepared blank solution (water). Development was monitored by documenting the
number of animals that had entered the four-legged stage. It has been found that animals
treated with the test solution metamorphosed more slowly than the control animals, i.e.
the effect of the homeopathically prepared thyroxin was opposed to the usual effect of
molecular thyroxin. The number of test animals that reached the 4- legged stage at
defined points in time was slightly smaller in the group treated with homeopathically
prepared thyroxin at some, but not at all points in time, compared to control. The results
in this study sustain the previous multi researcher findings that highly diluted
homeopathically prepared thyroxin is able to slow down metamorphosis of Rana
temporaria
KEY WORDS: amphibian, hormone, thyroxin, homeopathic dilution, curative effect

Graunke et al: Treatment of Lowland Frogs TheScientificWorldJOURNAL (2007) 7, xxx-xxx
2
INTRODUCTION
Intoxication – detoxification experiments are an important tool in research on homeopathy[1,2]. Due to
the relation between an enhanced thyroxin level during metamorphosis[3] and homeopathically prepared
thyroxin, a model with amphibians seems to fit well into this category[4,5].
Highland Animals and Thyroxin 10
-30
In previous experiments[7,8,9], we used Rana temporaria larves from alpine highland populations,
assuming that the highland animals’ natural thyroxin level is comparatively high and that the increase in
environmental temperature causes an artificial acceleration of metamorphosis, when the animals are
brought from the highland biotopes to the laboratory. Animals were treated with homeopathically
prepared dilution of thyroxin 10
-30
(final concentration in the basin water 10
-35
) from the two-legged stage
on. In five independent laboratories in Austria and the Netherlands, it was found that the dilution
diminished the speed of metamorphosis (3-11%). Differences were statistically significant (p < 0.01 or <
0.05 at the measuring points in time). Frequencies of animals treated with homeopathically prepared
thyroxin reached the values of control animals with a delay of about ½-1 intervals between points in
time[7,8,9].
0
200
400
600
800
1000
1200
1234567
Figure 1. The effect on highland Rana temporaria, added
from the two-legged stage on; pooled data from 1990 and
2000 [7-9]. Ordinate = cumulative frequency of 4- legged
tadpoles (N). Abscissa = points in time. Black squares =
frequencies of animals treated with homeopathically
prepared thyroxin; white squares = of animals treated with
analogously prepared water.
Lowland Animals and Thyroxin 10
-30
When Rana temporaria from lowland biotopes were used in the same experimental set-up, starting
treatment at the two- legged stage, no difference was found between the test and the control group (p >
0.05)[6].
Hyperstimulation
In further experiments we observed that hyperstimulation of lowland animals with molecular thyroxin
(final concentration in the basin water 10
-8
) was able to enhance animals’ responsiveness to
homeopathically prepared thyroxin[6]. However, this was applicable only to treatment with

Graunke et al: Treatment of Lowland Frogs TheScientificWorldJOURNAL (2007) 7, xxx-xxx
3
homeopathically prepared thyroxin 10
-8
, but was not applicable to treatment of lowland animals with
thyroxin 10
-30
. Nor did hyperstimulation enhance the observed effect of thyroxin 10
-30
in highland
animals[10].
In the following study, the influence of thyroxin 10
-30
on metamorphosis in hyperstimulated lowland
Rana temporaria was studied. Different to our previous studies, where treatment started at the two-legged
stage, in the study presented here, animals were treated from the spawn stage on.
METHODS
Researchers and Blinding
The experiments were carried out at the laboratory of the Interuniversity College and the study planned by
Graunke and Endler. Application of the homeopathically prepared thyroxin or solvent was done blind.
Animals, Staging, Water and Further Laboratory Conditions
Four days old Rana temporaria spawn were taken from an Austrian lowland pool. The larvae (tadpoles)
were treated and observed during seven weeks until both, the hind and forlegs, broke through the skin and
the animals had thus entered the four-legged stage.
20 eggs were allotted to each of a total of 75 white plastic basins according to a random procedure.
Basins contained 6 l of dwell water each. Indirect natural light was used. Room temperature was 21 +
1°C. The tadpoles were fed with blanched greens (lettuce) ad libitum. The experiment was carried out
between April and June 2006.
Preparation and Administration of Hyperstimulation and Test Solutions
Two groups of animals were exposed to the stock solution of tetra-iodo-thyronin sodium pentahydrate
(T
4
, Sigma, 10
-4
parts by weight in double distilled water, diluted in the basin water down to a final
concentration of 10
-8
) (immersion in thyroxin 10
-8
, hyperstimulation). This is a standard procedure in our
laboratory derived from studies with Rana temporaria from lowland biotopes, where it speeds up
metamorphosis by about 15%[6]. One group of the hyperstimulated highland amphibians was then treated
with the homeopathically prepared test dilution, while the other was treated with the analogously prepared
solvent:
• thyroxin 10
-8
+ thyroxin 10
-30
(hyperstimulated test group)
• thyroxin 10
-8
+ water 10
-30
(hyperstimulated control group)
The third group was treated with
• water 10
-30
(inert control group).
For preparation of the test dilution thyroxin 10
-30
, the stock solution (10
-4
), was diluted with pure double
distilled water in 26 steps of 1:10, and agitated after each step of dilution according to a standardized
protocol. Analogously prepared solvent was used as a control (water 10
-30
).

Graunke et al: Treatment of Lowland Frogs TheScientificWorldJOURNAL (2007) 7, xxx-xxx
4
3 μl of molecular stock solution (10
-4
) was added per animal and 300 ml of basin water of the
hyperstimulated groups (final concentration of the hyperstimulation solution was 10
-8
after the first
application) at intervals of 2 weeks. 3 μl of probe dilution (test or control) was added per animal and 300
ml of basin water (final concentration of probe dilution was 10
-35
after the first application) at intervals of
48 hours.
Comparison and Evaluation of Data
After reaching the four-legged stage the cumulative frequency of animals treated with test or control was
aggregated for each day. Treatment was started on April 29
th
. On June 10
th
the first animals in a four-
legged stage were observed. According to the reasons described in study[14], chi-square tests were used
to compare groups at the points in time. Further statistical methods used in connection with the amphibian
model have been described in[6,9].
RESULTS
500 animals were treated with thyroxin 10
-8
plus thyroxin 10
-30
(hyperstimulated test group), 500 animals
with thyroxin 10
-8
plus water 10
-30
(hyperstimulated control group) and 500 animals with water 10
-30
(inert
control group).
As shown in figure 1, animals treated with the test solution (black squares) metamorphosed more
slowly than the control animals (white squares). The number of test animals that reached the four-legged
stage at defined points in time was smaller in the group treated with homeopathically prepared thyroxin at
most points in time. Differences were statistically significant (p < 0.01) at 3 points in time. Standard
deviation was about + 2.5 to + 3.0 (a high value for SD is typical for hyperstimulated groups[6]. Animals
treated with water 10
-30
only (not hyperstimulized inert control group, black lines) were the slowest group.
0
100
200
300
400
500
1357911
Figure 2. The effect of thyroxin 10
-30
, on lowland Rana
temporaria, added from the spawn stage on. For further
explanation, see legend to figure 1.
The data in figure 2 shows a difference of about 1-10%, i.e. frequencies of animals treated with
homeopathically prepared thyroxin reached the values of the control animals with a delay of about 0.1 – 1
day (e.g. June 15
th
) or did not reach the values of the control animals at the end of the experiment
respectively. P-values are given in table 1.

Graunke et al: Treatment of Lowland Frogs TheScientificWorldJOURNAL (2007) 7, xxx-xxx
5
TABLE 1
For explanation se text
hyp+WD30: hyp+WD30: hyp+TD30:
time (days) hyp+TD30 inert inert
1 0,48 0,48
2 0,62 0,62
3 0,77 0,37 0,54
4 0,57 0,00 0,02
5 0,26 <0,001 0,01
6 <0,001 <0,001 0,01
7 0,05 <0,001 0,07
8 0,46 0,06 0,24
9 0,64 0,06 0,17
10 0,56 0,25 0,57
11 <0,001 <0,001 0,61
12 <0,001 <0,001 0,93
DISCUSSION
In 1990, inspired by older literature on the influence of metal salts[11], we standardized an experimental
model based on the influence of highly diluted thyroxin (10
-30
) on inert Rana temporaria from highland
biotopes. Results from a study in 1990[7] were confirmed by ourselves and others in 2000[6,9]: speed of
metamorphosis was reduced by diluted thyroxin (10
-30
) in all five laboratories involved.
In order to gain a physiological explication for these results, and further to optimize them, we added
hyperstimulation with molecular thyroxin to our initial standard protocol.
Using such hyperstimulation, we were able to include animals from lowland biotopes into our study
line. This was interesting with regard to the limited availability of highland amphibian. However, lowland
animals, even when hyperstimulated, reacted to homeopathically prepared thyroxin only in 2/3 of
experiments and, furthermore, only to a low dilution of thyroxin (10
-13
). Thus, our hope to have found a
robust experimental set-up involving lowland animals was deceived[6]. On the other hand, we learned a
lot about the model, namely that if a difference between test group and control group is found in one
laboratory at all, this difference is considerably greater when hyperstimulated animals than when inert
animals are used[6]. This led to the possibility to explain our amphibian model as an intoxication-
detoxification model analogous to other models used in homeopathy research[4].
From our studies we conclude that there appears to be a relationship between the effect of
homeopathically prepared thyroxin and a naturally or artificially elevated thyroxin level in the animals
during metamorphosis. It is reasonable to suppose that highland larves of Rana temporaria have become
adapted to an environment which necessitates a comparatively high thyroxin level or high sensitivity to
thyroxin[12,13]. This would be a plausible explanation for their consistent response in experiments with
diluted thyroxin.
This assumption is backed by the observation that, in normal lowland animals, immersion in
thyroxin 10
-8
parts by weight leads to an increase of 4- legged animals of about 15% compared to
immersion in plain basin water, whereas in highland animals, thyroxin 10
-8
causes an increase of only
around 5%[14].

Graunke et al: Treatment of Lowland Frogs TheScientificWorldJOURNAL (2007) 7, xxx-xxx
6
Furthermore, the increase in environmental temperature causes an acceleration of metamorphosis
when animals are brought from the highland biotopes to the laboratory. This acceleration is likely to be
“cured” by homeopathically prepared thyroxin.
The present results permit the conclusion that administering homeopathically prepared thyroxin to
amphibian larves during their thyroxin-controlled metamorphosis is in a certain sense analogous to the
intoxication - detoxification concept used in other models of homeopathy research[4], although the
intoxication dose and its effect on responsiveness do not seem to correlate in a linear way.
Apart from its inherent interest for physiology and medicine research, the amphibian model seems to
be a helpful tool to investigate environmental influences on homeopathic drugs[14].
ANNOTATION
The Interuniversity College would be pleased to collaborate with other institutions in order to prepare
further independent repetitions of the experiment standardized in 1990 and repeated in 2000[7,9]. Such
collaborations should include initial training at the laboratory of the College plus an independent
experimental phase at the guest’s own laboratory. Reprints of published papers can be downloaded at
www.inter-uni.net/edition. The process of “trial and error”, but also of “trial and success” in amphibian
research has been described in the journalistic book “Homeopathy Research – An Expedition
Report”[15].
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Graunke et al: Treatment of Lowland Frogs TheScientificWorldJOURNAL (2007) 7, xxx-xxx
7
15. Endler, P.C. (2003) Homeopathy - An Expedition Report. An old healing system gains plausibility.
edition@inter-uni.net, Graz.
This article should be cited as follows:
Graunke
,
H., Endler, P.C., Scherer-Pongratz, W., Frass, M., and Lothaller, H. (2007) Treatment of lowland frogs from the
spawn stage with homeopathically prepared thyroxin (10
-30
) TheScientificWorldJOURNAL: TSW Holistic Health & Medicine
7, xx–xx. DOI 10.1100/tsw.2007.220.