A series of experiments have been conducted with ten adult rabbits, drowning them to death in a ditch those water contains diatoms in abundance. The bones (selected ones are the femur, humerus, riHand vertebra) of these drowned rabbits have been buried underground, wrapped tightly in cellophane bags and left there for three years, and the detection of diatoms has been conducted with these bones either as they are or after cremating them in the electric'oven at 300°C, 500°C, 800°C or 1,000°C, for 20 minutes. As the results it has been clarified that diatoms can be detected in a considerable number in the bones of four limbs, and of these detectable diatoms some of them can be found even after cremation at 1,000°C for 20 minutes. This clearly proves diatoms are detectable from the bones even after a long period of time·after burial and even after cremation at high temperatures.</p

Okuyama, Masami

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Experimental studies on diagnosis of death from drowning by means of detection of vegetative planktons (diatoms) II. Detection of diatoms from putrefied and cremated bones of drowned bodies

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Acta Medica OkayamaVolume 15, Issue 4 1961 Article 5AUGUST 1961Experimental studies on diagnosis of deathfrom drowning by means of detection ofvegetative planktons (diatoms) II. Detection ofdiatoms from putrefied and cremated bones ofdrowned bodiesMasami Okuyama∗∗Okayama University,Copyright c©1999 OKAYAMA UNIVERSITY MEDICAL SCHOOL. All rights reserved.Experimental studies on diagnosis of deathfrom drowning by means of detection ofvegetative planktons (diatoms) II. Detection ofdiatoms from putrefied and cremated bones ofdrowned bodies∗Masami OkuyamaAbstractA series of experiments have been conducted with ten adult rabbits, drowning them to deathin a ditch those water contains diatoms in abundance. The bones (selected ones are the femur,humerus, riHand vertebra) of these drowned rabbits have been buried underground, wrappedtightly in cellophane bags and left there for three years, and the detection of diatoms has been con-ducted with these bones either as they are or after cremating them in the electric’oven at 300◦C,500◦C, 800◦C or 1,000◦C, for 20 minutes. As the results it has been clarified that diatoms canbe detected in a considerable number in the bones of four limbs, and of these detectable diatomssome of them can be found even after cremation at 1,000◦C for 20 minutes. This clearly provesdiatoms are detectable from the bones even after a long period of time·after burial and even aftercremation at high temperatures.∗Copyright c©OKAYAMA UNIVERSITY MEDICAL SCHOOLActa Med. Okayama 15, 261-267 (1961)EXPERIMENTAL STUDIES ON DIAGNOSIS OF DEATHFROM DROWNING BY MEANS OF DETECTIONOF VEGETATIVE PLANKTONS (DIATOMS)11. DETECTION OF DIATOMS FROM PUTREFIED ANDCREMATED BONES OF DROWNED BODIESMasami OKUYAMADepartment of Legal Medicine, Okayama University Medical SchoolOkayama (Director: Prof. Y. Mikami)Received for publication, April 20, 1961As introduced briefly in the first report, for the diagnosis and the determi-nation of the place of death from drowning, the detection of vegetative plank-tons (diatoms) from the organs and bones now serves the purpose in legal medi-cine. The theoretical basis for the detection of diatoms has been established bymany investigators (1895-1955)1-9. In Japan TOMONAGA et al. (1954)10-20, andMIKAMI and his co-workes (1959)21.22 made some detailed studies on this problem.However, similar studies on the possibility of detecting diatoms from the crema-ted bones and the bones that had been·buried for a considerable period of timehave not yet been studied much in details.In the previous experiments attempts were made to detect diatoms fromthe incinerated bones of drowned rabbits and also it was possible to detectdiatoms even from the bones cremated in the electric oven at high temperature.It was then pointed out that the diagnosis of death from drowning was possibleby examining the bones of the corpse (1961)23. In the present paper the resultsof the detection of diatoms are described about a series of experiments conductedwith the bones of drowned rabbits, buried for a relatively long period of timeand also with the bones cremated or incinerated.MATERIALS AND METHODSThe site of drowning is the same as in the previous report (1). Ten adultrabbits weighing about 2.5 kg. served as the experimental animals. The waterof the ditch at time of drowning rabbits was used for the detection of diatoms, i.e. about 100 ml. water was centrifuged and the sediment placed on object glasswas examined under microscope. Thus the kinds and approximate number ofdiatoms were estimated, and these served as the control.The drowned rabbits were immediately dissected for extracting the bones(the femur, humerus, rib and vertebra), and ,these bones of each rabbit were2611Okuyama: Experimental studies on diagnosis of death from drowning by meansProduced by The Berkeley Electronic Press, 1961262 M. OKUYAMAwrapped tightly in a cellophane bag and buried in the ground about a meterdeep. Three years later these bones were unearthened, and the detection ofdiatoms was conducted. These were divided into five groups of a, b, c, d and e(each group consisted of the bones of two rabbits). The bones in each groupwere washed with distilled water and the surface of the bones dried pri<?r to theuse. The amounts of each bone used were about 11 g. of the femur, about 6 g.of the humerus, about 5 g. of the rib and about 10 g. of the vertebra. In thecase of Group "a", the bones were used as they were just as in the previousexperiments. Each bone is thoroughly disintegrated with 30 ml. fuming nitricacid in the identically same manner as in the previous experiments. To this 30per cent hydrogen peroxide (10-20 m!.) is added until the liquid becomes com-pletely colorless. This transparent liquid is transferred to spitz glasses and centri-fuged at 3,000 r. p. m. for 20 minutes. The number and kinds of diatoms inthe entire field are examined in the same manner as in the previous report.The bones of Groups "b", "c", "d" and "e", are cremated it the electricoven at 300°C, 500°C, 800°C and 1,000°C, respectively for 20 minutes as'inthe previous experiments, and these are disintegrated, centrifuged and examinedfor the detection of diatoms.RESULTSThe diatoms detected in the water of the ditch where the rabbits weredrowned: These proved to be the same 11 kinds as in the previous report.Namely, Cymbella, Navicula, and Nitzshcia, 2 kinds each, Melosira,Cyclotella, Rhopalodia, Diploneis and Tabellaria, one kind each. Of themthe most numerous kinds are Melosira islandica, Navicula placentula, Na-vicula radiosa, Cymbella parva and Cyclotella comta.The rabbit bones (Group "a") that had been buried underground for threeyears demonstrated several kinds of the diatoms that had been detected from theditch water at the time of drowning them. Namely, it was possible to detect 4kinds from the femur and 8 diatoms in the entire field of 5 samples; and thisnumber proved to be greatest. Next, 4 kinds from 5 samples of the humerus tothe total of 7 diatoms, followed by 3 kinds and 4 diatoms, from the rib sam-ples, 2 kinds and 3 diatoms from the vertebra samples, the number and kindsof diatoms decreasing in the order mentioned (Table 1).In the case of the rabbit bones (Group "b"), buried underground for 3 yearsand cremated in the electric oven at 300°C for: 20 minutes, likewise 5 sampleseach revealed 4 kinds and 7 diatoms in the femur; 3 kinds and 5 diatoms in thehumerus; 2 kinds and 2 diatoms in the rib; and 3 kinds and 4 diatoms inthe vertebra (Table 2).In the case of those bones (Group "c"), cremated at 500°C ·in the electric2Acta Medica Okayama, Vol. 15 [1961], Iss. 4, Art. 5http://escholarship.lib.okayama-u.ac.jp/amo/vol15/iss4/5Diagnosis of Death from DrowningTable 1. The number and kinds of diatoms detected from drowned rabbitbones after being buried in the ground for three years (Group "a")~I Femur I Humerus I Rib I VertebraDiatomsCymbella naviculiformis 0 0 0 0Cymbella parva 2 2 1 0Navicula placentula 0 0 0 0Navicula radiosa 1 3 1 2Nitzschia filiformis 0 0 0 0Nitzschia vermicularis 0 0 0 0Melosira islandica 2 1 2 0Cyclotella comta 3 I, 0 1Rhopalodia gibba 0 0 0 0Diploneis elliptica 0 0 0 0Tabellaria fenestrata 0 0 0 0Table 2. The number and kinds of diatoms detected from the drowned rabbitbones after being buried for three years (Group "b")---:-~I Femur 1 Humerus I Rib I Vertebra IDIatomsCymbella naviculiforrnis 0 0 0 0Cymbella parva 2 1 0 0Navicula placentula 0 0 0 0Navicula radiosa 3 2 1 1Nitzschia filiformis 0 0 0 0Nitzschia verrnicularis 0 0 0 0Melosira islandica 1 2 1 2Cyclotella comta 1 0 0 1Rhopalodia gibba 0 0 0 0Diploneis elliptica 0 0 0 0Tabellaria fenestrata 0 0 0 0Table 3. The number and kinds of diatoms detected in the bones of Group "c"~I Femur I Humerus I Rib I VertebraDiatomsCymbella naviculiforrnis 0 0 0 0Cymbella parva 0 1 0 1Navicula placentula 0 0 0 0Navicula radiosa 2 1 0 2Nitzschia filiformis 0 0 0 0Nizschia vermicularis 0 0 0 0Melosira islandica 3 2 3 1Cyclotella comta 1 0 1 0Rhopalodia gibba 0 0 0 0Diploneis elliptica 0 0 0 0Tabellaria fenestrata 0 0 0 02633Okuyama: Experimental studies on diagnosis of death from drowning by meansProduced by The Berkeley Electronic Press, 1961264 M. OKUYAMAoven for 20 minutes, 5 samples each, revealed 3 kinds and 6 diatoms in thefemur, 3 kinds and 4 diatoms in the humerus, 2 kinds and 4 diatoms in the rib,and 3 kinds and 4 diatoms in the vertedra. (Table 3).In the case of the bones (Group "d")·cremated in the electric oven at 800°Cfor 20 minutes, 3 kinds, 6 diatoms in the femur samples; 3 kinds, 5 diatomsin the humerus; 3 kinds, 3 diatoms in the rib; and 2 kinds, 2 diatoms in thevertebra (Table 4).Table 4. The number and kinds of diatoms detected in the bones of Group "d"------___ OrgansI Femur I Humerus I Rib I VertebraDiatoms ~Cymbella naviculiformis)0 0 0 0Cymbella parva 0 0 1 0Navicula placentula 0 0 0 0Navicula radiosa 2 2 1 1I Nitzschia filiformis 0 0 0 0! Nitzschia vermicularis 0 0 0 0Melosira islandica 2 1 1 1Cyclotella comta 2 2 0 0Rhopalodia gibba 0 0 0 0Diploneis elliptica 0 0 0 0Tabellaria fenestrata 0 0 0 0In the case of those bones (Group "e") cremated in the electric oven at1, 000 QC for 20 minutes, 4 kinds and 5 diatoms in the femur samples; 2 kindsand 4 diatoms in the humerus; 2 kinds and 3 diatoms in the rib; and 2 kindsand 3 diatoms in the vertebra (Table 5).Table 5. The number and kinds of diatoms detected in the bones of Group "e"i~~~ ------~l Femur I Humerus I Rib I VertebraI Cymbella naviculiformis 0 0 0 0i Cymbella parva 1 2 0 0Navicula placentula 0 0 0 0Navicula radiosa 2 0 2 0Nitzschia filiformis 0 0 0 0Nitzschia vermicularis 0 0 0 0Melosira islandica 1 2 0 2Cydotella comta 1 I 0 1 1IRhopalodia gibba 0 0 0 0Diploneis elliptica 0I0 0 0Tabellaria fenestrata 0 0 0 0In everyone of these cases the greater number of diatoms can be detectedin the femur followed by that in the humerus. In addition, as the temperature4Acta Medica Okayama, Vol. 15 [1961], Iss. 4, Art. 5http://escholarship.lib.okayama-u.ac.jp/amo/vol15/iss4/5Diagnosis of Death from Drowning 265of the cremation is higher, there is a tendency to show a greater number ofbroken diatoms.COMMENTSIn the case of death from drowning it is well established that diatoms ofthe river water can be detected in the organs of the drowned corpse. However,Mikami and others have contributed considerably for the legal medicine in thediagnosis of the death from drowning, especially where the corpse or remainshad been buried in the ground.In the present experinents it has been possible to detect such diatoms asNavicula placentula, MeLosira islandica, NaaJicula -radiosa, and Cycto*tel/a comta, which are numerically greater among such diatoms as Cynbellanaviculi/ormis, Cymbella parva, Navicula placentula, Navicula radiosa,Nitzschia filiformis, Nitz$chia vermicularis, Melosira islandica, Cye/o-teUa comta, Rhopalodia gibba, Diploneis elliptica and Tabellaria /enut-rata, the same found in the ditch water where the rabbits had been drowned3 years previously.It is assumed that these diatoms have been detected because they are morenumerous and also because they are the kinds that are more resistant to putre-faction. In addition, those six kinds of diatoms, such as Cymbella parva,Cymbella naviculiformis, Navicula placentula, Nitzschia filiformis,Nitzschia vermicularis, Rhopalodia gibba, Diploneis eltiptica and Tabel-laria fenestrata that could not be detected, seem to be the kinds of diatomsthat are not resistant to putrefaction. These data agree with those of MIKAMI,et al. of our department.These diatoms buried underground for three years did endure and could bedetected even after the cremation in the elctric oven at 300°C, 500°C, 800°Cand 1,000°C for 20 minutes, although the number of detectable diatoms is lessthan in the case of the buried but not-cremated bones (Group a). The decreasein the number of detectable diatoms after cremation seems to be due to the dest-ruction of diatoms bv high temperature. Nonetheless, it is noteworthy that somediatoms do remain intact even after the cremation at 1,OOOC° for 20 minutes.The fact that diatoms can be detected more numerously in the femur andhumerus indicates that long bones of the four limbs should be selected for thedetection of diatoms for such purpose in the legal medicine. These results alsosuggest that the detection of diatoms is possible for determining the actual causeof death from drowning so long as there remain some bones, even if a considera-bly long time has elapsed after the burial and even the bones have been exposedto a high temperature. This fact seems to have a great significance in the legal5Okuyama: Experimental studies on diagnosis of death from drowning by meansProduced by The Berkeley Electronic Press, 1961266 M. OKUYAMAmedicine as well as· in criminology for obtaining an evidence to prove deathfrom drowning.CONCLUSIONA series of experiments have been conducted with ten adult rabbits, drow-ning them to death in a ditch those water contains diatoms in abundance. Thebones (selected ones are the femur, humerus, riHand vertebra) of these drownedrabbits have been buried underground, wrapped tightly in cellophane bags andleft there for three years, and the detection of diatoms has been conducted withthese bones either as they are or after cremating them in the electric'oven at300°C, 500°C, 800°C or 1,000°C, for 20 mi~utes.As the results it has been clarified that diatoms can be detected in a con-siderable number in the bones of four limbs, and of these detectable diatomssome of them can be found even after cremation at 1,OOO°C for 20 minutes.This. clearly proves diatoms are detectable from the bones even after a longperiod of time·after burial a~d even after cremation at high temperatures.ACKNOWLEDGEMENTThe author expresses his profound thanks to Prof. Y. Mikami and Ass. Prof. M. Kandafor their kind guidance and encouragement throughout these experiments.REFERENCES1. HOFFMANN,: Ertinken, Eulenburgs Realencyclop~die d. ges. Heilkunde 7, 311, 1895..2.. REVENSTORF, G.: Der Nachweis der aspirierten Ertrinkungsflussigkeit als Kriterium desTodes durch Ertrinken. vjschr. gerichtl. Med. Zl, ·274, 1904.3. BUHTZ, U. BURKHRDT, W.: Die Feststellung des Ertdinkungsortes aus dem Diatome~-Befundder Lungen. Ibid. 29, 469, 1938.4. Kasparek, B.: Beitrage zur Diagnose des Ertrinkungstodes durch den Nachweis von Plank-ton-Organismen in Lunge und Duodenum. Dtsch. Z. gerichtl. Med. 27, 132, 1937.5. Mueller, B.: Studien Uber das Eindringen von corpuscularen Wasserbestandteile aus denLungenalveolen in den Kreislauf wahrend des Eintrinkungsvorganges. Dt:sch. z. gerichtL Med.39, 715, 1949.6. Incze, Gy., Tamaska,L and Gyongyosi, J.: Zur Blutplanktonfrage beim Tod durch Ertrin-ken. Dtsch. Z. gerichtl. Med. 43, 517, 1955.7. Ogawa, H. and Miura, K.: Studies on the microorganisms in water available for the dia-gnosis of drowning. Jap. J. Legal Med. & Criminol. 15, 285, 1941. (in Japanese)8. Ogawa, H. and Miura, K.: Ibid. 15, 475, 1941. (in Japanese)9. Ogawa, H.: Studies on the microorganisms in water available for the diagnosis of drown-ing. Hokkaido J. of Med. Sience. 20, 684, 1942. (in Japanese)10. Tomonaga, T.: An application of incineration method for the detection of planktons inthe inspired water by drowning. Jap. J. Legal Med. Criminol. 14, 293, 1940. (in Japanese)11. Tomonaga, T. and Suyama, H.: On a few cases of medico-legal examination in regard todeciding proof of drowning. Jap. J. Legal Med. 4, 177, 1950 (in Japanese)12. Tomonaga, T.: An application of incineration method for the detection of planktons in6Acta Medica Okayama, Vol. 15 [1961], Iss. 4, Art. 5http://escholarship.lib.okayama-u.ac.jp/amo/vol15/iss4/5Diagnosis of Death from Drowning 267the inspired water by drowning. Ibid. 8, 143, 1954. (in Japanese)13. Tomonaga, T.: A study on the evidence to prove death from drowning by the incinera-tion method. Jap. J. Legal Med. 8 (3), 143. 1954. (in Japanese)14. Suyama, H.: Study on determination of drowning in the sea water. Nagasaki IgakkaiZasshi. 29, 847, 1954. (in Japanese)15. Suyama, H.: Ibid. 30, 493, 1955. (in Japanese)16. Hironaka, M.: Study on diatoms essential to determining drowning as a cause of death.Ibid. 30, 1348, 1955. (in Japanese)1~. Hironaka, M.: Ibid. 30. 1362, 1955. (in Japanese)18. Shinzawa, Y.: Intrusion of water into the lungs of drowned corpse by water pressure.Ibid. 32, 256. 1957. (in Japanese)19. Shinzawa, Y., Furuno, J. and Fukuda, H.: Studies on changes of the vegetative planktonin drowned death body at spring and autumn. Jap. J. Legal Med. 11, 320, 1950, (inJapanese)20. Shinzawa, Y., Furuno, J. and Fukuda, H.: Studies on the number and distribution of dia-toms contained in the drowned and not drowned corpse, especially in lung. Jap. J. Legal, Med. 11, 320, 1957. (in Japanese)21. Mikami, Y., Kanda, M., Kamimura, O. and Okuyama, M.: Experimental study and. practice on the detection of vegetative plankton in the bone marrow of the drowned deadbody. Acta Med. Okayama. 13, 259, 1958.22. Shiragami, K.: The experimental studies on the changes of the vegetative planktons (dia-, toms) in the organs of drowned bodies. Okayama Igakkai Zasshi. 71, 295, 833, 1959. (inJapanese)23. Okuyama, M.: Diagnosis of Death from Drowning. Acta Med. Okayama 15, 250, 19617Okuyama: Experimental studies on diagnosis of death from drowning by meansProduced by The Berkeley Electronic Press, 1961

A study on the evidence to prove death from drowning by the incineration method.

An application of incineration method for the detection of planktons in

An application of incineration method for the detection of planktons in the inspired water by drowning.

Beitrage zur Diagnose des Ertrinkungstodes durch den Nachweis von Plankton-Organismen in Lunge und Duodenum.

Der Nachweis der aspirierten Ertrinkungsflussigkeit als Kriterium des Todes durch Ertrinken.

Diagnosis of Death from Drowning. Acta Med.

Die Feststellung des Ertdinkungsortes aus dem Diatome~-Befund der Lungen.

Eulenburgs Realencyclop~die d. ges.

Experimental study and . practice on the detection of vegetative plankton in the bone marrow of the drowned dead body.

Intrusion of water into the lungs of drowned corpse by water pressure.

On a few cases of medico-legal examination in regard to deciding proof of drowning.

Studien Uber das Eindringen von corpuscularen Wasserbestandteile aus den Lungenalveolen

Studies on changes of the vegetative plankton in drowned death body at spring and autumn.

Studies on the microorganisms in water available for the diagnosis of drowning.

Studies on the number and distribution of diatoms contained in the drowned and not drowned corpse, especially in lung.

Study on determination of drowning in the sea water.

Study on diatoms essential to determining drowning as a cause of death.

The experimental studies on the changes of the vegetative planktons (dia, toms) in the organs of drowned bodies.

Zur Blutplanktonfrage beim Tod durch Ertrinken.

Experimental studies on diagnosis of death from drowning by means of detection of vegetative planktons (diatoms) II. Detection of diatoms from putrefied and cremated bones of drowned bodies

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