The Materials and Methods of Palaeobotany

THE MATERIALS AND METHODS OF PALAEOBOTANY

WILLIAM CULP DARRAH 122 Lincoln Hoad, Medford, Massachusetts

INTRODUCTION impossible, even ridiculous, duplication of names, creation of artificial form genera, and NDER the guidance of the late establishment of purely arbitrary systems of Prof. Birbal Sahni, the plant-bearing nomenclature. U rocks of India were subjected to Consider but one example of this tendency, scrutiny by every technique known to the Palaeozoic spores and pollens, more precisely palaeobotanist. The most unpromising and called palynology. In utter disregard of controversial sediments were investigated parent sporangia, taxonomic group or bio with complete freedom from prejudice logical origin, the multitudinous forms are (SAHNI, 1941, 1946, 1948; SITHOLEY, 1946). binomially identified, classified, and inter In memory of our lamented colleague, I have preted. Gradually we are erecting a colossus chosen as the subject of this contribution, of meaningless terms which will haunt our The Materiats and Methods of Pataeobotany. synonymies for years to come. The prag New techniques - and improvements of old matic or empirical temporary advantage of techniques - have not only wrought a a workable system for coal correlation and revolution in the interpretation of fossil similar purposes is of little long-range im plants but also have provided methods for portance. Existing techniques are sufficient investigating sediments hitherto considered ly refined and perfected to determine within unwor.thy of attention. the space of a few years the biological Some years ago while turning the pages of identities of hundreds, perhaps a thousand, a venerable tome History of Egyptian Mum Palaeozoic spores and pollens. mies by Dr. Thomas Pettigrew (1833), I The changing views of petrifaction merely stumbled upon a discovery made more than reflect a growing realization of the rela a century before, that bituminization may, tive indestructibility of organic materials under certain circumstances, preserve animal ( DARRAH,1941 ). The ex cathedra pronounce tissues indefinitely. The similarity between ment "too delicate for preservation" has embalming with bitumens to coalification or passed unchallenged for a century. No bituminization of plant tissue,; convinced me greater obstacle has impeded exploration of that a vast opportunity to investigate fossil new techniques, new approaches to old plants with greater accuracy than heretofore problems, and pioneering in new problems. merely awaits the improvement of known Dr. Pettigrew in 1833 was examining the techniques or the invention of new ones. heart of an Egyptian mummy estimated to During the past thirty years palaeobotany be 3000 years old. He had removed the has tended to fragment into several special bi tuminous embalming preservatives by isms, each with its own methodology and maceration of the heart in " spirits". 'vVithin tradition. Although this is the inevitable a few days the softened tissues began to result of activity and progress, in such decompose and in a week putrefied. The fragmentation there is a loss of coherence. chemical composition of the macerated heart Taxonomic systems become inordinately arti was such that normal putrefaction occurred ficial, terminologies grow complex and the promptly. On several occasions I have philosophy of the science becomes confused. observed thriving colonies of yeasts and Palaeobotany, however narrowly or inclu molds growing on macerated residues of sively we may wish to define it, is the study bituminous coals but in these cases the of fossil plants. This is the only common polysaccharides and "humic" materials of denominator in our chosen field of research. the coals had been fundamentally altered by Yet we have contributed to the separation of prolonged chemical processing. the specialisms by disregarding the nature In order to repeat Pettigrew's observation of fossils. A proper correlation between I sought the co-operation of Dr. Ernest form and structure would minimize the Hootan and Dr. Donald Scott of the Peabody 1+6 THE PALAEOBOTANIST

Museum of Harvard University. These Briefly then by 1850, the occurrence of the plant gentlemen provided me with tissues of a embedded plant bodies was not only recog original st female Egyptian mummy and dry mummy niz~d,but also fairly generally understood. denied, w~ Arizona Indian, the former being of the 21st It was known also that resistant cutinized long-stand Dynasty ca. 1000 B.C. and the latter ca. parts could be recovered from lignitic and tion by l\> A.D. 1000. Portions of the pterygoid muscle bituminous coals. masses of from the oral cavity of the Egyptian mummy Solms-Laubach (1891, pp. 2-13) distin by demine were macerated by three methods: mineral guished two types of mineral petrifaction, the Indeed, acids and alcohol, benzene and petroleum first in which there is no organic residue ( i.e. deminerali ether. Well-preserved tissues were recovered all apparent structure is pseudomorphic), silicified " by all methods. On the other hand, no and the second, embedded fossils. This excellent recognizable tissues were recovered from the distinction, which today is recognized as an obvious t dry mummy, which incidentally was not over-simplification, epitomized the results of replacemel impregnated with bituminous substances but Goeppert's extensive investigations. wood was rather was completely desiccated. All these chemical methods involve the Goeppert The preserving qualities of bituminization dissolution of inorganic materials and the form of sil are well known. Forinstance, Bn~ggerin pre degradation of organic residues. The degree determine! paring wooden objects found in the Oseberg of chemical alteration resulting from such recognizab Ship used the following technique: wood treatment has long been in dispute and, of Rhynie steamed and soaked with creosote followed by except for certain simple and obvious reac and from a bath in linseed oil in which carbolineum tions, no conclusive evidence has ever been calcareous (an anthracene oil) is dispersed. Soft woods presented. Laboratory experiments on the In this were hardened with alum. This procedure degradation of cellulose and lignin offer little certain cht simulates a common type of fossilization. direct evidence on this question because they yielded all These examples draw attention to the fact do not duplicate natural conditions in which early Pala' that normally destructible tissues have been the organic materials are already degraded. considered preserved by artifactual bituminization and Direct observations on the individual consti Macerat that tissues can be deliberately preserved by tuents of naturally occurring sediments are employed bituminization. extremely meagre. many year For many years the thin ground section to macera MACERATION TECHNIQUES has been respected as the ideal preparation and nitric because, by its very nature, it portrays the sium chlol More than a century ago Goeppert success true condition of the fossil structure. Any vealed the fully removed the petrifying substance of technique which taJ!lpers with the matrix or rials such, silicified and calcified specimens recovering modifies the plant, no matter how little, is and relati" in these experimen ts organic residues of the viewed with suspicion. There is, to be sure, The proce~ original plant bodies. In several cases he a measure of truth in this assumption. has prove( observed the cellulose reaction with sulphuric Certain thin sections, as of the Rhynie chert, The apr acid. Goeppert (1864) performed many the Black Hills cycadeoids and the Permian que to" c( macerating experiments and derived from ferns of Chemnitz, exhibit magnificent pre refinemenl these investigations a better understanding servation. Fungal sporangia, mycelia, sto attempts of the nature of fossils than any other palaeo mata, rhizoids and even cell contents may impressior botanist of the 19th century. As with most be recognized. Yet a silica pe trifaction is COdol1othec prophets, his opinions were generally un oftimes disappointing. Many woods from methods h appreciated. the Triassic of Arizona and the Cretaceous of a large per Other maceration techniques were develop Wyoming preserve nothing of interest, indi which rea ed by several investigators of whom Schultz cating scarcely any features except gross tested for and GUmbel deserve special men tion. form, growth rings, and scattered cell masses. of the bas Schultz in 1855 devised the mixture ·of Such petrifactions are little more than of only a mineral acids (which still bears his name) pseudomorphs. importanc for macerating low rank coals. Subsequent By 1880, despite the work of Goeppert, it on the cal: alkalination was accomplished with sodium was generally assumed that the silicified ( or bryophyte hydroxide. Caustic soda tended to cause calcified) petrifaction was in reality molecular and corda separation of cell masses and, if concentrated, mineral replacement and the structure purely and Arnol dissolved certain constituents. GUmbel mini pseudomorphic. Chemical analyses disclosed oPteris. ' mized this disintegration by gentle alkalina insignificant carbon content, a fact which was b'e recove tion with ethyl alcohol. in terpreted as proof of the disappearance of shown in: DARRAH - THE MATERIALS AND METHODS OF PALAEOBOTANY 147

:urrence of the plant body. The possibility that the TRANSFER TECHNIQUES only recog original structure could be present while not It has been assumed generally that such mderstood. denied, was ignored. The first break in this cutinized or resinoid structures as the exo t cutinized long-standing prejudice was the demonstra spore of spores and pollens, cuticles, and ignitic and tion by Marie Stopes that relatively large heavily lignified xylem elements are so resis masses of original tissue could be recovered tant to hydrolysis and chemical attack by 13) distin by demineralization. mineral and organic acids that their frequence faction, the Indeed, Stopes (1918) had succeeded in and fidelity of preservation are greater than esidue ( i.e. demineralizing completely a specimen of other types of tissue. Although this is )morphic ), silicified wood recovering the tssues in an relatively true, the assumption tends to ssils. This excellent state of preservation. It was obscure the occurrence, indeed the pre lized as an obvious that so-called mineral molecular valence, of more delicate structures. The e results of replacement had not occurred but rather the possibility that a whole mount or " transfer" s. wood was embedded in silica, essentially as of a bleached or cleared structure might pro nvolve the Goeppert had claimed. How prevalent this vide mnch more information than macerated s and the form of silica petrifaction is has not yet been residues was ingeniously explored by Walton The degree determined, but I have been able to recover ( 1923 ). The results exceeded expectations. from such recognizable plant tissues from macerations The Walton technique in basic terms ;pute and, of Rhynie chert, and Triassic silicified woods, involves four steps: vious reac- and from several hundred specimens of ever been calcareous and pyritized coal balls. 1. Affixing the specimen, using balsam as nts on the In this connection it is noteworthy that the adhesive, to a glass slide. offer little certain cherts macerated for microfossils have 2. Coating the exposed glass with wax. cause they yielded algae. The possibilities inherent in 3. Macerating the matrix with mineral IS in which early Palaeozoic sediments have scarcely been acids, usually hydrofluoric, to remove the degraded. considered. associated rock. ual consti Maceration techniques, of course, have been 4. Washing the specimen thoroughly. iments are employed in the investigation of coals for By way of final preparation the specimen many years. Subjection of bituminous coals is dehydrated, covered with balsam, and nd section to maceration by a mixture of hydrochloric protected permanently by a cover glass. >reparation and nitric acids ( to which crystals of potas Thus the specimen is " transferred" from the >rtrays the sium chlorate are sometimes added) has re rock to a glass slide or plate. :ure. Any vealed the presence of many identifiable mate There are two limitations to this method, matrix or rials such as spores, pollens, cuticles, mycelia, the specimen is generally dense and opaque, w little, is and relatively large masses of discrete tissues. and further manipulation is difficult, if not to be sure, The process of coalification, like petrifaction, impossible. The limitation due to opacity ;sumption. has proved to be extremely complicated. may be overcome to some extent by photo ynie chert, The application of this maceration techni graphing with infra-red light (LECLERCQ, e Permian que to " coalified" compressions was a logical 1933 ). ficent pre refinement in technique. One of the earliest The transfer technique was a notable con 1celia, sto attempts to recover pollen from a so-called tribution to the methodology of palaeobotany tents may impression was Sellards' experiment with because it demonstrated, beyond dispute, that 'ifaction is COdol1otheca( 1903). Since that date, similar the so-called carbonized impression was in Dods from methods have been widely used so that today many cases a compression of the original ~taceousof a large percentage of promising fructifications plant body. Of equal significance was the ~rest,indi which reach the hands of researchers are proof that in compression lateral distortion cept gross tested for spores or pollens by some variation is negligible. Thus freed from prejudice ;ell masses. of the basic maceration technique. Mention and preconception it was possible to turn lore than of only a few examples will indicate the to still greater refinements in technique. importance of this procedure: Hartung (1933) In view of the achievements of maceration oeppert, it on the calamarians, Harris (1939) on Triassic and transfer techniques, the recovery of licified (or bryophytes, Florin (1933) on the conifers mummified compressions with an inter , molecular and cordaites, Sahni ( 1923 ) on Glossopteris, mediate transfer process is not a very great ure purely and Arnold ( 1936 ) on the Devonian Archae departure. If the bituminous impregnating :s disclosed opteris. Typical of the structures which can substances were dissolved and the specimen which was be recovered by this method are the spores loosened, the objective could be gained with earance of shown in Figs. 1 and 2. a minimum of manipulation and destruction. 148 THE PALAEOBOTANIST

Various procedures more or less similar these solvents would be outside the scope of any more I have proved to be successful. The following this paper, but mention of three - benzene, the objecti schedule is typical. pyridine and aniline - will suggest the follow the 1. Slowly macerate the specimen in a nature of their action. Successful recoveries The Deel solution of equal parts of nitric acid, hydro have been made in somewhat less than 60 pounded fl chloric acid and water. Maceration should per cent of the attempts, but these reveal a is no univE continue from 3 to 7 days. Some types of wide range of structure: cuticle, epidermis, determines matrix require longer periods and a change palisade and mesophyll parenchyma, xylem with a wic

of fresh solution. elements, ray parenchyma, sporangial cells, prepared a~ 2. Wash the specimen gently in run as well as the expected spores, pollens, etc. 115 gm. ning water for 4 hours, or immerse in water A promising use for maceration may pro 12-13 per c for 6 hours with at least four changes of perly be considered dissection. I have used 1,000 C.c water. it for studying seeds in calcareous and 200 C.c. , 3. At this stage the matrix should be pyritized coal balls. The specimen is immers 10 C.c. tc tested for softness by means of a fme needle. ed in a suitable acid bath in a shallow vessel. 5 c.c. de If it has become soft or somewhat mushy and With the aid of fine sharp needles the This mi the specimen has loosened along the edges, tissues may be peeled off, one by one, either approximal it should be possible to lift the specimen in entirety or in large cell masses un til the dry slowly gradually by prying gently around the peri seed spore is recovered, nearly always com bubbles. ] phery working a little at a time. If, on the plete and unbroken. Fig. 10 shows the seed the additio other hand, the specimen does not yield, spore of Cordaicarpus florini Darrah isolated of the othe further maceration is desirable. by this method. solvent seI' 4. When the acid maceration and rinsing Inasmuch as some mechanical deformation cellulose, a have been accomplished, the specimen is and destruction result from the liberation of thinner fila placed in a 3 per cent solution of sodium gas and the crumbling of attacked crystals, recommend hydroxide or a 5 per cent solution of ammo minimization of these effects is desirable. best possib nium hydroxide. Gradual dissolution of the Some compensation can be gained by dispers and be free " humic" constituents occurs with concom ing gum Arabic in the acid bath. Silicone prepared cc mitant clearing. Experience will indicate oils serve the same purpose. able. If f how long alkalination ~houldbe continued. excellent p Two to ten minutes is usually sufficient, but PEEL TRANSFER TECHNIQUES The proc in some cases 8 hours is not excessive. the followil 5. The specimen is then transferred, Concurrently with refinement of m:lcera 1. Grine manually by forceps or lifter, to a shallow tion techniques there has been developed the surface to 1 watch crystal, washed repeatedly with water so-called peel techniflue. The term" peel" 2. Etch and then covered with a 50 per cent solution is unfortunate because it has become as acid (comm of ethyl alcohol. After 5 minutes in the sociated with the primitive experiments which 3. Wash 50 per cent solution, the specimen is dehy depended upon unstandardized cellulose care not t drated gradually by successive changes of acetates and nitrates which tended to em surface. 70, 85, 90 and 95 per cent and absolute brittle and discolour with age. The basic 4. Air d alcohol. teclmiClue was devised by Koopmans and 5. Covel 6. Following immersion in xylol for one improved by \Valton whose inspiration came surface wit minute, the specimen is placed on a micro from an accidental observation by Nathorst 6. AHo\', scope slide, wetted with a drop of xylol, ( ca. 1908) that a dried film of collodion would 7. Peel j covered with balsam or damar dissolved in peel oft a little of the carbonaceous material The leSt xylol, and permanently sealed with a cover from a compression. Within recent years actually a glass. the method has been highly perfected, parti as 0'5-1 ml Most tissues investigated by this method cularly in the United States ( DARRAH, 1936). thickness ] show surprising mechanical strength and can Two distinct problems are involved in this the embedc be manipulated, with reasonable care, with technique: (1) selective maceration ( " etch lucence of out difficulty. Fig. 5 is a macerated compres ing ") of the surface to be peeled and (2) to that of sion of Neu1'Opteris rarinervis. In this speci compounding of a plastic colloidal mixture quality ent men translucency was increased by a rinse in which upon setting will give an exact transfer mineral anc benzene between steps (5) and (6) described of the structure of the fossil. Etching may To obtain T above. Many organic solvents may be used be accomplished by various mineral acids clarity the to remove selectively the impregnating though for silicified specimens hydrofluoric or damar bituminous compounds. A discussion of is necessary. Since the film cannot transfer glass slide. DARRAH - THE MATERIALS AND METHODS OF PALAEOBOTANY 149

Ie scope of any more detail than is exposed by etching, The possibilities of the peel transfer method - benzene, the objective is to prepare the surface and are suggested by Figs. 7-9,11-14. The degree 19gest the follow the procedure with extreme care. of preservation is faithfully recovered, as the recoveries The oeel solution, so-called, can be com undamaged condition of the archegonium ;s than 60 pounded from various constituents. There ( FIG. 12) and young embryo ( FIG. 13) will ;e reveal a is no universal or ideal mixture, the purpose show. epidermis, determines the most suitable. A mixture It i:; not my intention to enumerate the rna, xylem with a wide range of application may be discoveries made possible by refined tech ngial cells, prepared as follows: niques, but mention of a few examples, lens, etc. 115 gm. nitrocellulose (pyroxylon, grade demonstrate the possibilities of these I may pro- 12-13 per cent nitrogen) methods. The number of instances of com have used 1,000 C.c. butyl acetate pressions which reveal excellent cellular reous and 200 c.c. amyl alcohOl preservation is so great that they are no I is immers 10 C.c. toluol longer reganied as rare curiosities. Perhaps ow vessel. 5 c.c. dehydrated castor oil. the most remarkable discovery of this kind leedles the This mixture, which must be aged for was made by Harris ( 1939) who macerated one, either approximately two weeks before use, will Triassic sediments for specimens of the small s until the dry slowly without the inclusion of air liverwort Naiadita lanceolata (Bruckmann ). ways com bubbles. It may be thinned if desired by This curious plant, known since 1844 by vs the seed the addition of butyl acetate without addition external form, revealed through Harris' ah isolated of the other ingredients. Jnasnmch as the preparations archegonia, perianth parts and solvent serves only as a vehicle for the nitro gemma cups. Obviously, bryophytes, tradi eformation cellulose, a more dilute solution results in a tionally regarded as extremely delicate, afford )eration of thinner film. The highest purity pyroxylon is opportunities for research as great as any d crystals, recommended. The film should have the other group. desirable. best possible chemical and optical properties Selaginellites amesiana (DARRAH, 1938) by dispers and be free from foreign inclusions. Ready a compressed bituminized strobilus from Silicone prepared commercial" dopes" are undepend Mazon Creek, Illinois, exhibited megaspores able. If these precautions are heeded, an with contained gametophytes, some with cell excellent product will be obtained. nuclei and supposed nucleoli (FIG. 8). A QUES The procedure may be indicated briefly by few gametophytes had free nuclei and cell the following schedule: plates (which precede normal cell-wall )f macera 1. Grind and smooth (not polish) the formation). The preparations were nitro 'eloped the surface to be studied. cellulose peels while the preservation was m "peel" 2. Etch the surface with a suitable mineral strictly bituminization. ~come as acid (commonly hydro~loricor hydrofluoric). Animal tissues as well as the more I:esistant lents which 3. Wash gently with running water, using plant structure may also be preserved by cellulose care not to touch or agitate the etched bituminization. We need only to recall the ed to em surface. discovery of a mummified frog (from the The basic 4. Air dry the etched surface. Eocene brown coal of northern Germany) in ,mans and 5. Cover, usually by pouring, the etched which the skin contained expanded chromato ~tioncame surface with nitrocellulose solution. phores (VOIGHT, 1935). I Nathorst 6. Allow film to dry at least 8 hours. dion would 7. Peel film from matrix. MICROTOME METHODS IS material The I esulting film, as noted above, is :ent years actually a thin transfer. Sections as thin Once the presence of recognizable, albeit :ted, parti as O'5-1 micron have been measured. The collapsed, tissues was demonstrated, the AH, 1936 ). thickness refers to the fossil structure, not compression type of preservation assumed ved in this the embedding nitrocellulose. The trans new meaning. If tissues could be recovered n ( " etch lucence of such preparations is far superior by transfer or flotation subsequent to macera ~dand (2) to that of conventional ground sections, a tion, then they could be embedded and sec 11 mixture quality enhanced by the removal of soluble tioned by establiShed microtome technique. ct transfer mineral and opaque insoluble clay particles. Experiment, indeed we may say practice, ching may To obtain maximum permanence and optical has proven this to be true. Two distinct ,eral acids clarity the film should be mounted in balsam embedding methods are practicable: (1) the ydrofluoric or damar on a standard "non-corrosive" celloidin method of Jeffrey ( 1916) originally )t transfer glass slide. devised for sectioning xyloid lignites and 150 THE PALAEOBOTANIST other woody structures, and (2) a paraffin circular disposition. In this case the speci tistry and method devised by Halle ( 1933 ). men had been sawn prior to X-ray photo available e< In either method, it is necessary first to graphing and it was known that the basal mation re, demineralize the specimen completely because region was present. The radiograph con in troductof' the presence of crystallites will damage the firmed the whole structure "in the round" the Eastma cutting edge of the microtome knife and which previously had been reconstructed Various I tear the section. Foreign particles such as from serial sections of three other specimens. chemical, h; quartz or feldspar are particularly detri So far as I am aware there have been no palaeobotar mental. previous published reports of the application These expel Unlike the methods discussed previously, of radiography to fossil plants, consequently purely eXJ= the celloidin and paraffin embedding techni a few of the problems encountered will be 1941 ) 1 ha ques involve rather costly supplementary discussed briefly. A radiograph is simply a thest:' hold. equipment, however, such as is available in shadow picture recorded by a sensitized film. most biological laboratories. The celloidin The object is photographed by X-rays, SUMM method requires a sliding microtome of the differences in density being recorded by J ung or J effrey-Thompson type while the corresponding differences in shadow effects. Each of paraffin procedure utilizes a rotary micro The radiograph, being a shadow formation, brief surve tome. Free-hand sectioning, while possible, is subject to distortion, particularly magni understand! is extremely difficult because of the toughness fication which varies directly as the distance tion and j of some materials and the friability of others. of the object from the sensitized film. The cance, their The materials are manipulated in accordance specimen shown in Fig. 15 measures 15·0 mm. tration of with standard laboratory practice with the in maximum width, the radiograph measures body, in exception of staining which, in most speci 15·4 mm. The distortion is, therefore, 0·4 is an integr mens, would be superfluous. mm. or approximately 2 per cent. The frequently' Microtome sectioning of fossil tissues has clarity or contrast of the image depends Some of not gained general recognition, probably upon many factors of which two are critical: obvious, ot because of the prolonged and complicated radiographic energy and focus. The amount stated bold proceeding involved in demineralizing and of radiation which strikes the object may be 1. The p embedding. However, the remarkable pre expressed by the empirical formula: chemical < parations of Jeffrey, Halle, and others degree resis justify much wider application. Re = Voltage' X Current X Time position. Distance 2 2. Jn err tions in w' X-RAY PHOTOGRAPHY where, Voltage = kilovolts (kV) Current milliamperes The question naturally arises. in what = (mA) Time seconds condition do the plant tissues occur in the = Distance ~ inches or millimetres fossil specimen? Ground thin sections, of course, exhibit the undisturbed structure but The exponent x is a variable which for a because of inherent limitations cannot show given set of conditions may be taken as = 1. thick or large features. During the summer The values most generally satisfactory of 1948 I commenced experiments with for calcified and silicified specimens with ARNOLD, C. thickness varying between 5 and 15 mm. Plants fr, X-ray photography in an attempt to identify America. structure in certain calcified seeds removed were: Bay, Que from coal balls. The objective was to orient 5 : 37-48. X X R =.5 kV 5 mA 6 sec. _ 1.00 DARRAH, W. promising specimens so that in subsequent e 122 in. cutting and peeling, critical structures would botanv. E Idem (1938 not be destroyed or by-passed. Equipment capable of large focus was used Preserved The gametophyte with the archegonium for most of the investigation despite the fact Leafi. Ha shown in Fig. 12 was oriented by this method that good radiographs were obtained with Idem (1941 prior to cutting ( it should be noted, however, fine focus equipment of the type commonly Pan-Amei Idem (1949 that the archegonium itself was not recog used in dentistry. The example shown in bryos. P, nizable in the radiograph). Another radio Fig. 15 was taken with a large focus arrange • ERDTMAN, graph (FIG. 15) of the base of the large ment. Analysis. embryo of Lepidocarpon (Lepidocystis) gla is not necessary to en ter into an elemen FLORIN, R. ( It des Meso brum (DARRAH, 1949), shows the lobed tary discussion of the techniques of radiology. 3rd Ser. construction with strong indication of the The many applications of X-radiation and upturned arms, which appear to have a radiology in industry, medicine, and den- • Not cite, DARRAH - THE MATERIALS AND METHODS OF PALAEOBOTANY 151

e the speci tistry and the diversity of commercially embedded in silica, calcite, or other mineral, -ray photo available equipment have made such infor this body may be recovered by dissolution of t the basal mation readily accessible. An excellent the matrix. The original tissues, somewhat Igraph con introductory guide has been published. by degraded, remain and can be recovered the round" the Eastman-Kodak Company. subjected to microscopic examination and constructed Various other techniques, chieAy micro chemical analysis. - specimens. chemical, have been applied to the study of 3. In bituminized fossils, i.e. ·so-called. lve been no palaeobotanical materials to a slight extent. carbonized impressions, mummifications, application These experiments have been preliminary or compressions, etc., the plant tissues are onsequently purely exploratory. Elsewhere (DARRAH, impregnated with their own decomposition :red will be 1941) 1 have suggested the promise which products. Many coals are composed princi is simply a these hold. pally of compressed plants which, though sitized film. variously altered, are recoverable by macera by X-rays, SUMMARY AND CONCLUSIONS tion techniques. ecorded by 4. Many organic compounds are virtually dow effects. Each of the methods described in this indestructible under conditions favourable for 'I formation, brief survey has contributed to a better fossilization. A number of such compounds arty magni understanding of the conditions of preserva have been positively identified chemically. the distance tion and fossilizatio:1.. Of greater signifi 5. Radiographs of compressions and em I film. The cance, their combined effect has been demons bedded petrifactions show that in many 'es 15·0 mm. tration of the fact that the original plant specimens the tissues are neither crushed nor )h measures body, in varying degrees of degradation, distorted. lerefore, 0·4 is an integral part of many fossils, far more 6. With proper understanding of the cent. The frequently than is commonly believed. nature of a given specimen, and the applica ge depends Some of the following conclusions are tion of suitable technique, considerable are critical: obvious, others are opimon, but they are knowledge hitherto unsought may be attain The amount stated boldly at the risk of pedantry. ed. Many specimens assumed to be worthless ject may be 1. The plant body is constructed of many may contribute significant data towards ula: chemical constituents which in varying the correlation of external form and internal degree resist subaerial oxictation and decom structure. Time position. 7. There are no structures " too delicate 2. Jn embedded fossils, that is petrifac for preservation". Our methods have been tions in which the plant body is literally inadequate, not our materials. : kV) res (mA)

millimetres which for a REFERENCES ken as = 1. satisfactory imens with ARNOLD, C. A. (1936). Observations on Fossil GOEPPERT, H. (1864). Die fossile Flora der per- ld 15 mm. Plants from the Devonian of Eastern North mischen Formation. Palaeontographia. 12, America. I. Plant Remains from Scaumenac Chap. 1. Bay, Quebec. Cont>·ib. jl1us. Pal. Univ. lvlich. HALLE, T. G. (1933). The Structure of Certain 5 : 37-48. Fossil Spore-bearing Organs Believed to Belong = 1·00 DARRAH, W. C. (1936). The Peel Method in Palaeo to Pteridosperms. Kungl. Svensk. Vet. Handl. botany. Bot. jl1us. Leafl. Harv. Univ. 4 : 69-83. 3rd Ser. 12 (6) Idem (1938). A Remarkable Selaginella with HARRIS, T M. ( 1939). Naiadita, A Fossil Bryo us was used Preserved Female Gametophyte. Bot. Mus. phyte with Reproductive Organs. Ann. Bryol. Jite the fact Leafl. Harv. Univ. 6: 113-136. 12: 57-70. tained with Idem (1941). Changing Views of Petrifaction. HARTUNG, W. (1933). Die Sporenverhaltnisse der : commonly Pan-Amel-. Geol. 76 : 13-26. Calamariaceen. Arb. Inst. Palaob. u. Petrog. Idem (1949). Palaeozoic Lepidodendroid Em Brennsteine. 3 : 97-149. e shown in bryos. Palaeobotanical Notices II. JEFFREY, E. C. (1916) Methods of Studyll1g ;us arrange- * ERDTMAN, E. (1936). New Methods in Pollen CoaL Sci. Conspectus. 6: 71-76. Analysis. Svensk. Bot. Tidsk. 30 : 154-164. LECLERCQ, S. (1933). Application de la lumiere ,an elemen FLORIN, R. (1933). Studien Uber Die Cycadales infra-rouge a L'etude microscopique des des Mesozoikums. Kungl. Svensk. Vet. Handl. vegetaux fossiles. A nn. Soc. Geol. Belg. d radiology. 3rd Ser. 12 (5). 56: 351-356 jiation and PETTIGREW, T. ( 1833). History of Egyptian MU'ill • and den- * Not cited in the text . mies. London. 152 THE PALAEOBOTANIST

SAHNI, B. ( 1923). On the structure of the Cuticle SOLMS-LAUBACH, H. (1891). Fossil Botany. 7. Neuropter in Glossopteris angustifolia Brongn. Rec. Geol. Oxford peel of a pinn SUrtl. Ind. 54 (3): 277-280. STOPES, M. C. & WHEELER, R. V. (1918). Mono County, Iowa. Idem (1941). Indian Silicified Plants. 1. Azolla graph on the Constitution of Coal. London. that a longitu intertrappea. P"oc. Ind. Acad. Sci. 14: 489 * THOMAS, H. H. (1912). On Some Methods 8. Selaginell 501. in Palaeobotany. New Phyt. 11: 109-114. with gametopl Idem (1946). Microfossils and the Salt Range VOIGHT, E. ( 1935). Die Erhaltung von Epithel nucleoli. Nitr Thrust. Proc. Nat. Acad. Sci. Ind. 16 (2-4) : zellen mit Zellkernen, von Chromatophoren und strobilus. Car 1-50 Corium in fossiler Froschhaut aus der Mitte X 200. Idem (1948). The Pentoxyleae: A New Group leozanan Braunkohle des Geiseltales. Nov. Act. of ] urassic Gymnosperms from the Raj mahal Leopold. (N.F.) 3 : 339-360. Hills of India. Bot. Gaz. 110 : 47-80. WALTON, ]. (1923). On a new method of investi SELLARDS, E. H (1903). Codonotheca, A New gating fossil plant impressions or incrustations. 9. Cordaicar Type of Spore-bearing Organ from the Coal A nn. Bot. 37 ( 147 ): 379-391. peel, longitud' Measures. Am. Jour. Sci (N.S.) 16: 87-95. Idem (1936). On Factors which Influence External showing micr, SITHOLEY, R. V. (1946). Microfossils from a Form of Fossil Plants. Phil. Trans. Roy. tissues of the Kerogen Shale of the Saline Series in the Soc. Lond. (B) 226: 219-225. Dallas County Khewra Gorge, Salt Range. Proc. Nat. Acad. 10. Cordaica Sci. Ind. 16 ( 2-4) : 220-225. * Not cited in the text. macerated fr< Coal ball, Shul

THE ILLUSTRATIONS

The specimens selected to illustrate this form which, incidentally, has been observed brief summary suggest how in many cases in attachment with Myeloxylon petioles. it is possible to correlate external form with High fidelity peel transfers can be obtained internal structure. Not only can a petrifac with both compressions and petrifactions. tion be subjected to microtechnique, but a A single example of the former category is compression, in favourable instances, can sufficient to indicate the thinness and fidelity be prepared to recover comparable data. attainable, the endosporal gametophyte of Maceration readily yields spores, pollens, Selaginellites amesiana (Darrah) shown cuticles, etc. Figs. 1, 2 and 10 show typical in Fig. 8. Peels of coal balls are innumer recoveries. The important factor is positive able. I have for instance made several identification - known forms are subjected thousands from specimens of Lepidocarpon to maceration. With appropriate processing glabrum ( DARRIIH, 1949) among them many ( strictly speaking, arrested or partial macera gametophytes and embryos. A fine arche tion ) relatively large fragments of leaves or gonium is shown in Fig. 12 and a recently pinnules can be removed from associated discovered young embryo, the smallest yet rock matrix. The two specimens illustrated observed, in Fig. 13. The large embryo in by Figs. 4 and 5 are compressions isolated Fig. 14 has been oescribed previously but is by this method. The pinnule of Neuropteris illustrated here for comparison with the rart"nervis ( Bunb. ) is particularly interesting radiograph shown in Fig. 15. because it compares directly (FIG. 6) with The completeness, with which N eUl'opteris a nitrocellulose peel transfer of a calcified rarinervis and 'Lepidocarpon glabrum are pinnule preserved in a coal ball (FIG. 7). known, by benefit of diverse techniques, Considerable work has been devoted to this presages a rich return for our labours.

EXPLANATION OF PLATES

PLATE 1 entire and have been stripped by moderately deep etching in the surface. X 264. 1. Spores of Ptychocarpus uHitus (Brongn.) 4. Sphenopteris sp. Macerated and bleached macerated from a compression. Mazon Creek, compression. Carboniferous (Conemaugh): Fair Illinois. x l!50. Oaks, Allegheny County, Pennsylvania. X 2. 2. Seed spore macerated from a compression 5. N europteris rarineJ'vis (Bun b. ). Macerated of Cordaicarpus fairchildi White. Henry County, and bleached compression. Carboniferous: Henry Missouri. x 5. County, Missouri. X 2. 3. Pollen of Cordaianthus shuleri Darrah. Coal 6. Same specimen as No.5. Pinnule enlarged ball, Shuler Mine, Dallas County, Iowa. Nitro to show venation and lacunar structure of meso cellulose peel. Note that the pollen grains are phyll. X 22. DARRAH - THE MATERIALS AND METHODS OF PALAEOBOTANY 153

.ssil Botany. 7. Neuropteris rarinervis (Bun b.). Nitrocellulose 11. Lepidocarpon glabrum Darrah. SporangIum, peel of a pinnule. Coal ball, Shuler Mine, Dallas bract, pedicel and abortive spores. Nitrocellulose 1918). Mono County, Iowa. The specimen has been so ground peel. Carboniferous: Shuler Mine, Dallas County, ,I. London. that a longitudinal section was obtained. X 60. Iowa. X 5. lme Methods 8. SelagineUites arnesiana (Darrah). Megaspore 12. Lepidocarpon glabrurn Darrah. Nitrocellulose : 109-114. with gametophyte preserving nuclei and supposed peel showing a portion of the gametophyte with von Epithel nucleoli. Nitrocellulose peel, compression of a an archegonium. Carboniferous: Shuler Mine, ctophoren und strobilus. Carboniferous: Mazon Creek, Illinois. Dallas County, Iowa. x 60. us der Mitte X 200. 13. Lepidocarpon glabrum Darrah. Nitrocellulose es. Nov. Act. peel showing young embryo Carboniferous: PLATE 2 Shuler Mine, Dallas County, Iowa. X 60. lod of investi 14. Lepidocarpon glabrum Darrah. Nitrocellulose incrustations. 9. C01'daicarpus Fiorini Darrah. Nitrocellulose peel. .. Mature" embryo with lobed base. Vas peel, longitudinal section through apex of seed cuLarization is evident. Carboniferous: Shuler ence External showing micropyle. summit of megaspore, and Mine, Dallas County, Iowa. x 5. Trans. Roy. tissues of the pericarp. Coal ball, Shuler Mine, 15. Lepidocarpon glabrurn Darrah. Radiograph Dallas County, Iowa. x 60. of the specimen shown in Fig. 14. Note the 10. Cordaicarpus Fiorini Darrah. Seed spore more dense area, the base, with strong indication macerated from a mechanically isolated seed. of the lobes. Approximately natural size (see Coal ball, Shuler Mine, Dallas County, Iowa. X 22. text for explanation of linear distortion).

:n observed 'etioles. be obtained etrifactions. category is and fidelity ~tophyteof h) shown e innumer Ide several ~pidocarpon them many fine arche a recently mallest yet embryo in 'usly but is with the rveuropten's abrmn are :echniques, tbours.

moderately ld bleached lUgh): Fair a. X 2. Macerated rous: Henry ule enlarged He of meso-

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