CHLOROPHYLL-CORPUSCLE AMYLOI DEPOSITSD S ......CHLOROPHYLL-CORPUSCLES AND AMYLOI DEPOSITSD 22. 9 On the CHLOROPHYLL-CORPUSCLE and AMYLOI DEPOSITSD S of SPONGILL and HYDRAA By. E. LANKESTER

CHLOROPHYLL-CORPUSCLES AND AMYLOID DEPOSITS. 2 2 9

On the CHLOROPHYLL-CORPUSCLES and AMYLOID DEPOSITSof SPONGILLA and HYDRA. By E. RAY LANKESTER,M.A., F.R.S., Jodrell Professor of Zoology in UniversityCollege, London. (With Plate XX.)

QUITE recently (November, 1881) Dr. Karl Brandt1 hasadduced reasons for considering the green-coloured corpuscleswhich occur in the cells of Spongilla Jluviatilis and of Hydraviridis as parasitic Algae, and has given to those occurring inthe former the name " Zoochlorella parasitica," to thoseoccurring in the latter the name " Zoochlorella conductrix."

Professor Semper, of "Wurzburg, had previously expresseda similar opinion as to the nature of these green-colouredcorpuscles. In his remarkable volume, ' Animal Life"(International Scientific Series, p. 73), Semper brings for-ward facts which he thinks " may soon require us, if wefind true chlorophyll in animal tissues, to recognise in itspresence a singular and interesting case, either of parasiticismor of the community of two organisms so different, as ananimal with true tissues and organs and a one-celled plant."Semper is led to this view by the analogy of Lichens, theduplex nature of which was demonstrated by Schwendener.He is further influenced by and cites Cienkowsky's obser-vations and conclusions as to the "yellow corpuscles" ofRadiolarians. But the value of Semper's judgment in thematter is very much diminished by the fact that he expresslydeclares that the chlorophyll bodies of plants do not divideand multiply (p. 74). Upon this erroneous conception ofthe chlorophyll bodies of plants Semper bases his argumentas to the probability of the green corpuscles found in animalsbeing independent Algae, since in some cases these greencorpuscles have been observed to multiply by transversedivision. It is, however, well known that the chlorophyllbodies of green plants also multiply by transverse division.

The probability in favour of the independent algal natureof the green corpuscles of Spongilla and Hydra was largelyincreased between the publication of Professor Semper'sconclusions and the memoir of Dr. Karl Brandt by theobservations of the Hertwigs on the yellow corpuscles ofRadiolarians and on yellow corpuscles, which occur abun-dantly in the endoderm of Anthozoa (Actinise). The Hert-

1 ' SiUungsbericlit der Gesellsch. Naturforscli. Freunde zu Berliu,'No. 9, 1881.

2 3 0 PROFESSOR E. RAY LANKESTER.

wigs confirmed the observations of Cienkowsky as to theindependent nature of the Radiolarian yellow corpucles,and held them to be parasitic. They also demonstrated thatthe yellow corpuscles of Anthozoa had the structure andproperties of unicellular Algse, and were inclined to regardthem also as parasitic.

The term " symbiosis/' introduced I believe originally toexpress the relation of the green algal gonidia of Lichensand the associated colourless Inophyte, has been recentlyextended with great effect to include the relationship of theRadiolarians to their yellow cells, and also of Anthozoa totheir yellow cells.

Whilst there appears to be very nearly sufficient groundfor accepting the conclusion thus formulated in regard tothe Radiolarians and the Anthozoa, I shall endeavour toshow in the following pages that there is no evidence tojustify the extension of the doctrine of symbiosis to Spon-gilla and Hydra, as advocated by Professor Semper and Dr.Brandt. It appears to me, on the contrary, that an examina-tion of the green-coloured corpuscles of Spongilla andHydra demonstrates those corpuscles to be similar in natureto the " chlorophyll bodies " of green plants, and that thereis no more reason to regard them as symbiotic Algae thanthere is to regard the green corpuscles in the leaf of a butter-cup as such. The results of my examination of the greencorpuscles of Spongilla and Hydra, which have been madeat intervals during the past seven years, are given below.I have already in this Journal (vol. xiv, p. 400, 1874) pub-lished a note relative to the chlorophyll-corpuscles ofSpongilla. It is, perhaps, worth pointing out, that in advo-cating the view that what I venture to consider as a normalproduct of the living activity of animal cells is in reality aparasite, Dr. Brandt is exactly in the converse position tothat occupied by Dr. Gaule, of Leipzig, whose opinion thatthe parasitic Drepanidium phase of a Gregarina is a normalproduct of the living activity of the blood-corpuscles andother cells of the Prog, I controverted in the last number ofthis Journal.

Nature of chlorophyll.—The green colouring matterwhich gives its characteristic tint to foliage is not a purechemical substance, but a somewhat variable mixture ofcoloured substances, none of which have yet been properlyisolated and characterised by the chemist. Hence there isno little difficulty encountered when we attempt to definitelyand satisfactorily identify the green-coloured substanceswhich appear in lower plants and in some animals with the

CHLOROPHYLL-CORPUSCLES AND AMYLOI0 DEPOSITS. 2 3 1

green-coloured substance which tints the " chlorophyllbodies " of higher plants.

The introduction of the spectroscope, as a means of iden-tifying coloured substances in solution, has resulted in thediscovery of the complex nature of " chlorophyll." Start-ing from the important observations of Stokes, followed bythose of Kraus and other continental observers, we come tothe latest and most complete investigation of " chlorophyll,"which is that of Mr. Sorby.

Sorby had the advantage of having made an extensivestudy of vegetable and other colouring matters by the aid ofhis beautiful micro-spectroscope and the systematic use ofchemical reagents. His method of investigation is primarilybased upon that of Stokes. He distinguishes, in relation tothe green leaves of the higher plants, colouring matterssoluble in water (erythrophyll and chrysotannin group)and those soluble in absolute alcohol and often in carbonbisulphide (chlorophyll group, xanthophyll group, andlichnoxanthine group). The green substances are thoseknown as the " chlorophyll group/' and occur in the so-called chlorophyll bodies in association with the yellowsubstances of the xanthophyll and lichnoxanthine groups.The chlorophylls are distinguished as blue and yellowchlorophyll, found in higher plants, and chlorofucine, whichoccurs in association with the two preceding, in certain Algae.Blue chlorophyll is separated from yellow chlorophyll by itsgreater solubility in certain media. The absorption spectraof these and of chlorofucine present bands differing in posi-tion ; they are in each case precisely fixed and representedin a diagram by Sorby, as are the absorption bands of eachspecies of the xanthophyll and lichnoxanthine groups.Upon all these substances acids and alkalies have certaindefinite effects, in some of them profoundly modifying theabsorption spectrum, and clearly giving rise to new chemi-cal compounds; whilst in other cases a slight modificationonly is produced by such reagents. According to Sorby,various observers previous to himself had, in studying theabsorption spectrum of the alcoholic extract of a green leaf,considered the seven-banded spectrum due to a mixture ofblue and yellow chlorophyll, together with the products ofthe action of acids upon those substances—as characterisingone simple substance1—the so-called " chlorophyll." Accord-ing to Sorby, such a mixture occurs ordinarily in alcoholwhen allowed to act upon a green leaf. Various analyticalprocesses must be used to separate the various bodies thusoccurring in company, and precautions adopted to prevent

# 3 2 PROFESSOR E. RAY LANKESTJER.

the modifying effects of acids in solution. The deep absorp-tion band in the dark red, which is seen even in weak solu-tions of green colouring matters spoken of by other observersas " chlorophyll " or '•' chlorophylloid substance," is, accord-ing to Sorby, present in a slightly varied position in the caseof blue chlorophyll, in that of yellow chlorophyll, and inthat of chlorofucine, and also in a totally distinct substancecalled Bonelleine. Hence no very precise conclusion can bebased upon the occurrence alone of this strong absorptionband in the red (as, for instance, had been done by myselfin the case of Hydra, Bonellia, Idotea, and Chsetopherus).^

Moreover, the fine red fluorescence which is seen in analcoholic solution of leaf green is exhibited, according toSorby, equally by blue and yellow chlorophyll and bychlorofucine, and, as we now know, by Bonnelleine and byMoseley's Pentacrinin, the red rays having, however, in eachcase a different and characteristic refrangibility.

Thus, it appears that the investigation of the claims ofany given greenish coloured pigment to be regarded as" chlorophyll" is by no means a simple matter. Supposingthe pigment to be soluble in alcohol, we still have to ascer-tain which of Sorby's three groups (chlorophylls, xantho-phylls, lichnoxanthines) are present, and which of each ofthe species distinguished by him within those groups.

In order to do this we have to rely on :1st. Variations in degree of solubility in such media as

alcohol, ether, benzine, carbon bisulphide.2nd. Absorption spectra of the series of solutions obtained.3rd. Fluorescence and spectrum of the fluorescent light of

such solutions.4th. Reactions of the solutions with acids, alkalies, and

oxidising and reducing agents, which give rise to new com-pounds or change the spectra characteristically.

There are, however, yet to be mentioned two categories ofphenomena in relation to the chlorophyll bodies of greenplants which comprise data of a nature to assist us injudging of the similarity or dissimilarity of the green pig-ments of animals compared with that of the chlorophyllbodies. These are, firstly, the physiological activities asso-ciated with the chlorophyll bodies of plants ; and, secondly,the morphological features of these bodies.

1 It is, however, very probable that blue chlorophyll and yellowchlorophyll, as well as the xanfchophylls and lichnoxanthines, are successivephysiological stages of metamorphosis of one (colourless ?) original mother-substance, since they appear always to be found associated with oneanother.

CHLOROPHYLI CORPUSCLES AND AMYLOID DEPOSITS. 2 3 3

If we find in an organism physiological processes asso-ciated with the presence of a green pigment, which processesare identical with those associated with the presence of thegreen pigment occurring in the chlorophyll bodies of plants,we have so far a certain amount of evidence in favour of theidentity of the green pigment in the two cases. And,again, if we find that the green pigment in an organismoccurs in corpuscles which are morphologically similar tothe chlorophyll bodies of plants, we have so far evidence infavour of the identity of the green pigment in the two cases.

This evidence, both as regards the physiological and themorphological phenomena, will acquire weight in proportionas those phenomena are elaborate or of a peculiar nature.

It has long been known that whatever may be the partactually taken by the green pigment of the chlorophyllbodies of plants in the process, the presence of this greenpigment is constantly associated with the decomposition ofCO2 by the green parts of plants in sunlight, accompaniedby the liberation of oxygen gas, and frequently with theformation in the green-coloured chlorophyll-corpuscles ofstarch. And conversely, it is true that these chemical pro-cesses are not known to occur except in the presence of thepeculiar complex of coloured substances which give to thechlorophyll bodies their green colour. The recent discus-sion by Pringsheim of the actual part played by the greenpigment in this physiological phenomenon is reported on atsome length in another part of this Journal. It is sufficientfor our present purpose that the process is constantly andexclusively associated with the presence of the green-coloured chlorophyll bodies. If this very remarkable andpeculiar physiological process can be shown to occur in thegreen-coloured parts of other organisms, we have groundfor supposing the green-coloured parts to be of the samenature as the chlorophyll bodies of plants. This line ofevidence we may therefore add to the four already estab-lished by Sorby, and distinguish it as the physiologicalevidence.

Similarly, the so-called chlorophyll of the chlorophyllbodies.of green plants has been shown to occur very usuallyas a pigment impregnating certain definitely shaped cor-puscles which are lodged in and form part of the proto-plasm of the vegetable cell. With exceptions amongst thelower Algee, these corpuscles are spherical bodies, which aredistinguished from the protoplasm in which they lie bytheir differing density. They appear to consist of albu-minoids and are not diminished in bulk when the green

2 3 4 PROFESSOR E. RAY LANKESTEIt,

pigment is completely dissolved away from them so as toleave them colourless. They have not the nature of " cellswithin cells," for they possess nothing within them compar-able to a cell-nucleus. Rather they have been comparedeach to a nucleus. They are devoid of special wall orcapsule, and have been observed in a state of transversedivision, from which it is inferred that they may multiply byfusion, just as nuclei may multiply in multinuclear cells. Asto the internal structure of the colourless basis of a vegetablechlorophyll-corpuscle, there appears to be less known thanmight be anticipated. The central portion appears to bethe seat of the formation of starch in certain cases, and thisimplies a differentiation of superficial and central substance.The green pigment is sometimes observed to impregnateonly the superficial layer of the corpuscle so as to form akind of green-coloured shell enclosing colourless contents(starch, &c). This is important for the comparison withthe supposed chlorophyll-corpuscles of animal tissues.

The development of the chlorophyll bodies of plants andthe effect upon them of physical agents is at present receiv-ing renewed attention. It has been observed that they arepreceded by colourless corpuscles which under the influenceof sunlight develop the green pigment in themselves.And it has also been observed that in certain saprophytesallied to green plants, but themselves devoid of green pig-ment, colourless corpuscles are present which appear torepresent the chlorophyll bodies, and these acquire a greencolour when acted upon by chemical agents (sulphuricacid).

If we find green-coloured corpuscles present in the cellsof an organism which exhibit the form and structure justdetailed, we have evidence so far in favour of such corpus-cles being of a nature identical with that of the chlorophyllbodies of plants.

This line of evidence in relation to the inquiry into thesupposed existence of chlorophyll in animals we may addas a sixth to those already enumerated, calling it the mor-phological evidence.

It may be added here that over and above the broad phy-siological and morphological characters of the chlorophyllbodies of green plants which have been above cited, thereare other morphological and physiological characters lessimportant or less familiar, which may of course be made useof in a comparison bet%veen these bodies and similar bodiesoccurring in animals.

Definite Droof of the occurrence of chlorophyll in an

CHLOROPHYLL-CORPUSCLES AND AMYLOID DEPOSITS. 2 3 5

animal.—There is at present only one example of an animalin which the frequent presence of that complex associationof pigmented bodies termed " chlorophyll " has been fullydemonstrated.

This animal is the freshwater sponge (Spongilla fluvia-tilis), which often exhibits a brilliant green colouration of itssubstance, though colourless or pale flesh-coloured growthsof it are very abundant under some circumstances.

Before Sorby had made his investigations on chlorophyll,I endeavoured to confirm the opinion which was current asto the probable identity of the green pigment of some loweranimals with the " chlorophyll" of plants, by the applicationof Sorby's micro-spectroscope and a comparison of the bandsof absorption afforded by the pigments in question withthose shown by a solution of leaf green in alcohol.

I obtained in the case of Hydra and in the CrustaceanIdotea a dark band identical with that of some solutions ofleaf green. In the case of Spongilla, I was able to procurelarger quantities of the colouring matter, and also in thecase of Chgetopterus (from the epithelium of the dark lobesof the alimentary canal) and in that of Bonellia. I was ledto the conclusion that the green colour of Spongilla wasallied to but not identical with " chlorophyll," and that thedark green pigment in Chsetopterus and in Bonellia wasactually " chlorophyll."

My results were unsatisfactory, owing to the fact that inthe first set of cases (Hydra, &c.) it was not possible toobtain a sufficient quantity of the colouring matter tooperate upon; and in the second set of cases my resultswere equally faulty, owing to the fact that there was not aproper standard of comparison in the knowledge which thenexisted as to leaf green or vegetable chlorophyll itself.

As soon as Mr. Sorby's researches on leaf green werepublished, I persuaded him to undertake the investigationof the two reputed cases of occurrence of chlorophyll inanimals in which I could promise him a sufficient supplyof material. These were Spongilla Jluviatilis and Bonelliaviridis. I supplied Mr. Sorby with a quantity of green-coloured Spongilla, and with a strong solution in alcohol ofthe green pigment of Bonellia. Mr. Sorby's researches onthe green pigments of these two animals were published inthis journal.(vol. xv, p. 47 and p. 166), and constitute theonly adequate investigation of reputed chlorophyll substancesfrom animals.

They resulted in the complete demonstration of theidentity of the green pigment present in Spongilla with that

2 3 6 PJIOFESSOR E, HAY LANttESTEH.

occurring in the chlorophyll bodies of normal green plants,•whilst what had been taken by Schenk as well as by myselffor chlorophyll in Bonellia, was shown to be distinct from anyof the pigments of the chlorophyll group, although muchresembling in its seven-banded absorption spectrum and itsred fluorescence certain combinations of that group.

To the pigment of Bonellia Mr. Sorby gave the nameBonelleine. The physiological evidence of correspondencebetween this substance and the green pigment of plants hasrecently been sought for by Mr. Geddes with a negativeresult. He has found that Bonellia does not in the livingstate when exposed to sunlight decompose carbonic acid withliberation of oxygen gas.

Mr. Sorby established the identity of the green pigmentof Spongilla with that of the chlorophyll bodies of higherplants by obtaining from it and identifying with the spec-troscope the following substances soluble in carbon bisul-phide, viz. blue chlorophyll, yellow chlorophyll, orangexanthophyll, xanthophyll, yellow xanthophyll, and lichno-xanthine. He also obtained a small quantity of a yellowsubstance soluble in water, differing from the chrysophyll ofhigher plants. The substances found in Spongilla solublein carbon bisulphide are precisely those which are presentin the ordinary chlorophyll bodies of green plants, in thesame association, but their proportions are a little differentin the two cases, according to Mr. Sorby.

No physiological evidence as to the nature of the greensubstance present in Spongilla has been brought forward byany observer, and I have not made any experiments in thatdirection. It is still not known whether the green parts ofSpongilla decompose carbonic acid and liberate oxygen inthe presence of sunlight.

The morphological evidence relating to it has also untilquite recently been neglected. An account of my observa-tions in this respect will be found below.

Thus, then, Spongilla remains the only animal in whichthe presence of a green pigment identical with that of thechlorophyll-bodies of plants has been definitely establishedby chemical and spectroscopical investigation. The fullcorroboration of the identity by physiological and morpho-logical evidence is still wanting.

Pigment of Clisetopterus and Pentacrinus.—It would beeasy to obtain the chlorophyll-like pigment of Chsetopterusin large quantity, and I anticipate that it would be found tobe similar to if not identical with Bonelleine.

In this connection I would draw attention to the interest-

CHLOROPHYLL-CORPUSCLES AND AMYLOID DEPOSITS. 2 8 7

ing green pigment (purple when acidified) obtained by Mr.Moseley in great quantity from species of Pentacrinusdredged by H.M.S. " Challenger." The banded absorptionspectrum of alcoholic solutions of this substance is figuredby Mr. Moseley in vol. xvii of this Journal, in a memoircontaining a great number of very important observations,which appear to have been overlooked by those who haverecently been studying the characters of animal pigments.

Chlorophylloid pigment of Hydra viridis.—As to thechlorophyll-like substance of Hydra viridis, it is to be ob-served that there is very great difficulty in obtaining sucha quantity of it as will suffice for complete spectroscopicstudy with reagents such as Sorby has carried out in thecase of Spongiila. Hence the general indication of chloro-phyll-like substances afforded by the single strong absorptionband in the red, acquires in this case importance. Still moreimportant is the physiological and morphological evidencewith regard to it which will be detailed below, and sufficesto render the identity of the pigment of the green corpusclesof Hydra with that of the chlorophyll bodies of plants highlyprobable.

Physiological evidence as to the occurrence of chloro-phyll in animals.—At present the attempt to prove theidentity of a green pigment occurring in an animal withthat of the chlorophyll bodies of plants, by proving thegreen tissues of the animal to be capable of decomposingCO2 in sunlight with liberation of oxygen gas, as the greentissues of plants are known to do, has only been made withsuccess in one case.

Mr. Geddes has shown that a green marine Planarianworm {Convoluta Schultzii), when exposed to sunlight inwater containing CO2 in solution, evolves an appreciableamount of oxygen gas. Supposing that the green pigmentof the chlorophyll bodies were itself the agent of the decom-position which is associated with them, this observationwould be tolerably conclusive as to the presence of the samegreen pigment in Convoluta Schullzii. But if Pringsheim'stheory that the chlorophyll acts only as a "screen" be true,and if the decomposition is actually effected by the proto-plasm of the vegetable cell, then it seems not very unlikelythat one green pigment might act as a screen as effectuallyas another, and the physiological evidence as to the identityof the two pigments would have little value. Some valuemust, however, be ascribed to it, for it is, on the whole, moreprobable—even if we accept the screen theory—that onlysubstances belonging to Sorby's groups of chlorophylls,

3 3 8 PROFESSOR E. RAY LANKESTER.

xanthophylls and lichnoxanthines can effectually act asscreens than that other green pigments of a different naturecan play that part.

Mr. Geddes has not adduced spectroscopic evidence simi-lar to that given by Sorby in relation to Spongilla, in favourof the view that the green pigment of Convoluta Schultzii iscaused by a mixture of blue and yellow chlorophyll, xantho-phyll and lichnoxanthine, or by any one or two of thesebodies; nor has the precise structural form in which thepigment occurs in Convoluta Schuttzii been described, so thatwe can compare it with the chlorophyll bodies of plants.

Physiological evidence in favour of the assimilation of thegreen pigment of Hydra viridis to that of green plantswas obtained two years ago by Mr. J. E. Blomfield, ofMagdalen College, Oxford, and University College, London.He has kindly supplied me with the following account ofthe experiments made by him :

" A number of Hydra viridis (some forty or fifty) wereplaced in a test-tube, and the test-tube filled with water wasinverted over water in a basin, the bottom of which waslined with clay so that the test-tube could be pressed into it{not so as to completely close the tube) and maintained inan upright position, and the Hydrse could either crawl upthe tube or remain resting upon the clay. Any gas givenoff from the Hydrse would necessarily ascend in the tubeand collect at the top of it, expelling a corresponding volumeof water from the inverted mouth of the tube. On exposingseveral tubes thus prepared to direct sunlight minute bubblesappeared on the Hydrae and on the side of the tube, whichascending formed a large bubble at the top of the latter.

" A test-tube containing nothing but water will, whenexposed to direct sunlight, be found to have its sides dottedwith minute bubbles, which collect into a larger bubble onrising to the top of the column of liquid, but the gas soseparated is reabsorbed iohen the temperature of the waterfalls from the point which it has reached by the heatingeffect of insolation.

"Several test-tubes containing Hydrse wereset up in thisway and left for a week, but owing to the unfortunatelysmall amount of direct sunlight at the time, only a smallbubble of gas was obtained at the top of each tube. Thebubbles so obtained were passed from the collecting tubesinto a smaller test-tube for analysis. The small test-tubecontaining some water and the collected gas was invertedover mercury, and a solution of pyrogallic acid in KHO wasintroduced into the tube. One third of the total volume of

CHLOROPHYLL-CORPUSCLES AND AMYLOID DEPOSITS. 2 8 9

gas was absorbed, and it was accordingly inferred that onethird part by volume of the gas evolved from the Hydrsewas oxygen gas."

Mr. Blomfield further observed that the green pigmentof the Hydrse was soluble in alcohol, and that the greensolution obtained in that medium was converted in thecourse of half an hour's exposure to direct sunlight into afaint yellow colour. Unfortunately nothing further wasobserved with the spectroscope than the presence of oneintense absorption band in the red similar to that affordedby solutions of leaf green.

Morphological evidence with regard to the occurrence ofchlorophyll in animals.

A. SPONGILLA.

Chlorophyll-corpuscles.—I now come to the statement ofmy own observations with reference to the form under whichthe green pigment of Spongilla presents itself, and I am ofopinion that these observations lead to the conclusion thatthe green pigment there present, which Sorby has demon-strated to agree in spectroscopic characters of a detailedkind with the green pigment of higher plants, is containedin " chlorophyll-corpuscles " or " chlorophyll bodies " whichhave the same nature as the bodies so designated in thetissues of green plants, and that these corpuscles are, as ingreen plants, formed by the activity of the protoplasm of thecells in which they occur.

In figs. 1 and 3, and others on Plate XX, amoeboid cellsfrom green-coloured specimens of Spongilla fluoiatilis arerepresented, showing the green-coloured corpuscles embeddedin their substance, which I consider as chlorophyll-corpusclesproper to these cells. The corpuscles are concavo-convexdiscs, averaging Tuys-oirth t° TT.-s's-oth inch in diameter. Theyare of a uniform green colour, and are often so abundant asto occupy a large bulk of the cell. Some cells, however,are observed in which they are much less abundant.

Rarely I have observed in the amoeboid cells containingchlorophyll-corpuscles of normal size and shape one chloro-phyll-corpuscle abnormally large and differing in shapefrom those usually seen. In fig. 2 ccc such a corpuscle ofspherical form is drawn. In fig. 12 a similarly large chloro-phyll-corpuscle from Spongilla is drawn. In this case thegreen pigment is confined to a superficial layer or shellinvesting a colourless substance and to a few grains within.

The amoeboid cells and the corpuscles containing themmay be well observed by simply teazing a piece of a living

2 4 0 PROFESSOR E. KAY LANKESTEll.

specimen of green-colouved Spongilla. But a valuablemethod is that of teazing the piece of Spongilla in a drop ofdilute solution of osmic acid (£ per cent.). Such prepara-tions may be subsequently stained by picro-carmine, asshown in PI. XX, fig. 13. When this treatment is adoptedit is noted that though the nucleus of the sponge-cell stainsvery intensely no staining of the chlorophyll-corpuscle, or ofthe protoplasm close to it, occurs.

There is no evidence of any nucleus-like body, eitherwithin the chlorophyll-corpuscle or in immediate relationwith it.

When a piece of green Spongilla is decolorised by theaction of strong alcohol and subsequently teazed the concavo-convex discs, which were before observed in a green state,are still to be found, but now they are colourless.1

When the amoeboid cells of Spongilla containing chlo-phyll-corpuscles are broken up on the stage of the micro-scope the corpuscles are set free, and are found to have aconsiderable degree of resistance to the action of water anda permanence of form, as is observed with the chlorophyll-corpuscles of plants.

Under such treatment it is further observed that the greenconcavo-convex discs do not separate cleanly from the pro-toplasm, but each carries with it a little sphere of protoplasm,upon which it rests as a cap (figs. 9 d, 10 b). The relation ofthe green cap to the sphere is such as to suggest a one-sidedformation of green matter upon one hemisphere of the pro-toplasmic particle. Were the formation of green mattersymmetrical we should have the protoplasmic sphere enclosedin a complete shell of green substance, as in the abnormalcorpuscle of fig. 1SJ, and as in the normal green corpuscles ofHydra viridis (figs. 17, 20).

I could not discover in the unbroken amoeboid cell thatthere was any differentiation of a protoplasmic sphere cor-responding to each chlorophyll-corpuscle. It would seemrather as though a piece of the surrounding protoplasmsimply adheres to the concavo-convex disc when the cell isbroken up. At the same time pieces of protoplasm may beobserved when such cells have been broken up which con-nect two or more (as many as six) concavo-convex discs ofgreen colour (fig. 10«), suggesting that the cap-like chloro-phyll-corpuscles are grouped around a centre of growth, and

1 Accordingly we distinguish in the concave chlorophyll-corpuscle ofSpongilla—(1) the chlorophyll itself, (2) the chromophorous substancewhich carries the chlorophyll. In Hydra, as will be seen subsequently, wehave to add to these two elements (3) colourless protoplasm, enclosed bythe chromophorous substance,

CHLOROPHYLL-CORPUSCLES AND AM VLOID DEPOSITS. 2 4 1

that they have originated by a process of cleavage of anoriginal layer of green substance which invested the particleof protoplasm. Evidence of the cleaving of the chlorophyll-corpuscles, so as to form two corpuscles from one, is given inthe accompanying drawings (fig. 9 c).

I am not acquainted with any chlorophyll bodies of plantswhich assume the form of concavo-convex discs as do thoseof Spongilla. At the same time there is nothing incon-sistent with what is known of chlorophyll bodies in this form,whilst in their simple negative characters the green cor-puscles of Spongilla are like the chlorophyll bodies ofhigher plants. They are quite unlike any known forms ofunicellular Algae.

Amyloid substance in Spongilla.—Neither before treatment•with alcohol nor after it did the addition of iodine solutionto the sponge-cells reveal any substance within the cor-puscles, which by its blue or violet coloration could suggestthe presence of starch.

I, however, obtained in both green and colourless speci-mens of Spongilla treated in this way with iodine solutionabundant evidence of the presence in other regions of thesponge-cell of an amyloid substance. My observationswere made on specimens taken late in the year (October),and I am inclined to believe, from my recollection of formerexperiments, that the amyloid substance is not so abundantin the early part of the year as in autumn.

The amyloid substance occurred in two forms—(1) as ahomogeneous substance occupying very large vacuoles—usually only one—in the protoplasm of the sponge-cell (PI.XX, figs. 3, 4), and (2) as fine spherical granules, which•were accumulated on the surface of some of the sponge-cells, and embedded in the superficial layer of the protoplasm(PI. XX, figs. 4, 5, 8).

The amyloid vacuoles of Spongilla and of other spongeswere discovered and described by Keller (• Zeitschr. furwiss. Zoologie,' vol. xxx, p. 572). Keller points out thatthe vacuoles contain a fluid which stains deep blue or violetwhen iodine solution is added (in my observations I obtainedonly violet staining), and that the substauce so coloured isinsoluble, either in ordinary or absolute alcohol or in coldwater, whilst potash solution decolorises the stained vacuoleand causes the cell to swell up.

Keller's observations undoubtedly prove that we have inthese vacuoles a starch-like substance in solution, but it isby no means to be concluded that this substance is identicalwith vegetable starch.

VOL. XXII. NEW SEfi. Q

2 4 2 PROFESSOR E. RAY LANKESTER.

An addition to Keller's observations on these "amyloiddeposits " which I have to record is that the vacuole whichstains violet with iodine is also deeply stained by a solutionof picro-carmine after previous treatment with dilate osmicacid (PI. XX, fig. 14).

This carmine staining would lead to the inference that analbuminoid as well as an amyloid substance is present inthe fluid vacuole.

Keller appears not to have seen the small granules ofamyloid substance which I observed in great abundanceboth in the superficial protoplasm of cells which containedan amyloid vacuole and in those which were devoid of anysuch vacuole. These granules may have been formed inthe protoplasm of the sponge-cells in the same way as thelarge vacuoles. On the other hand, it seems very possiblethat they are minute particles resulting from the burstingof a vacuole, and are taken into the substance of the neigh-bouring sponge-cells either as a normal process of nutritionor accidentally.

I may say that the amyloid vacuoles were exceedinglyabundant in specimens of Spongilla taken from the Thamesnear Windsor, in October, and that they were equally abun-dant in pale flesh-coloured specimens of Spongilla and inthose of a bright green tint.

It is of importance to notice that neither granules norvacuoles of amyloid substance appeared to have any relationto the chlorophyll-corpuscles. At the same time, it cannotbe denied, that the probability of the endogenous nature ofthe chlorophyll-corpuscles and of their non-parasitic cha-racter, is greatly increased by the demonstration of the factthat the sponge-cell is capable of forming amyloid substanceand depositing it in vacuoles in large quantities.

Definite observations, localising the formatioh of stareh-like deposits in the cells of an animal organism, have hither-to been wanting, although there are various indications inthe writings of previous observers of starch or starch-likesubstances having been obtained from animals.

There can be no doubt that a careful investigation by thephysiological chemist of the amyloid deposits of Spongilla,and of the substances by which they are preceded andaccompanied, and of the precise conditions under whichthey are produced, would be of great value and interest. Iam inclined to believe that this abundant formation of amy-loid substance—which is in fact most abundant in speci-mens of Spongilla which are actually breaking up anddying down at the in-coming of winter—has possibly a

CHLOHOPHYfcL-CORPUSCLES AND AMYLOID DEPOSITS. 2 4 3

relation to the foraiation of the winter " gemmules," andthe providing them with a store of food material.

Angular corpuscles of colourless Spongilla.—The fact thatSpongilla Jluviatilis occurs almost as frequently in a colour-less or rather pale salmon-coloured state as in the greenstate, is one of very great importance in relation to thenature and history of the chlorophyll-corpuscles found inthe latter form. Whenever Spongilla grows with deficientaccess of sunlight it does not develop a green colour, but itappears to be none the less vigorous. I have seen enor-mous growths (many pounds weight) of colourless Spongillaon the lower surface of a barge removed from the river atOxford. Frequently also in the locks on the Thames,sheet-like growths of Spongilla are seen which are onlymottled with green, their colour being in other parts lightbrown.

This fact, at first sight, seems to tell in favour of thetheory that the chlorophyll-corpuscles are parasitic or-ganisms, which can only attack and thrive in such growthsof Spongilla as are exposed to direct sun light.

An examination of the colourless specimens of Spongillawith the microscope at once, gives a very different significanceto the facts. In the amoeboid cells of the colourless Spongillait is true that no green-coloured corpuscles can be found,but colourless corpuscles are present, which appear to be thesame bodies as the chlorophyll-corpuscles in a modified con-dition. These are angular irregular corpuscles of the sameaverage diameter as the chlorophyll-corpuscles (figs. 6,11,14,PI. XX), and occurring iu the same abundance. If cells betaken from a piece of sponge of a pale green colour, whichis, so to speak, becoming green, individual cases may beobserved in which there are one or two chlorophyll-corpus-cles present amongst the angular colourless corpuscles. Insuch specimens too colourless corpuscles may be detected,which assume the concavo-convex shape of the normalchlorophyll-corpuscles (fig. 7).

It is difficult to avoid the conclusion that the colourlessangular corpuscles are capable of either directly developinginto chlorophyll-corpuscles under the influence of sunlight,or that in the process of their development they can be somodified by the influence of sunlight as to become, insteadof angular colourless corpuscles, concavo-convex chlorophyll-corpuscles.

An important fact in this connection, which I think goesfar to prove that the chlorophyll-corpuscles of Spongilla areformed by the protoplasm of the sponge-cell, was published by

2 4 4 PROFESSOR E. RAY LANKESTERj

me seven years ago in this Journal (1874, vol. xiv, p. 400)JI found that when a piece of colourless Spongilla is dippedinto sulphuric acid, it immediately assumes an intense greencolour. It is well known that sulphuric acid has a similataction upon some vegetable cells, which are remarkable forthe suppression of what may be considered their normalgreen pigment. The saprophyte, Neottia, is devoid ofchlorophyll, but when treated with sulphuric acid certainsubstances in the protoplasm of its cells appear to developrapidly a green-coloured body resembling (at any rate incolour) the green pigment of other plants.

The destructive nature of the reagent employed has pre-vented me from ascertaining, by observing the action underthe microscope, whether the green colour thus developedin colourless cells of Spongilla arises from a change of theangular corpuscles. It can hardly be doubted that this iathe case.

It does not seem possible to hold the view that thecolourless angular corpuscles are colourless parasitic Algaeready to develop into green varieties when exposed to sun-light ! They have even less of the form and structure ofindependent organisms than have the green corpuscles ofthe verdant varieties of Spongilla.

Dr. Brandt's observations and conclusions with referenceto the chlorophyll-corpuscles of Spongilla.—Dr. Karl Brandthas recently published certain observations with referenceto the green-coloured corpuscles of both Spongilla andHydra, which lead him to the conclusion that these bodiesare not " chlorophyll-corpuscles " similar in nature to the" chlorophyll bodies " of plants, but parasitic or " symbio-tic " unicellular Algse.

It is to be hoped that Dr. Brandt will soon publish hisobservations more in detail, together with illustrativefigures. In the memoir which he has already issued Dr.Brandt makes a series of statements, which are applied byhim both to the chlorophyll-corpuscles of Spongilla and tothose of Hydra.

He observes :(1) That he studied the chlorophyll-corpuscles when

isolated from the tissues of the animal by means of pressure.(2) That the corpuscles thus isolated are not equally and

completely green, but possess besides the green-colouredsubstance always some portion of hyaline protoplasm.

(3) In the hyaline colourless part of the green bodies acell nucleus could in all cases be detected with absolutecertainty by treatment with hEematoxylin. Sometimes more

CHLOROPHYLL-CORPUSCLES AND AMYLOID DEPOSITS. 2 4 5

titan one such nucleus was observed (2 to 6), which wereregarded as indications of a process of division.

(4) The green corpuscles were maintained on the objectslide after isolation from the surrounding cell-protoplasm,and were observed to retain their form for several days oreven weeks. When exposed to the light such green cor-puscles (from Spongilla as well as from Hydra) developstarch-grains within their substance.

(5) Isolated chlorophyll-corpuscles from Spongilla werebrought into association with ciliated Infusoria, which,swallowed the chlorophyll-corpuscles. The corpuscles wereeither digested or ejected unchanged.

On the other hand, the larger chlorophyll-corpuscles ofHydra viridis, when similarly swallowed, were found toremain unchanged in the Infusoria for a certain time. Dr.Brandt does not state that these latter corpuscles multipliedby division in the body of the Infusorian which had swal-lowed them.

Upon the grounds summarised in these five paragraphs,Dr. Brandt concludes that chlorophyll is never formed byanimal organisms, but when found in animal cells is due tothe presence of parasitic Algse, to which he has given genericand specific names.

It seems to me that even if we accept every word of Dr.Brandt's statement as to the structure of the chlorophyll-corpuscles there is not sufficient ground for adopting hisconclusions. With regard to the statement contained inparagraph 2,1 am in agreement with Dr. Brandt.

As to the existence of a cell-nucleus (paragraph 3) in thechlorophyll-corpuscles either of Spongilla or Hydra, I amat variance with him. I have not used hsematoxylin as astaining agent in this inquiry, but picro-carmine, and I hadvery fully satisfied myself that nothing like a cell-nucleusexists in connection with the green corpuscles in eitherSpongilla or Hydra previously to Dr. Brandt's statements.I have examined them in various ways, including that ofremoving the green pigment before treatment with picro-carmine. I have found that when a slight staining only isused, sufficient to colour well the nucleus of the amoeboidsponge-cell or of the endoderm-cell of Hydra, no colorationof the protoplasm in connection with the chlorophyll-cor-puscles is to be seen; but if a strong staining be allowed totake place, then the protoplasm of the general substance ofthe cell becomes pink as well as the nucleus of the cell, butin a less degree; and if the chlorophyll-corpuscles besqueezed out of cells of Spongilla so. treated, then the little

2 4 6 PROFESSOR E. RAY LANKESTER.

piece of protoplasm adherent to each (fig. 9 d, PL XX) willbe seen to have a pink colour, but is still perfectly homo-geneous.

Similarly the colourless protoplasm within the green cor-puscles of Hydra will take up a pink colour when strongstaining with picro-carmine is used; but nothing of thenature of a nucleus have I ever seen in these corpuscles,although the little granules within the corpuscles of Hydramight lead to the impression that a nucleus is present,(figs. 20, 23) if one were not acquainted with their truenature, as isolated granules of green-coloured substancelying within the corpuscle.

It seems to me possible that Dr. Brandt has been misledby these granules. At the same time it is possible thathsematoxylin brings into view a nucleus-like structure,which picrocarmine does not. Even if this were the case,when we remember that the chlorophyll-bodies of plantsare looked upon by botanists as similar in their nature tonuclei, it does not seem that we should have any groundfor regarding the chlorophyll-corpuscles as independentorganisms.

Dr. Brandt's observation of the formation of starch in theisolated chlorophyll-corpuscles (paragraph 4) is extremelyinteresting and important. It does not seem to me to tendin any way to prove that the corpuscles are independentorganisms. It would simply prove (if fully established) thata bit cf protoplasm with its associated envelope or cap ofgreen substance can retain its vital activity just as a pieceof an Amoeba can. At the same time what I note as espe-cially interesting is that Dr. Brandt does not state that hehas observed starch-grains in association with the chloro-phyll-corpuscles when observed in fresh living cells ofSpongilla (or of Hydra). I have failed to detect starch insuch position in living Spongilla-cells, though I have foundabundant amyloid substance in other parts of the sponge-cell. I have also, only in the rarest cases, found a minutetrace of starch in association with the chlorophyll-corpusclesof Hydra viridis.

This absence of starch from the living chlorophyll-corpuscles when in the sponge-cell, or Hydra's endoderm-cell, must necessarily appear remarkable. I have beendriven to the conclusion that the activity of the chlorophyll-corpuscles in these animals in sunlight gives rise to a bodysimilar to that which arises under the same conditions inplants, but that in place of being deposited in the corpuscleas starch-grains, it is rapidly diffused and chemically changed

CHLOROPHYLL-CORPUSCLES AND AMYLOID DEPOSITS. 2 4 7

in the surrounding protoplasm of the cell. In Spongilla,under certain circumstances, it is deposited as amyloid sub-stance (after diffusion) in the large vacuoles described aboveand figured in PI. XX, figs. 1, 3, 4, 14.

Now it appears not improbable that by removing thechlorophyll-corpuscles from the mass of surrounding proto-plasm, Dr. Karl Brandt has found a method by which theproduct of the activity of the chlorophyll-corpuscle may he, asit were, forced to remain in the corpuscle, there being nosurrounding protoplasm to take it up and operate furtherupon it. Hence, possibly enough, we get a deposit of starch-grains in the isolated corpuscle which would never occur inthe normal condition, since the product of assimilation is inthat condition rapidly diffused and so removed from thechloroph y 11-cor p uscle.

The inquiry suggested by Dr. Brandt's observation onthis point seems likely to have valuable results.

With regard to Dr. Brandt's experiments in infectingInfusoria with the supposed parasites of Spongilla andHydra (paragraph 5), it is at once apparent from his accountof them that they are opposed to and not in favour of theparasitic theory.

The chlorophyll-corpuscles of Spongilla were digested orelse ejected by the infected Infusoria. In other cases thechlorophyll-corpuscles of Hydra remained in the Infusorian'sbody unchanged. Had Dr. Brandt's view been confirmed,the green-corpuscle ought to have multiplied in its newhost, and even such evidence of a temporary manifestationof vitality after removal from the Hydra or Spongilla wouldnot in my opinion be at all conclusive to the effect that thechlorophyll-corpuscles are independent organisms, and notparts of the protoplasm of the cell in which they arenormally found.

HYDRA VIRTDIS.

Professor Nikolas Kleinenberg, in his memorable work on'Hydra / has given an account of the chlorophyll-corpusclesof H. viridis, which leaves little to be added on the subject.He has not, however, given any of the special features ofthe chlorophyll-corpuscles in his plates, and the figureswhich are given in my PI. XX, figs. 15—27, are, I believe,the first which adequately represent those bodies. Kleinen-berg says of them:

" They consist of (1) a dense ground-substance, very richin albumens, which stains dark brown with iodine, deepred with carmine or aniline; (2) and spread over this an

248 PROFESSOR E. RAY LANKESTER.

excessively thin coating of a green colouring matter, whichto judge by its chemical and optical characters is eitheridentical with chlorophyll or very near to it. These cor-puscles therefore exactly correspond, in regard to their con-struction, with the chlorophyll-bodies of plants. In acertain number of them the surface is quite smooth, othersacquire a segmented appearance from grooves and fissures.(See PI, X X , fig. 17a.) Related to the latter are smallcorpuscles in the course of breaking down, which exhibitangular forms, and instead of a bright green have a dirtydark colouring (PI. XX, fig. 21), and gradually pass overinto very small dark brown or even black granules, adheringto one another in little heaps (PI. XX, fig. 19). Thenumber of all these bodies varies greatly according to thenutritional condition of the animal. The green bodies arefound chiefly in the marginal region of the cell, and only•when they are very abundant in its basal portion; occasionallythey appear to be, as it were, stuck on to the cell-surface, butnevertheless always possess a thin envelope of protoplasm.The free end of the cell never contains chlorophyll-corpuscles ; on the other hand, the brown and black granulesare accumulated in that region.

" In H. auranti(tca and grisea1 the endoderm-cells of theToot and tentacle cavities are devoid of form-elements com-parable with the chlorophyll-corpuscles of H. viridis. Onlyorange, brown, and blackish spherical or angular corpusclesoccur, which all exhibit a remarkable resistance to chemicalreagents. The epithelium of the stomachal cavity containshowever—at least in well-nourished specimens—colourlessround or oval dense albuminous corpuscles (PI. X X , fig. 16),which, excepting the absence of chlorophyll, closely resemblethe coloured corpuscles of H. viridis, and also exhibit thesame transitional forms leading to the dark granules."

I have introduced into this quotation from Kleinenbergreferences to the figures of the plate accompanying thepresent memoir. The only important addition which I haveto make to Kleinenberg's statement, as to the structure ofa normal chlorophyll-corpuscle of Hydra viridis, is that veryusually there is within the shell or crust of the green-coloured substance one or more minute granules, alsocoloured green and embedded in the colourless protoplasmof the corpuscle.2

1 Apparently a synonym of our H.fusca.5 I would, however, analyse the chlorophjll-corpuscle of Hydra into

three elements as follows—(1) chlorophyll or green pigment, which can bedissolved out by alcohol, (2) chromophorous substance, which carries the

CHLOROPHYLL-CORPUSCLES AND AMYLOID DEPOSITS. 2 4 9

Sometimes these are absent, and it seems that specimensof Hydra viridis may be obtained, according to season andstate of nutrition, in which such internal granules arepresent in all the corpuscles, or, on the other hand, absentfrom all or nearly all of them.

The corpuscles average about twice the diameter of thechlorophyll-corpuscles of Spongilla, that is to say, from4-sVffth to Tnnnrth °f a n i n c n - Some are larger. The chiefdifference between the corpuscles in the two animals is foundin the fact that the green substance forms a concavo-convexcap upon its related protoplasmic base or corpuscle in Spon-gilla, whereas in Hydra the coloured cap is extended on allsides, so as to form a hollow sphere enclosing some protoplasm,and additional granules are developed within the sphere. Ithas been pointed out above that, as an exception, a greencorpuscle may be found in Spongilla having the form charac-teristic of Hydra (see PI. XX, fig. 12); so, too, in Hydrasmall green corpuscles may be found, which have the greeninvestment incomplete and cap-like (PI. XX, fig. 20 a,f, h).Kleinenberg observed the staining of the colourless proto-plasm within the green corpuscles of Hydra by iodine, car-mine, and aniline. This I have also observed, but foundit to be much less intense than that of the cell-nucleus, sothat it is possible to obtain a staining of the nucleus whilstthe corpuscles remain unstained (PI. XX, fig. 15).

Kleinenberg makes no mention of any nucleus-like body•within the green corpuscles, and I am in accord with him.Dr. Brandt states (loc. cit.) that a cell-nucleus can be clearlydemonstrated hi each chlorophyll-corpuscle, and makes thishis chief ground for regarding, the corpuscles as parasiticAlga?. I have not found such a nucleus in Hydra any morethan in Spongilla, and find it difficult to believe that, ifpresent, it would have escaped the careful examination madeboth by Kleinenberg and myself.

Kleinenberg makes no mention of starch in connectionwith the chlorophyll-corpuscles or other parts of the endo-derm-cells of Hydra. I have also not succeeded in findingstarch in these bodies. I have, however, very rarely ob-tained a blue coloration with iodine in the neighbouringprotoplasm, and in one specimen a few granules (not chlo-

chlorophyll, and is usually in the form of a spherical shell, enclosing factorNo. 3, but may also occur as granules embedded in that third factor; thechromophorous substance resists the action of staining agents. (3) Colour-less homogeneous protoplasm, enclosed by the shell of chromophoroussubstance, and capable of taking a stain with strong colouring agents,but devoid of nucleus or nuclear matter.

2 5 0 PROFESSOR E. RAY LANKESTER.

rophyllaceous), previously colourless, lying in the cell-protoplasm, gave a blue colour with iodine. I would venturethe surmise that in Hydra viridis, as in Spongilla, the pro-duct of assimilation due to the activity of the chlorophyll-corpuscles is very rapidly diffused, and does not take theform of starch as in green plants, or if it does take that form,it is after diffusion from the seat of assimilation.

A very strong argument against Brandt's theory of theparasitic nature of the chlorophyll-corpuscles is found in thefact noticed by Kleinenberg, that minute angular fragmentsof a green colour are often present, together with the normalcorpuscles (PI. XX, fig. 21). In fact, in Hydra, as inSpongilla, though there is a normal and fairly constant formof chlorophyll-corpuscle, yet other irregular forms appearside by side with these. How can such irregular forms beexplained on the parasite theory ? They present no diffi-culty if the corpuscles are regarded as products of theanimal's cell-protoplasm, for it may well be that such pro-ducts should sometimes be incompletely formed or ofmonstrous size and shape.

The brown and blackish granules noted by Kleinenbergas occurring both in H. viridis and in the "greenless"varieties of Hydra are important (PI. XX, fig. 19). Theyappear to result from the breaking down of the chlorophyllbodies or their colourless representatives.

Professor Jeffrey Parker, in his paper on the endoderm ofHydra (' Quart. Journ. Micr. Sci./ vol. XX), has advancedthe view that these dark granular bodies are ingested foodparticles in the course of digestion. In this view I cannotagree. They appear to me to be undoubtedly in the caseof JS. viridis connected with a degeneration of a chlorophyll-corpuscle, and in H.fusca I am inclined equally to attributethem to the formative activity of the cell-protoplasm. Similardark-coloured granules in the endoderm of Cordylophora andother hydroids (probably also the very scarce black granulesin the endoderm of the Medusa Limnocodium, ' Quart. Journ.Micr. Sci.J (vol. XXT, PI. VIII, fig. 1 b), are also to beregarded as products formed by the cell, and not as ingestedparticles. However, it is difficult to distinguish one kind of" dark granule" from another, and it is possible that somesuch dark granules, observed in the endoderm-cells ofHydrozoa, are really food particles which are undergoingintra-cellular digestion.

The representatives of chlorophyll-corpuscles in colourlessand olive green Hydrce. — It is an open question as towhether the white, brown, and orange-coloured specimens of

CHLOROPHYLL-CORPUSCLES AND AMYLOID DEPOSITS. 2 5 1

Hydra are to be regarded as species distinct from Hydraviridis or as varieties. I incline to take the latter view,since transitional forms are met with, namely, olive-greenand bluish-green specimens, which have "incompletelydeveloped chlorophyll.

There is no such clear evidence of the specific identity ofHydra viridis and Hydra fusca as there is of the identityof colourless and green varieties of Spongilla. In the lattercase one and the same piece of sponge may be fovind greenwhere exposed to sunlight, and pale flesh colour whereshaded from the light.

The experiment has not, I believe, been made of main-taining Hydra viridis in obscurity, though Max Schultzeobtained in this way colourless individuals of Vortex viridis.

Commonly in this country large numbers of a very palebrown Hydra are found associated without the presence ofany green specimens. These colourless Hydrse (H. fusca)are larger than the H. viridis usually is. But I know ofno character separating the two beyond colour and size.The larger size of H. fusca, if it be regarded as a variety ofH. viridis, may well be correlated with the non-develop-ment of chlorophyll, and a less active growth. For thoughthe individual is large it may possibly be less active in buddingin H. fusca than in the smaller H. viridis. Small size andrapid fission may go well together with the nutritional advan-tages represented by the possession of chlorophyll-corpuscles.

In the endoderm-cells of such specimens of H. fusca (asdescribed by Kleinenberg for his H. aurantiaca and H.grisea) there are angular and rounded colourless bodies (PI.XX, fig. 16 g), which appear to represent in a colourlessstate the green corpuscles of H. viridis. At the same timethey are not definitely spherical, as are the latter. Just thesame kind of difference is observed in relation to these cor-puscles between H. viridis and H. fusca as there is betweengreen and colourless Spongilla in relation to their corpuscles.

I have not made the experiment of subjecting Hydrafusca to the action of sulphuric acid, a reagent which, asnarrated above, develops a green colour in colourless sam-ples of Spongilla. But I have examined carefully somevery interesting specimens of Hydra which were found bymy assistant Mr. A. G. Bourne in company with Hydrafusca, but which were of an olive green or dull blue greeninstead of pale brown.

These exceptional individuals appear to me to have greatinterest. I found in their endoderm-cells that a certainamount of green pigment was developed, but instead of the

2 5 2 PROFESSOR E. RAY LANKESTER.

green bodies having the form of spherical corpuscles, theyformed irregular angular masses, as shown in PI. XX, fig. 18.Also, I was able to obtain, by squeezing from one and thesame endoderm-cell, groups of angular granules arrangedsymmetrically as parts of a sphere, of which some werecolourless, whilst others were green (PI. XX, fig. 22).

I consider this strong evidence in favour of the view thatthe colourless angular bodies of H. fusca are potentiallychlorophyll-corpuscles; that is to say, under certain cir-cumstances they may develop in themselves chlorophyll.What the conditions are precisely, we are as yet unable tosay.

It is noteworthy ihat both in Spongilla and in Hydra,when the pigment bodies remain in an abortive condition,they are irregular and angular; when they develop chloro-phyll green on the other hand by peripheral activity, theytend to the spherical condition. This may, it seems, beconnected with the fact that the formation of the chloro-phyll is essentially a surface activity probably dependent onthe access of sunlight, and this surface activity would, ifperfectly symmetrical, necessarily result in the productionof a sphere.

Dr. Brandt's conclusions with regard to the chlorophyll-corpuscles of Hydra viridis.—The summary given a fewpages back of Dr. Brandt's statements in reference to thecorpuscles of Spongilla applies equally to those of Hydra.

I am unable to see that he has adduced any facts, except-ing the presence of a nucleus, which. I doubt, which tend tothe conclusion which he has so definitely formulated byassigning to the green corpuscles of Hydra the name Zoo-chlorella conductrix.

There is in all that is known of the structure of thechlorophyll-corpuscles of Hydra, as of Spongilla, nothingwhich separates them in character from the known chloro-phyll bodies of plants. On the other hand, it is not onlyimpossible to characterise Dr. Brandt's genus " Zoochlo-rella " in botanical language, but there are a variety offacts known both as to the objects called by him Zoochlorellaconductrix and as to those called Zoochlorella parasitica,which lead to the conclusion that in thus dogmaticallyasserting the parasitic and algoid nature of those objects,Dr. Karl Brandt has wandered very far from the legitimateinferences warranted by the facts.

No distinct wall, either of cellulose or of other substance,exists external to the green-coloured cap or shell of thechlorophyll-corpuscles of Hydra and Spongilla. Their

CHLOROPHYLL-CORPUSCLES AND AMYLOID DEPOSITS. 2 5 3

form, especially in the latter, is very varied. In some thegreen colour is very partially deposited in granules andsuperficial caps, in others it is absent altogether, and thecorpuscle is irregular and angular in form.

Conclusion as to the parasitic or non-parasitic nature of thechlorophyll-corpuscles of Hydra and Spongilla—The finalconclusion to which we are led in relation to the chloro-phyll-corpuscles of Spongilla and Hydra is that a carefulstudy of these bodies reveals in both cases their correspon-dence with the known structure of the chlorophyll bodies ofplants, and that those who, like Semper and Brandt, havesupposed these chlorophyll-corpuscles to be parasitic, havebeen misled by, firstly, an imperfect acquaintance with thecharacters of chlorophyll bodies in general and of these inparticular, and, secondly, by the plausible but delusiveanalogy presented by the " yellow cells " of Radiolarianaand of Anthozoa.

As to the nature of these latter bodies, I have no observa-tions to offer.

The chlorophyll-corpuscles of Spongilla and Hydra inrelation to Pringsheim's theory of chlorophyll.—Now thatwe have established the occurrence of " chlorophyll/' or thecombined substances which together constitute that pigment,in Spongilla, and with nearly equal certainty in Hydra, andalso have come to the conclusion that the " chlorophyll" isformed in corpuscles in the cells of those animals just as itis in green plants, it becomes very important to knowwhether the chlorophyll in the animal is serving the samepurpose as it is in the plant; and if so, whether we may notbe able to get indications from the animals as to the disputedfunction of the green pigment, such as plants are unable tofurnish.

There is no doubt a field for experimental inquiry here,and with the memoir of Pringsheim in his hand the zoolo-gist may carry out a variety of inquiries upon " animalchlorophyll."

I would here only briefly insist on one or two remarkablefacts which are apparent, and which bear upon the generalquestion of the function of chlorophyll.

In the first place, what we may call " greenless " Spongillaand " greenless " Hydra flourish abundantly in the samewaters with green-coloured Spongillse and green-colouredHydrae. Hence, whatever value attaches to the chlorophyll-it cannot be a very great one in relation to the vital pro-cesses of these animals.

In the second place, no starch can be found in immediate

2 5 4 PROFESSOR E. RAY LANKESTEB.

relation to the chlorophyll-corpuscles of either green Spon-gilla or green Hydra, when in normal conditions. But, onthe other hand, an amyloid substance is formed by the greensponge-cell, and stored up in large quantities in vacuoles.

Equally large quantities of starchy matter are formed bygreeuless Spongilla. Accordingly, the Spongilla is notdependent upon chlorophyll for its power of forming amyloidsubstance. This formation of amyloid substance appears tobe due to a synthetical process resident in the colourlessprotoplasm of the sponge-cell. I t is not yet known thatthe process is a synthesis, or that the decomposition of CO2is connected with i t ; but if this could be proved to be thecase, we should have strong evidence in favour of the" screen theory " of chlorophyll. For we should then havethe amyloid synthesis going on equally both in green and" greenless " Spougilla, in the former the protoplasm beingprotected by chlorophyll from the direct sunlight, in thelatter no such protector being required nor developed, andthis because the greenless Spongilla exists in deep shadeaway from the reach of those rays which it is the businessof chlorophyll to intercept.