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Beijerinck has with great ability made use of the luminous bacteria for detecting the most minute quantity of an enzyme."

The following example will illustrate this: He takes advantage of the fact that Photobacterium phosphorescens displays light with maltose, while Photobacterium Pflügeri does not. He uses a thoroughly cooked mixture of sea water containing 8 per cent of gelatin, 1 per cent of pepton, and one-fourth per cent of potato starch. To a portion of this he adds an excess of Photobacterium phosphorescens and to the rest the same of Photobacterium Pflügeri, and prepares from these two similar gelatin plates equally illuminated. In both the starch remains unchanged, seeing that these bacteria are unable to secrete the necessary enzyme, diastase. If now some diastase preparation (such as maltose, pancreas-diastase, or ptyalin) is added to these plates, it distributes itself in all directions, transforms the starch into grape sugar, and upon the field of the Photobacterium phosphorescens there instantly appear strong, shining flecks, which later spread over the whole field of growth, while on the field of Photobacterium Pflügeri nothing of this kind is to be seen. In this way Photobacterium phosphorescens can be made to demonstrate through its luminosity the presence of maltose that is to say, of a diastase.

For an understanding of the nature of light development in plants it is above all necessary to state that the luminosity is absolutely dependent upon free oxygen. The light is conditioned on oxidation. The finest investigations of the dependence of luminosity upon oxygen are the brilliant experiments of Beijerinck. According to his researches the luminous bacteria afford the most sensitive tests for oxygen that we possess. Thus the extremely minute quantity of oxygen which unicellular algæ give off in sunlight by their assimilation of carbonic acid gas, is sufficient to instantly render these bacteria luminous. If we introduce these green cells into a glass tube filled with bouillon containing luminous bacteria, the light quickly disappears because the bacteria speedily consumes the oxygen contained in the liquid. If, now, such a tube is kept in a dark room and then the light of a match is allowed to fall upon it for a single second the entire mass grows luminous. The green cells give off oxygen, and this fabulously minute quantity of the free gas is sufficient to cause luminosity in the bacteria. It is a remarkable example of the fact that physiological methods not only compete well with the best physical and chemical methods, but plainly surpass them, and that a vital

a An enzyme is a product of certain plant or animal cells by means of which food material of a certain kind, such as starch, is transformed into another food material, such as grape sugar. Diastase is an example of an enzyme.Translator.

function itself can be used as a methodical factor of science of the highest value.

A demonstration of the importance of oxygen to luminosity can be made before a large audience in the following way: A glass tube closed at one end, having a diameter of about 8 mm. and a length of 1 to 1 m. is filled to within to 1 cm. of the top with strongly luminous bouillon (bouillon mixed with Bacterium phosphorium or Pseudomonas lucifera). Such a tube at the expiration of a quarter of an hour loses its light as the bacteria exhaust the oxygen, except the mere upper surface of the liquid in contact with the air. If, now, the tube is closed with the thumb and inverted, a bubble of air will ascend through the bouillon, making its entire course luminous and appearing in the darkness like a slowly ascending skyrocket. In a quarter of an hour or less the luminosity again disappears, and the experiment can be repeated.

Botanists, as a rule, teach that a direct relationship exists in the fungi between the development of light and respiration. Thus Sachs speaks of phosphorescence as the necessary consequence of respiration, of phosphorescence by means of respiration. But F. Ludwig has already demonstrated that luminous bacteria can be cultivated, and therefore made to grow and breathe without any luminosity; and we can therefore easily see how that at increasing temperatures the intensity of respiration may steadily increase, but the intensity of luminosity to only a limited degree. The relation existing between light development and oxygen is analogous to that between color development and oxygen. Most of the colorproducing bacteria show color only in the presence of oxygen, as can be seen in gelatin cultures into which an infected needle has been introduced. When the free gas can reach the bacteria, the color appears, but deeper in the gelatin the bacteria, cut off from oxygen, develop without color. Color production and light production are therefore oxydation phenomena.

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During recent times quite a number of investigators have been incidentally or directly engaged in throwing light upon the nature of luminosity--E. Pflüger, Radziszewski, Dubois, F. Ludwig, Katz, Tollhausen, Lehmann, Beijerinck, McKenney, and Nadson. However, their interpretations differ from each other considerably. The further our knowledge of the subject is extended the more probable appears the idea that within the cell is a hypothetical substance, photogen," which has the power of producing light in the presence of free oxygen. This idea receives substantial support from the fact brought out by Radziszewski that a long list of organic substances, such as aldehyde materials, ethereal oils, carbonic-acid, water, fatty oils, and certain of the alcohols, have the power of luminosity when brought into alkaline reaction with active oxygen.

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As the light produced by these substances has an external and spectroscopic likeness to that produced by living organisms, and as some of the substances in the list of Radziszewski which are capable of luminosity exist also in living cells (I mention lecithin, fats, cholesterine, ethereal oils, and grape sugar) that investigator has come to the conclusion that light development in living organisms can be explained as an oxydation of these same substances. Radziszewski looks upon the problem, therefore, as solved. We have, however, as yet hardly gotten that far. The question of whether Radziszewski is in the right could be definitely settled if we could extract from the living cell a photogen material which would show luminosity outside. of the cell. But up to the present time the attempt has not been successful. Furthermore, according to Pfeffer, no active light-causing oxygen exists in the living plant cell, which does not square with the theory on which Radziszewski's explanation rests, inasmuch as his light-producing substance is luminous only when in contact with active oxygen. Nevertheless, I look upon the photogen theory as the most plausible, though we at present have no knowledge as to the nature of photogen. Possibly it is a material in no sense similar to the luminous substances previously mentioned; perhaps something capable of giving light without active oxygen.

There are certain facts which appear to me to directly support the idea of a photogen. Thus certain animal organisms give out a noncellular luminous secretion, and certain cells together with their contents are capable of producing light when no longer living. Mention can be made of Pholas, certain of the insects, myriapods, and many of the worms. A fact of significance and too little noted is that certain tissues and cells have the power of producing light in a lifeless condition. Thus manuscript written with the luminous material obtained from Luciola italica gives off light when it is dampened. The light organs of Lampyris noctiluca lose their luminosity when thoroughly dried and kept in a vacuum. But, according to Bongardt, if after a year's time they are taken out and moistened with a drop of distilled water the light reappears. If filter paper is impregnated with the secretions of certain myriapods, it can, after two months' time, be made luminous by moistening. It is impossible in such instances to talk longer about "living cells" or "living cell contents," for it is impossible to describe as living the luminous material from an insect that has been dried and kept for a year in a vacuum. In such instances we are no longer dealing with a vital but with a purely chemical process; we are dealing with a substance which produces light in the presence of water and free oxygen.

In the case of luminous plants no such thing as a luminous excretion exists, though such is erroneously stated to be the case, for the light exists only within the cell. In other words, it has never been seen outside of the living plant cell, and to that extent luminosity in

plants must be spoken of as a genuine vital luminosity. But in the same way not long ago alcoholic fermentation was held to be inseparably connected with living yeast cells, while to-day, thanks to the brilliant biochemical discoveries of Buchner, we know that it is due to a certain material-the ferment zymase-which can of itself, although a lifeless substance, bring about the fermentation. We can suppose the same to be true for photogen. Although the isolation of such a luminous material has not as yet been accomplished, the failure is probably due to the material being present in such very minute quantity, to its extreme instability, and its destruction through the death of the cell. What photogen really is, and whether the giving of light represents a process of fermentation-these questions can not at present be answered. The future investigator must unearth these facts. To directly or indirectly prove the existence of photogen; to, if possible, isolate it from the cell, and then render it luminous—such efforts, in the light of other biochemical facts, appear to me most tempting and by no means unpromising.

Whoever has observed the swarms of fireflies flying through the darkness of the night like wandering stars, or the intense light of pure cultures of bacteria and the higher fungi, must involuntarily have been impressed by the peculiarity of these "living" lights. And therefore it is easy to see that at a time when the science of physics has surprised us with unexpected revelations, appearing at first like marvels, that we should with redoubled activity turn our attention to the nature of this light coming forth from life, and seek to discover its physical, chemical, and physiological activities.

I wish, first of all, to call attention to a noteworthy difference between the character of this light in the animals and in the plants. If we leave out of account the Peridineæ and confine ourselves to the plants alone, we see they are always steadily luminous. The bacteria and higher fungi give forth light for days, weeks, months—indeed, under some circumstances, as when supplied with abundant nourishment, for years, without cessation, day and night, while the animals, with few exceptions, shine only a short time, a few seconds or minutes, and mainly in response to some external irritation; so that the light gives the impression of a flash or spark. The light of the fungi is of a white, green, or blue character, and, contrary to earlier statements, never undulates like the light of phosphorus; never is inconstant or glimmering, but is in all cases quiet, steady, and constant, whether viewed with the naked eye or through the microscope. As a rule, its intensity appears to be low, and yet there are bacteria so intensely luminous that they can be seen on a bright day in the corner of a room without the eyes being accustomed to the darkness. A remarkable object in this respect is Bacterium phosphoreum (Cohn) Molisch, the luminous bacteria of butcher's meat, and, to an even

greater extent, Pseudomonas lucifera Molisch, a photobacterium which two years ago I discovered in marine fish, the light intensity of which surpasses that of any luminous bacteria heretofore known.

To R. Dubois is due the credit of having first attempted to utilize bacterial light in the form of a lamp, and I have renewed Dubois's attempt with the two already mentioned intensely luminous bacteria, and have constructed a bacterial lamp on the following plan: In a Florence flask, having a capacity of from 1 to 2 liters, is put from 200 to 400 cubic centimeters of salt-peptone-gelatin. It is then stopped with cotton wool and sterilized. When cool, but before the gelatin has quite solidified, it is infected with a culture, fresh and luminous, of Bacterium phosphoreum or Pseudomonas lucifera, a platinum needle being used. The flask is held horizontally and slowly rotated, so that the gelatin forms a coating on the entire inner surface of the flask and then hardens. After being kept for one or two days in a cool room, the entire inner surface of the flask is covered with colonies of bacteria, so that it gleams with an exquisitely beautiful bluish-green light, and presents with its soft, steady brilliancy a splendid appearance. I have lately found that the luminous power of such a lamp can be considerably augmented by applying the infection to the gelatin in parallel lines about 1 centimeter apart, running from the bottom to the neck of the flask, and adding to the gelatin 1 to 2 per cent of peptone and about one-half per cent of glycerin. Such a lamp will continue luminous in a cool room for about fourteen days, and when the eye is accustomed to the darkness will give light enough to see the face of a watch, the scale of a thermometer, or to read coarse print. Such a flask is visible on a dark night at a distance of 64 paces, and could in an emergency be utilized as a night lamp. Inasmuch as dead luminous flounders are successfully used as bait by fishermen on account of their light, a lamp of this kind could be made to serve as a valuable lure in catching fish.

My investigations warrant me in stating that in the future it will probably be possible, by means of exact formulas of nutriment and by selective breeding, to so increase the intensity of this exceedingly cheap source of light, so free also from heat rays, that on account of its cheapness, its long and uninterrupted luminosity, its freedom. from danger, and its lack of heat it can be turned to practical account in powder magazines, in mines that are not too warm, and in other places.

In connection with the investigations of F. Ludwig and Forster, I may state, in regard to the luminous bacteria and mycelial fungi, that their light spectra are continuous, without dark lines, and, as a rule, simply luminous spectra-that is to say, on account of their low intensity they are colorless; that the spectrum of the already named bacteria shows a more decided trend toward the violet end of the

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