| Description |
This study represents an investigation and comparison of the effects of various Veratrum alkaloids on nuclear mitotic division in an effort to determine 1. whether some purified alkaloidal mixtures or crystalline alkaloids possess the ability to modify the cycle of nuclear and cellular division, and 2. to determine how such activity compares in kind a degree to that already well established for colchicine. In addition, preliminary studies have be conducted to determine what effect a number of physical and chemical factors have on nuclear mitotic division being studied in order to eliminate their influences as experimental variables. Onion bulbs, Allium cepa var. White Portugal, were used as the experimental plant. Roots were sprouted on the bulbs using tap water and experiments were conducted under constant temperature (30°C.±0.5°C.) conditions. Roots from 1.5 to 6.0 cm. in length were used for all observations of nuclear division. Microscopic examination of the cells was performed by root tip smear method in which root tips were killed and fixed for 24 hrs. or longer, softened to separate the individual cells, rehardened, stained, and observed through a microscope at magnification of 440 diameters. All intact cells observed in a predetermined area of a glass slide, either resting or in any stage of mitotic nuclear division were recorded and the per cent of cell in the process of nuclear division at the time of observation was calculated. Suitable statistical technics were employed (chi-square, linear regression, and analysis of variance) to analyze the experimental results and permit conclusions to be drawn there from. No significant differences were noted from control values in nuclear division between times of exposure of 3 and 6 hrs. to a 2 per cent solution of 2-chloreothanol. Three hr. bulb soaking failed to stimulate germination of a significantly greater number of bulbs than was produce by tap water alone; however, production of new roots was significantly increased over results observed for tap-water controls. These data suggest that pre-soaking periods for Allium bulbs are not harmful to nuclear mitotic division. Further, the agent appears to be incapable of breaking bulb dormancy in the concentrations used and for soaking time employed by is highly effective in increasing the germination rate of new roots from non-dormant bulbs (approximately 19 per cent over control values). Although there was no significant difference in over-all nuclear mitotic division between roots up to 7.5 cm. in length, very short roots (up to 1.0 cm in length) were consistently low (2.4 per cent) in the number of dividing nuclei from that of a control (4.0 per cent) which suggests that short Allium roots up to 1. cm. long increase in length more from cell elongation than by meristematic mitotic production of new cells. To eliminate the influences of root length on nuclear division as an experimental variable, only roots from 1.5 to 6.0 cm. long were used for all observations. In a population sample of 40 Allium bulbs the number of mitotic dividing nuclei was not significantly changed for different times of day when observations were performed on roots obtained every 3 hrs. over a 24 hr. period of time. The effects of light, intermittent light, and darkness on nuclear division and root length over a 72 hr. test period indicated that prolonged exposures to artificial light had a noticeable but not-significant inhibitory effect (P>.05) on mitotic nuclear division and a marked inhibitory effect on root length (P<.001). In order to eliminate the influences of light as an experimental variable on the number of mitotic dividing nuclei, Allium roots should be exposed to known intensities of light which may be controlled at all times. Exposure of Allium roots to 7 different temperature gradients from 15 to 45°C. in increments of 5°C. for 24 hrs. revealed that the mean number of mitotic dividing nuclei exposed to 20, 25, 30, and 35°C. were not significantly different from the mean values obtained from roots exposed to the control temperature (30°C.), whereas means at temperature extremes of 15, 40, and 45°C. differed significantly (40 and 45°C., P<.01) from controls. This suggests that rather wide temperature fluctuations may be permitted without producing undesirable alterations in mitotic nuclear activity. Temperature extremes, however, may have marked effects on mitotic processes; therefore, whenever possible, experiment should be conducted under well controlled temperature conditions. Examination of Allium bulbs treated with concentrations of colchicine of 0.1, 0.2, 0.4, or 0.8 percent for predetermined periods of time of 3, 6, 9, or 12 hrs. revealed that the time and concentration factors both caused significant changes in nuclear mitotic division from control values. Independent of the time factor, all colchicine concentrations had an inhibitory effect. When time was examined as the major variable, significant differences in nuclear mitotic activity were also observed. The data also showed the existence of a significant interrelation between the effects produced by the time and concentration factors. Typical colchicine-mitotic (c-mitotic) effects were produced with all colchicine concentrations had an inhibitory effect. When time was examined as the major variable, significant differences in nuclear mitotic activity were also observed. The data also showed the existence of significant interrelation between the effects produced by the time and concentrations factors. Typical colchicine-mitotic (c-mitotic) effects were produced with all colchicine concentrations employed. The data suggests that colchicine, studied by these experimental methods, is capable of modifying nuclear mitotic activity by producing typical c-mitotic changes in the chromosomes of Allium roots. Successful inhibition of the spindle apparatus may be achieved and polyploidy cell (2n = 32) are prominent. When using colchicine experimentally, not only must be concentrations be taken into account but also the exposure time must be considered. The effects produced by Veriloid, a standardized mixture of hypotensively active alkaloids present in Veratrum viride Ait. Were examined in Allium bulbs treated with acidified solutions which contained concentrations equivalent to 0.1, 0.2., 0.4., or 0.8 per cent of Reference Standard Alkavervir (Riker Laboratories in frees tape water for predetermined periods of time (3, 6, 9, or 12 hrs.). The data suggests that alkaloidal agents present in Veratrum viride have and effect on nuclear mitotic division in roots of Allium cepa L. Chromosomal aberrations were frequently observed although their appearance suggested a markedly different mode of action than that observed with colchicine. Polyploid cells may be noted and the effects on nuclear mitotic division are in agreement with those reported by Witkus and Berger (1944) for veratrine, an alkaloidal mixture obtained from a different plant source (Schoenocaulon officinale A. Gray) but whose constituents are structurally similar to those found in Veriloid. The effects produced by protoveratrine A, a purified crystalline alkaloid present in Veratrum album L. were examined in Allium bulbs treated with a 0.025 per cent acidified solution in fresh tap water for predetermined periods of time (3, 6, 9, or 12 hrs.). The results suggest that protoveratrine A is capable of modifying nuclear mitotic division to produce polyploid cells in roots of Allium cepa. The changes observe red were indistinguishable from the changes reported in the Veriloid study. |
