The LSD molecule contains a number of pi-electrons. These pi-electrons absorb electromagnetic radiation very strongly, like many aromatic or conjugated systems. LSD absorption is maximal at 320 nm, and LSD fluorescence is maximal at 435 nm. These values are somewhat flexible depending on the spectrophotometer, with some authors reporting LSD absorption at 325 nm, and LSD fluorescence at 445 nm. The spectroscopic properties of radioactive [3H]-LSD are the same as LSD, with a maximum fluorescence at 445 nm, and an excitation wavelength of 325 nm. For LSD spotted onto chromatography plates, maximum excitation occurs at 330 nm, and emission at 410 nm. Spots of LSD on thin-layer chromatography plates give a violet-blue fluorescence under a UV lamp.
Mueller and Lang carefully studied the absorption and fluorescence of LSD as a function of pH. As shown in Figure 1 above, there is a minor excitation peak at 240-250 nm in addition to the major excitation peak of LSD at 330 nm. The major excitation peak of LSD could be shifted by placing the LSD sample in either acid or alkali conditions. The major LSD excitation peak was 327 nm in 0.01 N HCl and 319 nm in 0.05 M Na2HPO4. In basic solution, the 319 nm peak was approximately 25% greater in amplitude than the height of 327 nm in HCl. Major emission peaks of LSD were at 420-430 nm, with a shoulder at 536 nm.
In 1971, in situ fluorometry was performed on LSD using quinine as an internal reference. As seen in the figure below, quinine (A2, B2) and LSD (A1, B1) have the same excitation and fluorescence wavelengths.
LSD fluorescence can be used to check its potency. Niwaguchi and colleagues found a linear relationship between fluorescence emission intensity and the amount of LSD on thin-layer chromatograms. If the 9,10 double bond of the D-ring is intact, blue fluorescence is observed with a UV lamp.
Since the fluorescence emission intensity is proportional to the amount of LSD, the concentration of an unknown LSD solution can be determined with a Farrand or Bowman spectrophotometer. After blanking with water at 300 nm, solutions of LSD are scanned with 350 to 250 nm light, and the maximum absorbance, which occurs at approximately 330 nm, is compared to a standard solution of LSD. As little as 0.001 microgram of LSD, or 1/100,000 of a dose, can be analyzed this way.
LSD is among the most fluorescent substances known. According to A. Sperling, LSD is more strongly fluorescent than N,N-DMT, diethyltryptamine, psilocybin, or mescaline. For comparison, DMT absorbs at 280 nm, and fluoresces at 350 nm. Psilocybin absorbs at 270 nm and fluoresces at 340 nm.
It has been reported that LSD loses its fluorescence very rapidly upon strong ultraviolet irradiation, as shown in the decomposition curves below. After just 10-20 seconds of irradiation at 320 nm, the LSD fluorescence reading was significantly diminished.
If UV irradiation is continued for up to 60 minutes, a significant amount of decomposition of LSD can be shown by paper chromatography. Only 10% of LSD remained after 60 min UV irradiation while in the control experiment 90% of LSD remained after 17 h standing in the dark.
A decrease in LSD fluorescence with irradiation occurs when UV light catalyzes the hydration of LSD to a non-fluorescent derivative. A molecule of water added across the C9-C10 double bond of LSD produces the non-fluorescent lumi-derivative. In 1972, Upshall and colleagues described an easy-to-follow procedure for analyzing LSD in human plasma, by measuring the difference in fluorescence (318 nm excitation, 413 nm fluorescence) of plasma extracts before and after intense UV irradiation (at 254 nm). This analytical method using detection of change is greatly preferred to a direct reading of UV fluorescence in plasma, since the plasma blank reading has sufficient enough magnitude to seriously interfere with the determination of LSD. Upshall determined an average of 1-10 ng LSD per 1 mL human plasma in subjects who took a 160 ug dose.
When Aghajanian and colleagues measured LSD concentrations in human plasma in 1964, they found a concentration which corresponded to a level of LSD in the plasma that was higher than expected based on a known injection amount. In this case, the contents of plasma may have added to the native fluorescence of the LSD molecule because the researchers obtained a value of 6-7 ng/mL plasma, about 10X higher than expected. Assessment of the difference in fluorescence of plasma extracts before and after intense UV irradiation is the best way to measure LSD concentrations in human plasma.
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