Synthesis and antimalarial activity of (+)-deoxoartemisinin.

(+)-Deoxoartemisinin (2), a new and more active antimalarial agent, was successfully prepared from artemisinin in one step using NaBH4 and BF3.Et2O in THF. (-)-Deoxodeoxyartemisinin (5), a potential metabolite of deoxoartemisinin, was also prepared either from 2 or from artemisinic acid. 2 shows 8-fold increased antimalarial activity in vitro against chloroquine-resistant malaria as compared to artemisinin (1). Compound 2 possesses superior in vivo antimalarial activity to 1.

7-Caffeoylsedoheptulose from Nyssa sylvatica.

7-Caffeoylsedoheptulose has been isolated from the wood of Nyssa sylvatica, along with two ellagic acid derivatives and scopoletin. Its structure was established by chemical and spectral evidence.

Indolopyridoquinazoline alkaloid from Leptothyrsa sprucei.

The indolopyridoquinazoline alkaloid, 3-hydoxyrutaecarpine, was isolated from Leptothyrsa sprucei, along with 8-methoxypsoralen, 5-methoxypsoralen, imperatorin, isoimperatorin, kaempferol 3-O-alpha-L-rhamnopyranoside, clematine and cnidioside B. The usefulness of the gradient 1H-15N HMBC NMR spectroscopy in the structure elucidation of 3-hydroxyrutaecarpine is noted.

Local anesthetic effects of cocaine and several extracts of the coca leaf (E. coca).

Cocaine and a number of different fractions of a crude ethanol extract of the coca leaf (E. coca) were subjected to a local anesthetic screen using rat tail withdrawal from electric shock. Following an intradermal injection of 0.1 ml of a 2.0% (w.v) solution of cocaine HCl, an immediate response was observed. Two of the coca fractions also produced some local anesthesia. An alkaloidal fraction, containing an equivalent amount of cocaine, produced a maximum effect that was approximately 20% less than that observed with cocaine. The only other fraction producing any effect, a water soluble cocaine-free fraction, showed a maximum response that was approximately 30% of that observed with cocaine.

Comparative lethality of coca and cocaine.

The 24 hour lethal effects of cocaine were compared to those of a crude ethanol extract of the coca leaf (Erthroxylon coca) in male, Swiss mice. Various doses of cocaine HCl and coca leaf extracts suspended in a Tween 60, Arlacel 83, and distilled water vehicle were injected IP into groups of 10 mice. The LD50 for cocaine was 95.1 mg/kg. The LD50 for the coca extract was 3450 mg/kg. The LD50 of the extract based on its cocaine content was 31.4 mg/kg. The results clearly indicate that the coca leaf contains constituents other than cocaine that can contribute to a toxic effect of the plant.

Comparative stimulus properties of two fractions of the coca leaf (E. coca).

Male and female Wistar rats were trained to discriminate 5.0 mg/kg cocaine from 2.0 ml/kg saline using a two-bar food reinforcement (FR 30) drug discrimination paradigm. Once discrimination behavior had stabilized the subjects were tested (in extinction) with several doses of two different fractions of the coca leaf and four doses of cocaine HCl (1.0, 2.5, 7.5, 10 mg/kg). The fractions were prepared by extracting powdered coca leaves with 95% ethanol and then partitioning the residue between chloroform and water. Two doses of the water fractions (480, 960 mg/kg) and five doses of the chloroform fraction (7.5, 15, 30, 60 120 mg/kg) were tested. The water fractions was devoid of cocaine while the five doses of the chloroform fraction contained cocaine equivalent to 0.4, 0.83, 1.65, 3.3 and 6.6 mg/kg, respectively, as determined by gas chromatographic analysis. The 2.5, 7.5, and 10.0 mg/kg cocaine doses generalized to cocaine. The 1.0 mg/kg dose of cocaine generalized to saline. The water fraction at 480 mg/kg generalized to saline; however following pretreatment with the 960 mg/kg dose of this fraction, the animals failed to respond. The two largest doses of the chloroform fraction (60 and 120 mg/kg) generalized to cocaine while the other three doses did not. The 7.5 mg/kg generalized to saline; the 15 and 30 mg/kg doses engendered an intermediate level of responding on both the cocaine and saline lever.

Cross-reactivity of selected compounds in the Abbott TDx cannabinoid assay.

Immunoassay procedures, both enzyme immunoassay and radioimmunoassay, continue to be widely used to screen samples for recent marijuana use by analyzing the urine samples for 11-nor-delta 9-tetrahydrocannabinol-9-carboxylic acid (11-nor-delta 9-THC-9-COOH) (the major urinary metabolite of delta 9-tetrahydrocannabinol [delta 9-THC]). Using commercially available immunoassay reagents, the cross-reactivity of the antiserum utilized in Abbott's TDx cannabinoid assay (a fluorescence polarization immunoassay) was evaluated. This cross-reactivity was evaluated against a group of cannabinoids and noncannabinoid phenolic constituents of Cannabis, some cannabinoid metabolites, and other agents that appear in normal urine samples. In general, the antiserum was equally reactive toward 11-nor-delta 9-tetrahydrocannabinol-9-carboxylic acid, its glucuronide, and the corresponding delta 8-isomer, which was the acid moiety utilized in standards and controls of the assay prior to January, 1990. Reduced binding to the antiserum was observed with hydroxylated derivatives of delta 9- and delta 8-THC, and the other cannabinoids, in general, exhibited limited binding potentials toward the antibody. For the noncannabinoid constituents, no binding was observed at the highest concentrations evaluated (40 mg/L).

Poppy seed ingestion and opiates urinalysis: a closer look.

Review of scientific literature shows that ingestion of poppy seed containing products can result in a positive urinalysis test for opiates. In many cases the amount of seeds ingested is unrealistically high or is not specified. This study is designed to correlate the amount of seeds ingested with the urinary concentration of total morphine as a function of time. Two males and two females were involved in all four protocols, which were separated by at least one week. Subjects ingested one, two, or three poppy seed rolls, each containing 2 g of Australian seeds (108 micrograms morphine/g seed) in three protocols. In the fourth protocol subjects ingested two rolls per day for four consecutive days. Urine specimens were collected for 48 h after ingestion, analyzed by RIA, EMIT, and TDx, and selected samples were confirmed by GC/MS. The data show that the highest concentrations of total morphine in urine were found 3-8 h after ingestion or in the first-void samples. Of the 264 samples collected, there were only 16 specimens that exceeded 300 ng/mL by any of the methods used for analysis with only three samples exceeding 400 ng/mL by GC/MS (406, 611, and 954 ng/mL). In all cases, the total opiates level was less than 150 ng/mL 24 h after ingestion. Following these studies, one of the subjects ingested a poppy seed cake containing 15 g seed obtained from a bakery which analyzed for 169 micrograms morphine/g seed. Urine specimens were collected over 48 h, and all specimens were analyzed by GC/MS.(ABSTRACT TRUNCATED AT 250 WORDS)

Analysis of the major metabolite of delta 9-tetrahydrocannabinol in urine. V. Cross-reactivity of selected compounds in a radioimmunoassay.

The potential for cross-reactivity of an antiserum used to detect 11-nor-delta 9-tetrahydrocannabinol-9-carboxylic acid in a urine sample by a radioimmunoassay procedure has been examined. The degrees of cross-reactivity of 41 cannabinoids and non-cannabinoid phenolic constituents of Cannabis have been determined. In general, only (delta 8- or delta 9-THC type) cannabinoids of the dibenzopyran type structure were found to appreciably cross-react with the antiserum. Cross-reactivity was not observed with any non-cannabinoid constituent.

Analysis of the major metabolite of delta 9-tetrahydrocannabinol in urine II. A HPLC procedure.

A high performance liquid chromatographic (HPLC) procedure was developed for the analysis of 11-nor-delta 9-tetrahydrocannabinol-9-carboxylic acid in urine. Hydrolyzed urine samples were cleaned up using Bond-Elut columns. 11-Nor-cannabinol-9-carboxylic acid was used as internal standard. The analysis was carried out using a C8 reversed-phase column with acetonitrile:50 mM phosphoric acid (65:35) as the solvent. Detection was done by ultraviolet detector at 214 nm. The minimum detectable level of 11-nor-delta 9-tetrahydrocannabinol-9-carboxylic acid was 25 ng/mL.

Analysis of the major metabolite of delta 9-tetrahydrocannabinol in urine. IV. A comparison of five methods.

Urine samples from 29 subjects were simultaneously analyzed for 11-nor-delta 9-tetrahydrocannabinol-9-carboxylic acid by five previously published methods. The procedures employed were radioimmunoassay (RIA), enzyme immunoassay (EIA), high performance liquid chromatography (HPLC), gas chromatography with electron capture detector (GC/ECD), and gas chromatography/mass spectrometry (GC/MS). A comparison of the results showed significant correlations between the results obtained by HPLC, GC/ECD, and GC/MS procedures. As anticipated, quantitation by any of these procedures resulted in lower values than those obtained by either of the immunoassays.

Gas chromatographic/mass spectrometric analysis of morphine and codeine in human urine of poppy seed eaters.

In this study, poppy seeds were examined for a natural constituent that might serve as a maker for the seeds' ingestion as opposed to opiate abuse. Thebaine was selected as possible marker, since it was found to be a component of all poppy seeds examined and was not a natural component of different heroin samples. During the course of this investigation, a new extraction and cleanup procedure was developed for the gas chromatographic/nitrogen phosphorus detection (GC/NPD) and gas chromatographic/mass spectrometric (GC/MS) analysis of morphine and codeine in urine. A linear response, over a concentration range of 25 to 600 ng/mL, was obtained for codeine and morphine (r = 0.9982 and 0.9947, respectively). The minimum detectable level (LOD) and limit of quantitation (LOQ) for morphine were 10 and 30 ng/mL, respectively; whereas LOD and LOQ for codeine were 2 and 8 ng/mL, respectively. The coefficients of variance (CV, n = 6) for morphine and codeine analyses at the 100-ng/mL level were 13.3 and 4.6%, respectively. This procedure was used for the analysis of urine samples from five poppy seed eaters who each ingested 200 g of poppy seed cake. Results indicated that significant amounts of morphine and codeine are excreted in urine and that in all subjects, at least at one point in time, the apparent morphine concentration as determined by radioimmunoassay (RIA) analysis exceeded the cutoff value (300 ng/mL) established for screening. Thebaine was not detected in urine specimens collected following poppy seeds ingestion and thus could not be used as a marker.

Constituents of Cannabis sativa L. XXII: isolation of spiro-indan and dihydrostilbene compounds from a Panamanian variant grown in Mississippi, United States of America.

Three spiro-compounds, namely cannabispiran, dehydrocannabispiran and beta-cannabispiranol, and 2 dihydrostilbenes [3-(2-(3-hydroxy-4-methoxyphenyl)ethyl)-5-methoxyphenol and canniprene] were isolated from a polar fraction of a Panamanian variant of Cannabis sativa L. grown in Mississippi, United States of America. The plant material was extracted with 95% ethanol and the dried ethanol extract was then partitioned between chloroform and water. The chloroform fraction was fractionated between hexane and 3N sodium hydroxide solution. Acidification of the basic fraction followed by extraction with ether afforded a polar acidic fraction from which the above-mentioned compounds were isolated through repeated chromatography. The structures of the above compounds were determined by spectral means as well as by comparison with reference samples. The isolation of two dihydrostilbenes and three spiro-indan compounds from a single variant provides good support that the dihydrostilbenes are the natural precursors to the spiro-indan compounds.

Microbial metabolism studies of the antimalarial drug arteether.

Microbial metabolism studies of the antimalarial drug arteether (1) have shown that arteether is metabolized by a number of microorganisms. Large-scale fermentation with Aspergillus niger (ATCC 10549) and Nocardia corallina (ATCC 19070) have resulted in the isolation of four microbial metabolites which have been characterized using two-dimensional nuclear magnetic resonance (2D-NMR) techniques. These metabolites have been identified as "AEM1" (2), 3 alpha-hydroxydeoxyarteether (3), 3 alpha-hydroxydeoxydihydroartemisinin (4), and deoxydihydroartemisinin (5).

The effect of subacute marijuana smoke inhalation on experimentally induced dermonecrosis by S. aureus infection.

Dermonecrosis was induced in ICR mice by subcutaneous implantation of Staphylococcus aureus absorbed onto sterile cotton pellets. This model was used to assess the effects of marijuana smoke, marijuana placebo smoke and delta 9-tetrahydrocannabinol (delta 9-THC) on the local immune response to bacterial infection. Mice were exposed to 40 or 80 "puffs" of marijuana smoke, marijuana placebo smoke or air daily for 4 consecutive days. The estimated dose of delta 9THC per day generated from 40 or 80 puffs of marijuana smoke was 3.2 and 6.4 mg/kg, respectively. A group of sentinel (Shelf) control mice were included in each experiment. The necrotic index (NI) of mice exposed to 40 or 80 puffs of marijuana smoke were 67% and 44% of control, respectively. Air exposed mice showed a necrotic index comparable to the shelf control group. In chronically (60 days) exposed mice (80 puffs per day) the necrotic index was about 12% of control, while air-exposed mice were about 40% of control. Placebo marijuana smoke exposed mice had a NI comparable to that of marijuana smoke exposed mice which suggested that the reduction in NI was unrelated to the pychomimetic component delta 9THC. To further explore which of the constituents of marijuana were responsible for the decreased NI, the ethanol extract from marijuana leaves was partioned between water (cannabinoid free) and chloroform (cannabinoid rich). Injection of the cannabinoid free fraction produced comparable decrease in the NI as observed with whole marijuana smoke, while the cannabinoid rich fraction produced no effect. delta 9THC at a dose of 10 mg/kg per day did not alter the NI.

Microbial metabolism studies of the antimalarial sesquiterpene artemisinin.

Microbial metabolism of the sesquiterpene lactone antimalarial drug artemisinin [1] was studied. Screening studies have shown a number of microorganisms capable of metabolizing artemisinin [1]. Scale-up fermentation with Nocardia corallina (ATCC 19070) and Penicillium chrysogenum (ATCC 9480) have resulted in the production of two major microbial metabolites that have been characterized with the use of 2D-nmr techniques. These metabolites have been identified as deoxyartemisinin [2] and 3 alpha-hydroxydeoxyartemisinin [3].