Abstract

Crocus sativus stigmas are one of the widely known spices (saffron) and consist of unusually polar carotenoids.

Alzheimer’s disease is characterized pathologically by deposition of amyloid beta-peptide (Abeta) fibrils.

Oxidation is thought to promote Abeta fibril formation and deposition.

To identify agents inhibiting the pathogenesis of Alzheimer’s disease, we examined in vitro the antioxidant properties of extract of C. sativus stigmas and its effect on Abeta(1-40) fibrillogenesis.

The antioxidant properties were determined by measuring the ferric-reducing antioxidant power and Trolox-equivalent antioxidant capacity, while its effects on Abeta-aggregation and fibrillogenesis were studied by thioflavine T-based fluorescence assay and by DNA binding shift assay.

The water:methanol (50:50, v/v) extract of C. sativus stigmas possesses good antioxidant properties, higher than those of tomatoes and carrots, and inhibited Abeta fibrillogenesis in a concentration and time-dependent manner.

The main carotenoid constituent, trans-crocin-4, the digentibiosyl ester of crocetin, inhibited Abeta fibrillogenesis at lower concentrations than dimethylcrocetin, revealing that the action of the carotenoid is enhanced by the presence of the sugars.

Our findings suggest the possible use of C. sativus stigma constituents for inhibition of aggregation and deposition of Abeta in the human brain.

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Inhibitory Activity on Amyloid-Beta Aggregation and Antioxidant Properties of Crocus Sativus Stigmas Extract and Its Crocin Constituents

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Abstract

Crocus sativus L. is a plant cultivated in various parts of the world. Crocins are among the active components of Crocus sativus L.

The present study was designed to investigate in the rat whether or not crocins possess anxiolytic properties.

For this aim, the light/dark test was selected.

Either crocins, at a dose which did not influence animals’ motor activity (50mg/kg), or diazepam (1.5 mg/kg), significantly increased the latency to enter the dark compartment and prolonged the time spent in the lit chamber in the rats.

Conversely, lower doses of crocins (15-30 mg/kg) did not substantially modify animals’ behaviour.

The present results indicate that treatment with these active constituents of Crocus sativus L. induce anxiolytic-like effects in the rat.

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Effects of the active constituents of Crocus sativus L., crocins, in an animal model of anxiety

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Abstract

Rationale:

The expensive spice saffron originating from the stigmas of Crocus sativus L. and also applied in traditional Chinese medicine (TCM) constitutes a complex mixture of glycoconjugates varying not only in the aglycon structure, but also in glycosylation pattern. Therefore, various tandem mass spectrometric techniques were evaluated for their usefulness in structural elucidation.

Methods:

Three selected constituents of the stigmas of Crocus sativus L., trans- and cis-crocin-4 as well as picrocrocin, were isolated and purified by HPLC and finally analyzed by ESI-MS (ion trap, QqRTOF), IP-MALDI-MS (QqRTOF) and vMALDI-MS (TOF/RTOF) in combination with tandem mass spectrometry in collision energy regimes ranging from a few eV (LE) to 20 keV (HE) collisions for the first time. These data aid in structurally elucidating minor, unknown glycoconjugates originating from this plant-derived spice.

Results:

LE-CID of isomeric crocins on either an ion trap with ESI or a QqRTOF-instrument with ESI or IP-MALDI as desorption/ionization technique only yielded a limited number of structurally diagnostic sodiated product ions related to the carbohydrate moiety as well as to the intact aglycon in contrast to true HE-CID. The low MW constituent picrocrocin did not yield useful LE-CID spectra, but showed a high number of structurally diagnostic product ions by HE-CID utilizing a vMALDI TOF/RTOF-instrument.

Conclusions:

The highest number of structurally diagnostic product ions allowing also determination of the carbohydrate linkage of the gentiobiose-moiety of isomeric crocins ((0,4)A(2), (3,5)A(2) product ions indicating a 1→6 carbohydrate linkage) was only achievable by HE-CID. Fragmentation of the aglycon was not observed by any collision energy regime applied.

Comparison of Different Tandem Mass Spectrometric Techniques (ESI-IT, ESI- And IP-MALDI-QRTOF and vMALDI-TOF/RTOF) for the Analysis of Crocins and Picrocrocin From the Stigmas of Crocus Sativus L

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Abstract

Experimental evidence indicates that the non-competitive N-methyl-D-aspartate (NMDA) receptor antagonist ketamine impairs cognition and can mimic certain aspects of positive and negative symptoms of schizophrenia in rodents.

Crocins are among the active components of the plant Crocus sativus L. and were found to be effective in different models of psychiatric disorders including anxiety and depression.

The present study was designed to investigate the ability of crocins to counteract schizophrenia-like behavioural deficits produced by ketamine in rats.

Crocin’s ability to counteract hypermotility, stereotypies and ataxia induced by ketamine was evaluated in a motor activity cage.

The ability of crocins to reverse ketamine-induced memory deficits was assessed using the novel object recognition task (NORT).

The social interaction test was used in order to examine the effects of crocins on ketamine-induced social withdrawal. Crocins (50 but not 30 mg/kg, i.p.) attenuated ketamine (25 mg/kg, i.p.)-induced hypermotility, stereotypies and ataxia.

In a subsequent study, post-training administration of crocins (15 and 30 mg/kg, i.p.) reversed ketamine (3 mg/kg, i.p.)-induced performance deficits in the NORT. Finally, crocins (50 but not 30 mg/kg, i.p.) counteracted the ketamine (8 mg/kg, i.p.)-induced social isolation in the social interaction test.

Our findings show that crocins attenuated schizophrenia-like behavioural deficits induced by the non-competitive NMDA receptor antagonist ketamine in rats.

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Crocins, the active constituents of Crocus Sativus L., counteracted ketamine-induced behavioural deficits in rats

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Abstract

The stigmas of Crocus sativus L. have been used as spice and colorant agent (i.e. saffron) for more than 4000 years.

For an updated structural investigation of the aglycon present in the glycosylated crocetin apocarotenoids (i.e. crocins), seven representative derivatives ranging from one up to five glucosyl-residues with a maximum number of three monosaccharides per glycosylation site (glucose, gentiobiose, gentiotriose and neapolitanose) were isolated and purified by high-performance liquid chromatography.

The compounds selected for further mass spectrometric investigation include glucosyl-, bis-glucosyl-, gentiobiosyl-, gentiobiosyl-glucosyl-, bis-gentiobiosyl-, gentiobiosyl-gentiotriosyl- and gentiobiosyl-neapolitanosyl-crocetin.

Electrospray ionization in combination with low-energy collision-induced dissociation/tandem mass spectrometry of sodiated crocin precursor ions utilizing either a 3D-ion trap (MS(n) , n = 2, 3) or a QqTOF instrument, with the latter providing accurate mass determination with an accuracy of ±1-3 ppm or better at a resolution of 10,000 (full width at half maximum), was used.

Major fragmentation pathways included loss of either one or two carbohydrate substituents leading to the sodiated aglycon without interglycosidic bond cleavage during in MS(2) -experiments.

All sodiated precursor ions and major product ions were accompanied by a loss of 92 Da, which was elucidated as C7 H8 -loss from the aglycon by skeletal rearrangement via an eight-membered transition state as previously described for intact C40-carotenoids.

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In-chain Neutral Hydrocarbon Loss From Crocin Apocarotenoid Ester Glycosides and the Crocetin Aglycon (Crocus Sativus L.) by ESI-MS(n) (n=2, 3)

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Abstract

A chromatographic method was developed and fully validated for the determination of the major saffron constituents, i.e., picrocrocin and five major crocins.

Dried samples (styles of Crocus sativus and other Crocus taxa) were extracted with MeOH : water (1 : 1, v/v), and chromatographic separation of the analytes was achieved by reversed-phase chromatography using a gradient elution.

Full validation was performed using spiked samples with analytes, which were isolated, purified, and characterized by MS due to a lack of commercial standards.

The method showed a good fit (r2 > 0.999) for all analytes with limit of quantitation values in the range of 1-15 µg/mL, and demonstrated adequate intra- and inter-precision (< 15 % RSD) and accuracy (< 7 % RE).

The method was applied to the analysis of various commercial saffron samples and of indigenous Crocus taxa and allowed for the first time the absolute quantitation of several Crocus components.

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Quantitation of Crocins and Picrocrocin in Saffron by HPLC: Application to Quality Control and Phytochemical Differentiation From Other Crocus Taxa

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Abstract

In this study, the aphrodisiac activities of Crocus sativus stigma aqueous extract and its constituents, safranal and crocin, were evaluated in male rats.

The aqueous extract (80, 160 and 320mg/kg body wt.), crocin (100, 200 and 400mg/kg body wt.), safranal (0.1, 0.2 and 0.4ml/kg), sildenafil (60mg/kg body wt., as a positive control) and saline were administered intraperitoneally to male rats.

Mounting frequency (MF), intromission frequency (IF), erection frequency (EF), mount latency (ML), intromission latency (IL) and ejaculation latency (EL) were the factors evaluated during the sexual behavior study.

Crocin, at all doses, and the extract, especially at doses 160 and 320mg/kg body wt., increased MF, IF and EF behaviors and reduced EL, IL and ML parameters.

Safranal did not show aphrodisiac effects.

The present study reveals an aphrodisiac activity of saffron aqueous extract and its constituent crocin.

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The Effect of Saffron, Crocus Sativus Stigma, Extract and Its Constituents, Safranal and Crocin on Sexual Behaviors in Normal Male Rats

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Abstract

Crocus sativus L., is a plant cultivated in many countries of the world.

Crocins are among the active constituents of C. sativus and their implication in cognition has been proposed.

The present study was designed to investigate in the rat the effects of crocins on distinct recognition memory components (encoding, storage and retrieval). Subsequently, the potential use of crocins as adjunctive agents for the treatment of memory disorders was examined.

Thus, the effects exerted by a combination of subthreshold doses of crocins and memantine on recognition memory were evaluated.

To assess the effects of compounds on memory, the novel object-recognition task (NORT) was used. In a preliminary study, the influence of the retention time (the delay between the two trials) on the performance of rats was assessed.

Rats’ recognition memory abilities remained intact up to 6 h, but were extinguished when a delay of 24 h was utilized. Crocins, at any dose tested (5, 15, and 30 mg/kg), did not affect rats’ performance, whereas administration of higher doses (15 and 30 mg/kg) reversed delay-dependent deficits in the NORT.

The combination of subthreshold doses of crocins (5 mg/kg) and memantine (3 mg/kg) did not influence the performance, but counteracted delay-dependent deficits in the NORT.

These findings suggest that crocins counteract natural forgetting and may modulate different aspects of recognition memory, and that the combined use of crocins and memantine might represent a novel strategy to treat memory disorders.

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Effects of the Active Constituents of Crocus Sativus L. Crocins and Their Combination With Memantine on Recognition Memory in Rats

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