Determination of Residual Citrinin in Zhibituo Tablets by Solid Phase Extraction-UPLC-MS/MS / 中国药房

OBJECTIVE:To determine residual citrinin in Zhibituo tablet by solid phase extraction-UPLC-MS/MS. METH-ODS:The sample was processed preliminary by solid extraction. The determination was performed on Zorbax Eclipse Plus C18 col-umn with mobile phase consisting of water (0.1% formic acid)-acetonitrile (70:30,V/V,gradient elution) at the follow rate of 0.30 mL/min. The column temperature was 40 ℃,and analysis time was 7 min,the sample size was 2 μL. Ionization mode was positive ion mode. The ion source temperature was 150 ℃,drying gas velocity was 15 L/min,sheath gas temperature was 350 ℃, sheath gas flow rate was 12 L/min,capillary voltage was 3500 V,atomization device pressure was 40 psi,atomization voltage was 0 V. The acquisition mode was MRM mode. RESULTS:The linear range of citrinin was 0.1-20 ng/mL(r=0.9994);the limits of quantitation and detection were 0.05 and 0.01 ng/mL. RSDs of precision,stability and repeatability tests were all lower than 2.0%;recoveries were 98.868%-103.160%(RSD=1.5%,n=6). CONCLUSIONS:The method is rapid,sensitive,accurate and suitable for the determination of residual citrinin in Zhibituo tablet.

Discussion on quality standards of lipid-reducing Monascus product / 中草药

The status related to the quality standards of lipid-reducing Monascus product was introduced in this paper. The contents of several existing standards involved with lipid-reducing Monascus product in China were mainly discussed. Some of the standards lack inclusiveness and rationality, which makes too specific regulations on the fermentation strains and raw materials. Some of the standards lack scientificity in structure and test method of monacolin K. In some standards, the restricted concentration of citrinin is too harsh, which is out of the sensitivity of the existing instrument and also lacks operability. It is suggested that more scientific and reasonable standards should be established to promote the development of lipid-reducing Monascus product.

Inhibitory effect of various aqueous medicinal plant extracts on citrinin production and fungal biomass by Penicillium notatum and Aspergillus niger.

Citrinin is a nephrotoxic mycotoxin produced by several species of the genus Aspergillus, Penicillium and Monascus. Citrinin is generally formed after harvest and occurs mainly in stored grains The efficacy of different concentrations of aqueous leaf extracts of Aervalanata, Nerium oleander Rhazya stricta Decne and Cleome amblyocarpa (5 to 10 mg/mL) on growth and citrinin production in two fungal strains Penicillium notatum and Aspergillus niger was investigated. Mycotoxin production and fungal biomass by the isolates was suppressed, depending on the concentration of the plant extract added to culture media at the time of spore inoculation. Citrinin production in fungal mycelia grown for 15 days in culture media containing 5-10 mg/mL of the aqueous extracts of A. lanata, N. oleander, R. stricta Decne and C. amblyocarpa showed inhibition of approximately 14.2 to 91.8 % in Penicillium notatum and 13.4 to 90.3% in Aspergillus niger. Among the all four extracts Rhazya stricta Decne was more efficient than other tested plant extracts in inhibiting the citrinin production ranging from 22.4 to 91.8% in P. notatum and 32.6 to 93.2% in A. niger.

Effect of feeding graded doses of Citrinin on clinical and teratology in female Wistar rats.

Citrinin is the one of the well-known mycotoxins, which is possibly spread all over the world. The graded doses of citrinin (1, 3 and 5 ppm CIT in feed) in female Wistar rats 10 weeks prior to mating, during mating and during organogenesis resulted in resorptions and post implantation losses, decreased fetal body weights and crown-rump lengths in fetuses of all groups. Various developmental anomalies recorded in fetuses of treated rats included gross (wrist drop, curled tail, stretched forelimb, subcutaneous haematoma), skeletal (incomplete ossification of skull bones, incomplete fusion of vertebral bodies, complete and partial agenesis of sternaebrae, metacarpals, metatarsals and phalanges, fused ribs and swing out ribs) and visceral (internal and external hydrocephalus, cerebellar hypoplasia, microphthalmia, roundening of heart, contracted kidneys, dilated renal pelvis and cryptorchid testes). The results suggest that CIT has adverse effects on fetal development which may be due to the longer bioavailability of citrinin in the animals.

Cytotoxic and antibacterial substances against multi-drug resistant pathogens from marine sponge symbiont：Citrinin, a secondary metabolite of Penicillium sp

Objective: To Isolate, purify, characterize, and evaluate the bioactive compounds from the sponge-derived fungus Penicillium sp. FF001 and to elucidate its structure. Methods: The fungal strain FF001 with an interesting bioactivity profile was isolated from a marine Fijian sponge Melophlus sp. Based on conidiophores aggregation, conidia development and mycelia morphological characteristics, the isolate FF001 was classically identified as a Penicillium sp. The bioactive compound was identified using various spectral analysis of UV, high resolution electrospray ionization mass spectra, 1H and 13C NMR spectral data. Further minimum inhibitory concentrations (MICs) assay and brine shrimp cytotoxicity assay were also carried out to evaluate the biological properties of the purified compound. Results: Bioassay guided fractionation of the EtOAc extract of a static culture of this Penicillium sp. by different chromatographic methods led the isolation of an antibacterial, anticryptococcal and cytotoxic active compound, which was identified as citrinin (1). Further, citrinin (1) is reported for its potent antibacterial activity against methicillin-resistant Staphylococcus aureus (S. aureus), rifampicin-resistant S. aureus, wild type S. aureus and vancomycin-resistant Enterococcus faecium showed MICs of 3.90, 0.97, 1.95 and 7.81 μg/mL, respectively. Further citrinin (1) displayed significant activity against the pathogenic yeast Cryptococcus neoformans (MIC 3.90 μg/mL), and exhibited cytotoxicity against brine shrimp larvae LD50 of 96 μg/mL. Conclusions: Citrinin (1) is reported from sponge associated Penicillium sp. from this study and for its strong antibacterial activity against multi-drug resistant human pathogens including cytotoxicity against brine shrimp larvae, which indicated that sponge associated Penicillium spp. are promising sources of natural bioactive metabolites.

Cytotoxic and antibacterial substances against multi-drug resistant pathogens from marine sponge symbiont: Citrinin, a secondary metabolite of Penicillium sp

<p><b>OBJECTIVE</b>To Isolate, purify, characterize, and evaluate the bioactive compounds from the sponge-derived fungus Penicillium sp. FF001 and to elucidate its structure.</p><p><b>METHODS</b>The fungal strain FF001 with an interesting bioactivity profile was isolated from a marine Fijian sponge Melophlus sp. Based on conidiophores aggregation, conidia development and mycelia morphological characteristics, the isolate FF001 was classically identified as a Penicillium sp. The bioactive compound was identified using various spectral analysis of UV, high resolution electrospray ionization mass spectra, 1H and 13C NMR spectral data. Further minimum inhibitory concentrations (MICs) assay and brine shrimp cytotoxicity assay were also carried out to evaluate the biological properties of the purified compound.</p><p><b>RESULTS</b>Bioassay guided fractionation of the EtOAc extract of a static culture of this Penicillium sp. by different chromatographic methods led the isolation of an antibacterial, anticryptococcal and cytotoxic active compound, which was identified as citrinin (1). Further, citrinin (1) is reported for its potent antibacterial activity against methicillin-resistant Staphylococcus aureus (S. aureus), rifampicin-resistant S. aureus, wild type S. aureus and vancomycin-resistant Enterococcus faecium showed MICs of 3.90, 0.97, 1.95 and 7.81 µg/mL, respectively. Further citrinin (1) displayed significant activity against the pathogenic yeast Cryptococcus neoformans (MIC 3.90 µg/mL), and exhibited cytotoxicity against brine shrimp larvae LD50 of 96 µg/mL.</p><p><b>CONCLUSIONS</b>Citrinin (1) is reported from sponge associated Penicillium sp. from this study and for its strong antibacterial activity against multi-drug resistant human pathogens including cytotoxicity against brine shrimp larvae, which indicated that sponge associated Penicillium spp. are promising sources of natural bioactive metabolites.</p>

Review on Citrinin: Synthetic Methods, Molecular Biosynthesis and Effect of Plant Extracts.

Introduction: Citrinin is a mycotoxin originally isolated from Penicillium citrinum. It has been found to be produced by a variety of other fungi (Aspergillus niveus, Aspergillus ochraceus, Aspergillus oryzae, Aspergillus terreus, Monascus ruber, Monascus purpureus and Penicillium camemberti) which are found or used in the production of human foods (Abramson et al., 1999). The inhibitory effect of plant extracts on citrinin biosynthesis have been examined (Mossini and Kemmelmeier, 2008; Reddy et al., 2010). They found that all the tested plant extracts reduced the citrinin production. Shimizu et al. (2005, 2007) found that the pksCT gene was essential for citrinin biosynthesis in M. purpureus. Also Sakai et al. (2008) reported that introducing additional copies of an activator gene (ctnA), controlled by the Aspergillus nidulans trpC promoter, into the citrinin-cluster-containing transformants enhanced the transcription of all the genes in the cluster and resulted in an almost 400-fold higher citrinin production compared to that of the parental transformant. Aims: To give idea on physicochemical properties of citrinin, its production, effects of some plant extracts on it and gene involved in citrinin biosynthesis. Study Design: Review study. Place and Duration of Study: Department of Biology, Faculty of Science, Taif- Saudi Arabia and Department of Botany, Faculty of Science, Tanta-Egypt. 2011-2012. Methodology: Citrinin was produced in liquid potato-dextrose medium (PD) or in glucose medium. The citrinin was extracted three times with chloroform (1:1 v/v), pooled and concentrated in vacuo at 40ºC using a rotary evaporator. The crude extract was diluted in minimum amount of chloroform (2 ml) and citrinin was estimated by thin layer chromatography (TLC). Effects of some plant extracts like neem leaf extract and some medicinal plants were determined. Conclusion: This review was written with the aim of demonstrating the scope of citrinin production, various analytical techniques in citrinin detection and estimation and effects of some plant extracts and genes on citrinin biosynthesis. It was found that plant extracts can be used as a potential source of sustainable ecofriendly botanical fungicides to protect food grains from toxigenic P. citrinum and citrinin accumulation under storage conditions.

Antitumor metabolites produced by two Penicillium purpurogenum G59 mutants / 国际药学研究杂志

Objective: To investigate the antitumor metabolites of fungal mutants 2-2-3 and PDN-f-2, the two bioactive mutants of Penicillium purpurogenum G59 that do not produce antitumor metabolites. Methods: Bioactive metabolites newly produced by the mutants were isolated by a bioassay-guided separation procedure using iquid-liquid extraction, column chromatography and recrystallization methods through direct comparison with the sample from P. purpurogenum G59. The compounds obtained were identified by spectroscopic methods. The antitumor activity was assayed by the MTT method using K562 cells. Results: Two bioactive metabolites 1 and 2 were isolated from the fermentation products of 2-2-3 and PDN-f-2, respectively, and identified as ergone (1) and citrinin (2). Compounds 1 and 2 inhibited the proliferation of K562 cells with the IC50 values of 7. 4 and 48. 0 μg/ml, respectively. Conclusion: Compounds 1 and 2 are the antitumor metabolites newly produced by he mutants 2-2-3 and PDN-f-2, respectively, and have not been found in the metabolites of P. purpurogenum so far. It is revealed from the present result that the alteration of secondary metabolism of wild-type fungal strains without bioactivity for obtaining bioactive metabolite-producing mutants may become a new route to expand the source of new fungal strains for drug screening.

The Strategy of Citrinin Production Control in Monascus / 中国生物工程杂志

Pepole pay more attention to the medicinal value and health function of Monascus day by day,however,the existence of citrinin restricted the further development of Monascus products heavily. How to ruduce the content of citrinin is a problem need to be solved urgently.Briefly introduced the toxicity,the biosynthetic pathway,and the relative standards of citrinin in monascus. According to the research progress on citrinin,the strategies of citrinin control were described from the three aspects of fermentation technology,mutation breeding,and genetic engineering. The expectation about the direction of citrinin in the future was also discussed.

Screening Citrinin Mutants from the Transformants Library of Monascus ruber M-7 by Agrobacterium-mediated DNA transfer / 微生物学通报

200 citrinin mutants were screened with the inhibition zone method from the transformants library of Monascus ruber M-7 by Agrobacterium-mediate DNA transfer, which contains more than 5,000 transformants.Then 53 mutants, whose citrinin contents ranged from 0.04?g/g to 154.57?g/g in the red fermented rice (RFR), were achieved by high performance liquid chromatography (HPLC).Color values of RFR prepared by these mutants were also detected.The results showed that there was a positive correlation between the citrinin content and the color value among the mutants.These results provide materials and research bases for ferrther studying the relationship between the production of citrinin and pigment of Monascus ruber at molecular level.

Monascus Citrinin Analysis Methods and A Study on Formation of Citrillin by Monascus / 微生物学通报

A HPLC method of analysis of Monascus citrinin was established. More than 30 strains of Monascus spp. were cultured in steamed rice at solid state or in MSG liquid medium composed of monosodium glutamate as sole nitrogen source and glucose as sole carbon to investigate their ability of producing citrinin. The results indicated that most of the Monascus strains are able to produce citrinin. MSG medium can be used as a specific culture medium to qualitatively identify if the strain is the potential citrinin producer. But to confirm whether the Monascus strains are potential citrinin producers, these strains should be cultured in several cultivation methods, as the culture states and culture conditions influence the citrinin production greatly.