Monascus is one of the most essential microbial resources in China, with thousands of years of history. Modern science has proved that Monascus can produce pigment, ergosterol, monacolin K, γ-aminobutyric acid, and other functionally active substances. Currently, Monascus is used to produce a variety of foods, health products, and pharmaceuticals, and its pigments are widely used as food additives. However, Monascus also makes a harmful polyketide component called citrinin in the fermentation process; citrinin has toxic effects on the kidneys such as teratogenicity, carcinogenicity, and mutagenicity (Gong et al., 2019). The presence of citrinin renders Monascus and its products potentially hazardous, which has led many countries to set limits and standards on citrinin content. For example, the citrinin limit is less than 0.04 mg/kg according to the Chinese document National Standard for Food Safety Food Additive Monascus (GB 1886.181-2016) (National Health and Family Planning Commission of the People's Republic of China, 2016), and the maximum level in food supplements based on rice fermented with Monascus purpureus is 100 µg/kg in the European Union (Commission of the European Union, 2019).
Citrinin ; Dietary Supplements ; Fungi ; Monascus

Trace chemical constituents from the ethyl acetate extract of Red Yeast Rice were investigated. Four phenolic compounds were isolated by various column chromatographies, and their structures were identified on the basis of spectroscopic analysis including UV, MS, IR and NMR. The four compounds were identified as 2-methyl-5-(2'R-methyl-4'-hydroxy-butyl)-cinnamic acid(1), 5-(2'-hydroxy-6'-methyl phenyl)-3-methylfuran-2-carboxylic acid(2), daidzein(3), and genistein(4). Compound 1 was new and 2 was firstly discovered from the genus Monascus, while 3-4 were obtained from Red Yeast Rice for the first time.
Biological Products ; chemistry ; Magnetic Resonance Spectroscopy ; Monascus ; Phenols ; chemistry

The anti-obesity effects of fermented black bean were tested with mice fed a high fat diet for seven weeks. Body weight gain and feed efficiency ratio (FER) in the high fat diet control (HC) group were markedly higher, compared with those of the normal control (NC) group, but were significantly lower in the 2% black bean powder supplemented high fat diet (BB) group and 2% black bean powder fermented by M. pilosus supplemented high fat diet (BBM) group, compared with those of the HC group. Food intake in the HC and BB groups was significantly lower than that of the NC and BBM groups. Water intake in the HC group was significantly lower than that of the NC group, but was higher in the BB and BBM groups, compared with that of the HC group. On the other hand, relative liver and kidney weight in the HC group was lower than that of the NC group, but was higher in the BB and BBM groups, compared with that of the HC group. In addition, whereas epididymal fat weight in the HC group was markedly higher than that of the NC group, it was significantly lower in the BB and BBM groups, compared with that of the HC group. Meanwhile, hepatic GSH in the HC group was significantly lower than that of the NC group, but was slightly higher in the BB and BBM groups, compared with that of the HC group. Although hepatic LPO in the HC group was dramatically higher than that of the NC group, it was significantly lower in the BB and BBM groups, compared with that of the HC group. In addition, serum TG, total cholesterol, and LDL-cholesterol in the HC group was significantly higher than that of the NC group, but was significantly lower in the BB and BBM groups, compared with that of the HC group. On the contrary, HDL-cholesterol in the HC group was significantly lower than that of the NC group, but was higher in the BB and BBM groups, compared with that of the HC group. In addition, activity of XOR D type in the HC group was lower than that of the NC group, but was slightly higher in the BB and BBM groups, compared with that of the NC group. Activities of ROS scavenging enzymes, such as SOD, GPX, and GST in the HC group were significantly lower than those of the NC group, but were significantly higher in the BB and BBM groups, compared with those of the HC group. In addition, serum ALT activity in the HC and BB groups was higher than that of the NC group, but was significantly lower in the BB and BBM groups, compared with that of the HC group. In histopathological findings, hepatic fat accumulation in the HC group was higher than that of the NC group, but was lower in the BBM group, compared with that of the HC and BB groups. In particular, antiobese, hypolipidemic, and antifatty liver effect of black bean powder fermented by M. pilosus was specifically higher than that of non-fermented steamed black bean. In conclusion, the constituents of black bean fermented by Monascus pilosus have been proven to not only inhibit obesity and hyperlipidemia but also decrease hepatic fat accumulation in high fat diet-induced obese mice.
Animals ; Body Weight ; Cholesterol ; Diet, High-Fat ; Drinking ; Eating ; Hand ; Hyperlipidemias ; Kidney ; Liver ; Mice ; Mice, Obese ; Monascus ; Obesity ; Steam

OBJECTIVETo establish a method to test citrinin (CIT) in monascus products by immuno-affinity chromatography (IAC)-high performance liquid chromatography (HPLC), and to detect the content of CIT in monascus products in Fujian province.

METHODSIAC-HPLC was applied to detect the CIT content in monascus products. The conditions to use HPLC were as follows: C(18) reversed-phase chromatographic column, 150.0 mm×4.6mm×3 µm; mobile phase: the volume ratio of acetonitrile and 0.1% phosphoric acid solution at 65:35; isocratic elution; column temperature: 28°C; flow velocity: 0.8 ml/min; fluorescence detector, excitation wavelength (λ(ex)) was 331 nm and emission wavelength (λ(em)) was 500 nm. The standard curved was established by the linear regression of peak area (Y) to CIT content (X, ng/ml). The accuracy and precision of the method would then be verified. And 32 kinds of monascus products were determined and their color values were compared by this method.

RESULTThe standard curve established in this study was Y = 4634.8X-136.42, r = 1.000; whose limits of detection was 20 µg/kg and the limits of qualification was 64 µg/kg. In the range between 200 and 800 µg/kg, the standard recovery rate was 98.9% - 110.0% (n = 3), and the relative standard deviation (RSD) was 0.51% - 1.76%. Out of the 32 samples, CIT was detected from 11 samples of monascus rice, 9 samples of monascus powder and 5 samples of monascus pigments, the content was around 0.212 - 14.500 mg/kg. 4 out of 7 functional monascus samples were detected out CIT, whose content at 0.142 - 0.275 mg/kg.

CONCLUSIONThe method to detect CIT in monascus products by IAC-HPLC has been established.

Chromatography, Affinity ; methods ; Chromatography, High Pressure Liquid ; methods ; Citrinin ; analysis ; Drug Contamination ; Monascus

OBJECTIVETo study the chemical constituents of Monascus purpureus metabolite.

METHODThe compounds were isolated by column chromatography methods, and their structures were determined by spectroscopic methods.

RESULTEight compounds were isolated from the petroleum ether fraction of ethanolic extract and elucidated as stigmast-4-en-3-one (1), 3-oxo-24-methylenecycloarane (2), stigmasterol (3), 7beta-hydroxystigmasterol (4), 3beta-hydroxystigmast-5-en-7-one (5), 3beta-hydroxystigmast-5,22-dien-7-one (6), 5alpha, 8alpha-epidioxyergosta-6,22-dien-3beta-ol (7), sitosterol (8).

CONCLUSIONAll of the compounds were isolated from this genu for the first time except compound 3 and 7.

Drugs, Chinese Herbal ; chemistry ; isolation & purification ; metabolism ; Monascus ; chemistry ; metabolism ; Steroids ; chemistry ; isolation & purification ; metabolism

OBJECTIVETo establish an effective way for rapid identification of Monascus strains based on DNA molecular marker.

METHODA random amplified polymorphic DNA (RAPD) marker named F421 in genomic DNA of Monascus F strain was observed during a comparison of DNA fingerprints derived from 10 cultivated strains of Monascus. F421 was cloned and sequenced. Comparing the sequence of F421 (GenBank accession number EF063107) with other relative sequences in the GenBank databases, no distinct comparability was found. A pair of sequence characterized amplified region (SCAR) primers were designed based on the sequence of the cloned fragment and tested for the specific detection of Monascus F.

RESULTThe results of polymerase chain reaction showed that only a 421bp segment of Monascus F strain was amplified compared with other 9 cultivated strains of Monascus. And the acquired SCAR marker of strain F could be used as a specific DNA fingerprint to identify Monascus strain F within one day.

CONCLUSIONSCAR molecular marker technology is an effective new way to identify Monascus strains more rapidly. And also is an assistant tool to identify Monascus strains more accurately when disagreements come out using traditional classification. It could be applied widely to the protection of germ plasm resources, classification and identification distinguishing false strains of pharmaceutical fungi.

Molecular Sequence Data ; Monascus ; classification ; genetics ; Polymerase Chain Reaction ; methods ; Random Amplified Polymorphic DNA Technique

OBJECTIVETo obtain the full-length cDNA of a novel gene (named yp05) associated with citrinin production-related genes in Monascus aurantiacus.

METHODSTotal RNA was extracted from mycelium, 3' and 5' cDNA end of yp05 gene was amplified using smart trace cDNA amplification kit, and the full-length cDNA of a novel gene (named yp05) was obtained from the electronic assembly of 3'-RACE and 5'-RACE products.

RESULTSThis yp05 gene was 787 bp including a 597 bp open reading frame (ORF) and encoded a deduced protein with 199 amino acid residues, and the amino acid sequence of this protein was found similar with the sequences of many fungal manganese-superoxide dismutases in the GenBank with the aid of BLASTp. The transcription of yp05 gene in Monascus strains was analyzed with the aid of Northern blotting. The transcription of yp05 gene was only detected in Monascus strains, provided that citrinin was produced.

CONCLUSIONThe transcription of yp05 gene belongs to differential expression genes of citrinin yielded from Monascus and has no correlation with the biosynthesis pathway of red pigments.

Amino Acid Sequence ; Base Sequence ; Blotting, Northern ; Citrinin ; biosynthesis ; Cloning, Molecular ; DNA, Complementary ; chemistry ; Fungal Proteins ; chemistry ; genetics ; Gene Library ; Molecular Sequence Data ; Monascus ; genetics ; metabolism ; Mycelium ; genetics ; metabolism ; Pigments, Biological ; biosynthesis ; RNA, Messenger ; metabolism ; Sequence Alignment ; Sequence Analysis, DNA

OBJECTIVETo establish an instant determination method of citrinin in red kojic by high performance capillary electrophorphores for the first time.

METHODRed kojic was extracted with the mixtrue of Toluene and ethyl acetate (70:30). Separation was carried out in an uncoated fused silica capillary (50 microm x 45.0 cm). Meanwhile, a running voltage 15 kV, 20 mmol L(-1) borax buffer with 10.0 mmol L(-1) sodium deoxycholate (pH 9.3) and a UV detector at 212 nm were adopted.

RESULTRegression equation of citrinin was Y=9434 + 16781X (r =0.990), The lower limit of quantification (S/N > or =3) was 0.5 mg mL(-1). The assay coefficients of variation ranged from 98.8% to 101.1%. The intra and inter recovery ranged from 0.83 to 1.54% and from 1.86 to 5.09%. Twenty samples were determined with the method.

CONCLUSIONThe method is proved to be simple, rapid and accurate, and it can be used to determine citrinin in red kojic.

Citrinin ; analysis ; Electrophoresis, Capillary ; methods ; Hydrogen-Ion Concentration ; Monascus ; chemistry ; Reproducibility of Results

OBJECTIVETo construct a tag expression library of Monascus aurantiacus that could produce citrinin maximally on the thirteenth (0.966 mg/mL) day in the submerged culture.

METHODSTotal RNA was extracted from the mycelium, cDNA was synthesized using the SuperScript choice system, and then, a SAGE library was successfully constructed according to the MicroSAGE method.

RESULTSFive hundred and ninety eight clones were obtained in SAGE library, and 120 clones were picked out randomly for identification and sequencing purpose. Eighty nine clones had positive inserts, 26 clones had no inserts and the remaining 5 clones had no site of NlaIII enzyme in inserts. There were seven repeated clones.

CONCLUSIONWith the aid of SAGE2000 software, 901 tags were obtained from 89 clones, representing 686 unique transcripts. Six unique tags of them belong to highly expressed genes (Number of tags > or = 10) and 143 unique tags to moderately expressed genes (repeat tags > or = 2).

Anti-Bacterial Agents ; biosynthesis ; Citrinin ; biosynthesis ; Expressed Sequence Tags ; Gene Expression ; Gene Expression Profiling ; Gene Library ; Monascus ; genetics ; growth & development ; metabolism ; Polymerase Chain Reaction ; RNA, Fungal ; genetics ; isolation & purification

Extracts of Monascus purpureus have always been considered a natural source of lovastatin, the precursor of the world's largest selling class of drugs. In actual fact, the fungus contains many other substances (flavonoids, polyunsaturated fats, pyrrolinic compounds etc.) with a wide variety of other actions. The most recent studies have shown that it has an action on the glycemic metabolism, and on the mechanisms of adipogenesis, also an effects on the endothelium and on postprandial vasodilation. These effects are more extensive and complex than those of statins alone. And new strains of Monascus purpureus have recently been patented where the presence of statins is only one of the therapeutic components of the fungus. In particular, the increase in secondary components, such as flavonoids, which coincides with a more complex therapeutic action, probably making the new extracts of Monascus purpureus, the ideal candidate for the treatment of the metabolic syndrome.
Aged ; Coronary Disease ; drug therapy ; Humans ; Hydroxymethylglutaryl-CoA Reductase Inhibitors ; therapeutic use ; Monascus ; Phytotherapy ; Plant Extracts ; therapeutic use