红曲霉所产桔霉素及其检测与控制

红曲霉能够生成多种生理活性而备受关注，但大多红曲霉同时也产生一种真菌毒素—桔霉素。红曲产品的桔霉素问题已经使我国红曲出口受到了很大的影响。介绍了红曲霉所产桔霉素的产生、合成途径、检测方法以及控制红曲霉产生桔霉素的几种措施。     

红曲色素和桔霉素代谢调控方法的研究进展

在食品安全问题日益受到关注的今天,取代人工合成色素的天然色素具有广阔的市场前景。红曲色素是国际范围内生产量及使用量最大的天然食用色素之一,广泛应用于肉制品加工和水产品加工领域的着色。福建红曲以品质优、色价高的特点,在国内外享有极高声誉,但在红曲霉代谢过程中同时会合成真菌毒素桔霉素,使红曲色素在使用中存在安全隐患。我国红曲色素产品中桔霉素含量普遍偏高,制约了我国红曲产品的出口和使用领域的拓展。红曲色素和桔霉素合成呈伴生现象,常出现提高红曲色素产量的同时,桔霉素含量也相应提高的情况,因此调控红曲霉代谢过程中促进红曲色素的产生同时抑制桔霉素的产生成为解决这一矛盾的关键。文章介绍了国内外红曲色素和桔霉素的代谢调控方法的研究概况,主要包括菌种选育、发酵工艺优化及基因工程等来达到调控目的,通过掌握红曲菌代谢调控网络,研究红曲菌产桔霉素的代谢途径及途径中的代谢机制,进而综合分析调控红曲霉合成代谢桔霉素存在的问题并提出应对措施,对推动红曲产品的出口和红曲产业的健康发展具有重要意义。     

红曲霉代谢产物的研究进展与应用

红曲霉(Monascus)是散囊菌目中的子囊菌曲霉科真菌,一种丝状腐生真菌,具有较高的糖化能力和酯化能力。文章主要介绍了红曲霉产生淀粉酶、糖化酶、蛋白酶、酯化酶等初级代谢酶及次级代谢产物红曲色素、莫纳克林K、桔霉素与γ-氨基丁酸的研究进展;简要介绍了代谢产物在食品行业、医药行业以及酿酒行业的应用,并对红曲霉代谢产物的研究进行了展望。     

5种金属离子对紫色红曲霉主要次生代谢物谱的调节作用

目的:研究金属离子对紫色红曲霉生长和主要次生代谢物合成的调节作用。方法:将紫色红曲霉孢子接种到含不同浓度的金属离子（Cu2+、Zn2+、Mg2+、Mn2+、Co2+）的固体培养基,观察不同离子对其生长和色素合成的影响;同时,将不同浓度的上述离子及金属螯合剂8-羟基喹啉分别接种到液体培养基中,测定紫色红曲霉生物量、红曲色素、莫纳克林K、γ-氨基丁酸和桔霉素等主要次生代谢物含量及发酵液中代谢物谱变化。结果:Cu2+（0.5 mmol/L）处理导致水溶红、橙、黄色素产量分别提升了2.01、2.07和1.73倍;醇溶色素分别增加了2.83、1.74和1.95倍;莫纳克林K和桔霉素分别增加了1.78和5.53倍。Zn2+（0.5 mmol/L）促使水溶、醇溶色素分别增加了93.3%、78.6%、57.6%和2.9、1.54、0.54倍。莫纳克林K和桔霉素则分别增加了0.92和6.91倍。0.5 mmol/L的Mg2+处理导致生物量有所下降,但无显著差异;水溶、醇溶色素则分别增加了2.68、2.25、2.02倍和1.26、1.23、1.82倍;莫纳克林K则增加了51.8%。0.25 mmol/L的Mg2+则使桔霉素提升了1.79倍。Mn2+（0.5 mmol/L）刺激水溶、醇溶色素产量分别增加了48.3%、29.9%、24.8%和131.1%、83.6%、49.2%,莫纳克林K增加了1.01倍。而桔霉素则在1 mmol/L Mn2+刺激下增加了92.3%。Co2+（0.5、1 mmol/L）导致生物量分别降低了47.9%和53.7%,而醇溶色素则分别提高了4.07、5.04、2.62倍和3.07、3.41、2.70倍,对莫纳克林K合成无作用;桔霉素产量分别降低了90.8%和93.9%。上述离子均不利于γ-氨基丁酸的合成。金属离子螯合剂8-羟基喹啉（20 μmol/L）使红曲霉生物量降低了26.9%,水溶色素分别下降了40.6%、34.3%和26%。莫纳克林K和桔霉素分别降低了56.7%和90.1%。发酵液的波谱（200~600 nm）扫描结果发现,金属离子仅调节代谢物的产量,并不改变紫色红曲霉代谢物种类。结论:金属离子不仅影响紫色红曲霉的生长,并显著调节其主要次生代谢物谱。     

不同液态发酵基质对红曲霉产红曲色素及桔霉素的影响

在前期分离纯化出的10株红曲霉基础上,以18种液体培养基对10株红曲霉进行发酵培养,通过分光光度计和质谱分析仪测 定各发酵液中红曲色素和桔霉素含量,分析不同地区红曲霉的红曲色素和桔霉素的代谢特性,筛选出高产红曲色素低产桔霉素的 红曲霉菌株。 结果表明,产红曲色素最适的液体培养基配方为葡萄糖3%、蛋白胨1%,其中新疆地区红曲霉所产红曲色素量最高,为 6.81×10-2 mg/mL;新疆地区红曲霉仅在18种培养基中的面筋碱性蛋白酶水解液+葡萄糖发酵液中产生了桔霉素,而红曲霉ZBX天津 在所有培养基发酵液中均未产生桔霉素。     

温度对红曲霉15-1产桔霉素的影响

试验对不同温度对红曲霉15-1在YES培养基上产桔霉素的影响进行了研究。通过HPLC法对桔霉素的生成量进行动态检测,结果表明,温度对红曲霉生长及桔霉素浓度有较大影响。在试验温度范围内,35℃有利于红曲霉的生长,此温度条件下桔霉素产量则相对较低,而且随着培养时间的延长,桔霉素还可被红曲霉分解利用,降解率可达76.22%。因此,红曲生产中可适当延长发酵时间,以降低红曲发酵产物中桔霉素的浓度。     

桔霉素与红曲霉的安全性争议

红曲霉能代谢生成天然红色素和多种生理活性物质而备受国际关注,但同时也发现其生长过程产生桔霉素.桔霉素是一种真菌毒素,作用的靶器官是肾脏,可以致畸、诱发肿瘤、突变等.红曲霉中桔霉素的发现使得我国的红曲产品的安全性引起争议.介绍了红曲中产生的桔霉素的理化性质、毒性、产生途径和检测方法等的研究现状,并就如何消除红曲中桔霉素提出了对策.     

物理场辅助法对红曲霉次级代谢产物影响的研究进展

近年来,红曲霉次级代谢产物的实际应用价值越来越高。同时物理场辅助法具有选择性好、周期短、应用潜力大的特点。综述了不同物理辅助手段(超声波、微波、光照、磁效应)对红曲霉产红曲色素、Monacolin K、桔霉素等的影响情况,并且对复合物理场手段调控红曲霉次级代谢产物进行了展望。     

高产开环型monacolin K红曲霉菌株的筛选及其在红曲酒酿造中的应用

该研究采用38株红曲霉菌株固态发酵制备功能红曲米,采用高效液相色谱法测定红曲米中monacolin K和桔霉素含量,筛选高产开环型莫纳可林K(monacolin K)红曲霉菌株,通过形态观察及分子生物学技术对筛选菌株进行鉴定,并将其应用于红曲酒酿造,分析红曲酒的品质。结果表明,筛选得到1株高产开环型monacolin K、不产桔霉素的菌株ABQ2,经鉴定,其为丛毛红曲霉（Monascus pilosus）。采用该菌株发酵制备功能红曲米中的monacolin K含量达到25.10 mg/g,开环率为90.5%,且桔霉素未检出。采用菌株ABQ2酿造的红曲酒酒精度为12%vol,酸度为1.91 g/100 mL,monacolin K含量为0.48 mg/mL,其理化指标均符合相关行标要求;与小曲酒相比,红曲酒中总酯含量（433.65 mg/L）极显著增加（P<0.01）,其中乳酸乙酯和乙酸乙酯含量最高,高级醇类含量（145.52 mg/L）略低,主要为正丙醇、异丁醇、正戊醇、β-苯乙醇等。     

高产Monacolin K红曲霉的诱变选育及其与酿酒酵母共酵培养

为提高红曲中莫纳克林K含量,以实验室保藏紫色红曲霉（Monascus purpureus）MY-11为原始菌株,利用紫外诱变筛选红曲霉,并进行固态发酵制备红曲,采用高效液相色谱（HPLC）法对红曲中的莫纳克林K含量进行测定,选出高产莫纳克林K突变株,并与食源性菌株酿酒酵母（Saccharomyces cerevisiae）进行共酵。结果表明,通过紫外诱变筛选出2株高产莫纳克林K突变株M7和M8,固态发酵25 d后莫纳克林K含量分别为12.42 mg/g、12.49 mg/g,分别比原始菌株MY-11提高了26%、27%。将诱变菌株M7、M8分别继续与不同添加量的酿酒酵母共酵培养,结果发现加入酵母菌液添加量3%时莫纳克林K含量最高,分别为9.35 mg/g、8.94 mg/g,比对照组提高了2.63%、16.41%。紫外诱变筛选结合共酵培养方法可提高红曲中莫纳克林K含量。     

Characteristics of Monascus sp. isolated from Monascus fermentation products

To screen for the strains suitable for industrial applications, 15 Monacus sp. were isolated from various Monascus fermentation products. The isolated strains were belonging to Monascus pilosus, Monascus purpureus, Monascus albidulus, and Monascus femeus based on the ITS sequence and morphological characteristics. They produced globular and hyaline conidia, and red, orange, or hyaline ascomata except for M. fumeus, in which ascomata was not observed in malt extract media. Each strain produced monacolin K and citrinin at different levels on different media: 15.79–20.05 mg/L of monacolin K and 0–255 μg/L of citrinin in submerged culture of monacolin K producing media, 1.27–2.69 mg/L and about 507.4 μg/L in YES medium, 0–0.78 g/kg and 0–9.7 mg/kg in Monascus red rice products, suggesting that it is necessary to adopt the similar conditions to a specific application for screening as well. These various isolated strains would be great resources for industrial application to develop Monascus fermented functional products.     

Edible Filamentous Fungi from the Species Monascus: Early Traditional Fermentations,Modern Molecular Biology,and Future Genomics

Monascus spp. are filamentous fungi famous for their fermented products, especially red mold rice (RMR), a traditional fermented food in East Asian areas with a very long edible history documented back to the Han dynasty (BC 202‐AD 220) in China. Nowadays, RMR and its related products involve a very large industry from artisanal traditional fermentations to food companies to medicine manufacturers, which are distributed worldwide. Modern studies have shown that Monascus spp. are able to produce abundant beneficial secondary metabolites, such as monacolins (cholesterol‐lowering agents), γ‐amino butyric acid (an antihypertensive substance), dimerumic acid (an antioxidant), and pigments (food‐grade colorants), and some strains can also secrete citrinin, a nephrotoxic metabolite. Monascus‐related studies have received much attention because of their wide applications. However, to our knowledge, no systematic review on the progress of Monascus research has ever been published. In this review, the progress of research on Monascus is summarized into 3 stages: Monascus fermentation, Monascus molecular biology, and Monascus genomics. This review covers the past history, current status, and future direction of Monascus research, contributing to a comprehensive understanding of Monascus research progress.     

Study on the fingerprint profile of Monascus products with HPLC–FD,PAD and MS

Monascus products have been widely used as food additives and pharmaceuticals. Citrinin is a toxic metabolite produced during the Monascus fermentation. In this work, a quick extraction method with ultrasonic treatment was studied for a complete extraction of citrinin from the fungi cells. Furthermore, the proposed HPLC method has the advantage of the simultaneous determination of Monascus pigments and citrinin without sample pretreatment. Under the optimized HPLC conditions, a baseline resolution of pigments, citrinin and other catabolites was achieved. The detection limit of citrinin reached 0.5 ng mL⁻¹ (S/N > 3/1) with a fluorescence detector (FD). Eight Monascus pigments were detected, with photodiode array detector (PAD), two of which not yet described. The HPLC fingerprint profile of the eight Monascus pigments and citrinin appears significant for the quality control of the Monascus product.     

红曲的生产工艺及Monacolin K和桔霉素的检测

综述红曲的生产工艺及Monacolin K和桔霉素的检测方法,指出采用新型生产工艺、优化发酵条件和应用菌种筛选等现代技术是生产既富含活性成分Monacolin K又不合桔霉素的功能性红曲的重要途径,同时预示Monacolin K和桔霉素检测技术的进展将为功能性红曲产品的质量、安全使用及国际市场的开拓提供技术支撑.     

Biochemical and morphological characterization of a new fungal contaminant in balsamic and cider vinegars

In this study we have determined the morphological, some physiological and biochemical characteristics of a fungal strain isolated from balsamic and cider vinegars. The examination of fungal contamination of 100 balsamic and cider vinegar samples showed that 90% contained more than 10⁴ colony forming units (CFU ml^?1), but in most samples (65%) values of more than 10⁵ CFU ml^?1 were obtained. The most frequent contamination was with Monascus sp. (80% of the samples examined showed 2 × 10³ to 6 × 10⁵ CFU ml^?1). This mould can produce secondary metabolites such as citrinin and monacolin K_L (lactone form), and monacolin K_A (acid form). Using high-performance liquid chromatography (HPLC) with fluorescence detection, citrinin was found in 68% of the samples. The concentration of these mycotoxins varied between 1.6 × 10^?2 µg ml^?1 and 7.2 × 10^?2 µg ml^?1. The concentrations of citrinin were very low, and we can anticipate that this compound at these concentrations has no toxic effects on renal cells. Monacolin was not detected in any sample studied.     

组学技术在红曲霉研究中应用的进展

组学技术的发展拓展了人们对红曲霉生长发育、代谢产物的合成及代谢调控机制的认知。总结了近年来红曲霉组学研究的现状及热点问题,从基因组学、转录组学、蛋白质组学及代谢组学等多个维度剖析了组学技术在红曲霉研究中的应用状况。基因组学主要用于探讨红曲霉的进化地位、基因簇的预测与挖掘、同源基因的比对及代谢途径分析等;红曲霉生长和代谢过程中基因或基因簇的转录表达、代谢调控机制及新转录调控因子的预测和挖掘是转录组学的应用范围;蛋白质组学主要涉及红曲霉代谢过程中重要酶类的功能解析及其参与代谢产物的机制解析;而代谢组学则以红曲霉代谢产物为基础,解析其潜在的代谢机制。阐述了红曲霉代表性次级代谢产物之一——红曲色素在研究和生产中面临的问题,并围绕调控红曲色素生产的高效化、精细化、安全性等方面,总结了基于组学技术的菌种选育、培养基优化、化学和物理因素调节及菌株的代谢改造等调控技术。对综合运用组学技术围绕红曲色素代谢调控的研究提出了展望,以期为推动红曲霉研究及其代谢产物的生产提供参考。     

A 90-D Toxicity Study of Monascus-Fermented Products Including High Citrinin Level

Abstract: Monascus is one of the traditional fermentation fungi and has been used in many kinds of food for thousands of years. Although Monascus-fermented red mold rice performs cholesterol-lowering effects, blood pressure-lowing effects, and antioxidant effects, another metabolite, nephrotoxic and hepatotoxic citrinin, causes the concerns for safety. Various citrinin concentrations (1, 2, 10, 20, and 200 ppm) in the red mold rice are, respectively, estimated for safe use in animal tests. According to the results of serum biochemistry assays of liver and kidney in each group, citrinin did not reveal any nephrotoxicity and hepatotoxicity. Furthermore, the results of histopathological slices of liver and kidney in each group did not show any significant differences from control histopathological findings. As a result, we presume that citrinin concentrations in Monascus-fermented products within 200 ppm will not affect the functions of liver and kidney or cause any nephrotoxicity and hepatotoxicity. According to safety factor, it is proposed that 2 ppm citrinin in Monascus-fermented products may be a safe concentration.     

红曲菌中桔霉素的控制策略及研究进展

红曲菌在我国的使用历史悠久,其在食品添加剂、发酵食品、酿酒和中药等方面应用广泛。目前,虽然液 态发酵技术基本解决了红曲色素产品中的桔霉素含量高的问题,但固态发酵红曲米产生的桔霉素含量高的问题依然 是制约红曲企业及产业发展的主要技术难题之一。本文对红曲菌代谢产物中桔霉素的毒性、限量标准、检测方法、 合成途径及调控基因进行了综述,并从传统方法和生物方法两个层面阐述了控制红曲菌中桔霉素含量的主要策略, 旨在为有效降低红曲菌中桔霉素的含量和促进红曲产业健康发展提供参考。     

Thermal degradation kinetics analysis of monacolin K in Monascus-fermented products

The objectives of this study were to investigate the thermal degradation kinetics of monacolin K in Monascus-fermented product (MP) solution. The results indicated that monacolin K content in MP solution remained fairly constant in the range of pH 3-9. However, heat treatment significantly decreased the content of monacolin K in MP solution, for example, from 75.8% under 100 °C, 60 min to 54.3% under 100 °C, 90 min at pH 9. Moreover, if the heating temperature was increased to 121 °C for 90 min at pH 9, the content of monacolin K rapidly decreased to 14.1%. The thermal degradation of monacolin K followed a first-order reaction kinetic. The temperature dependence of rate constants followed Arrhenius relationship, with an activation energy value of 126.64 kJ/mol (90-121 °C, r² = 0.99). Our results suggest that monacolin K is easily degraded when the MP solution is heated to a high temperature (such as 121 °C). However, more than 50% of monacolin K could be remained when the MP solution is heated under the pasteurization temperature used for food processing.     

Improved functionality of soft soybean curd containing Monascus fermented soybean ethanol extract

Monascus fermented soybeans (MFS) were produced by solid state fermentation and ethanol extractions were carried out to recover monacolin K (MK) and isoflavones from the MFS. About 99 and 87% of monacolin K (891 mg/kg) and isoflavones (895 mg/kg) were recovered by 80%(v/v) ethanol extraction. The 80% ethanol extract also showed significantly higher antioxidant activities (DPPH, ABTS radical scavenging activity, and ferric reducing antioxidant power) than the other ethanol extracts prepared by either 40 or 60% ethanol. The 80% Monascus fermented soybeans extract (MFSE) also contained significant α-glucosidase inhibitory activity (2.7 acarbose g equivalents/g MFS). Based on the results, MFSE can be used to enrich bioactive MK and isoflavone aglycones in soft soybean curd products.