Optimisation of ultrasound-assisted extraction conditions for maximal recovery of active monacolins and removal of toxic citrinin from red yeast rice by a full factorial design coupled with response surface methodology

Highlights

• A practical method of ultrasonic extraction was developed in red yeast rice.

• Response surface methodology was used to optimise the process variables.

• This method reached maximal recovery of monacolins and removal of toxic citrinin.

• This method may be applied for industrial in manufacturing of monacolins.

Abstract

This study optimised the ultrasound-assisted extraction (UAE) conditions to achieve maximal recovery of active monacolins with minimal contents of citrinin from red yeast rice (RYR). A central composite design after a full factorial design was utilised to examine the different UAE parameters. The studies revealed that HAc%, extraction time and EtOH% had significant influences on the recovery yield of monacolins, while HAc% and EtOH% were key factors for the elimination of citrinin. The resulting optimal conditions were as follows: ultrasound power of 250 W, HAc% of 7.7%, RYR amount of 0.2 g (solvent-to-solid ratio 40 mL/g), extraction time of 50.7 min, EtOH% of 57.2% and extraction temperature of 20 °C. Under these conditions, at least 94.7% of monacolins was recovered and 87.7% of citrinin was removed from RYR. This optimised UAE condition was further evaluated for potential industrial application in manufacturing of RYR as pharmaceuticals and nutraceuticals.

Introduction

Natural products as pharmaceuticals and nutraceuticals with positive effects on human health have been popular in our daily life. Red yeast rice (RYR), a well-known functional natural product from fermentation of steamed rice using fungus Monascus purpureus, has been widely used, especially in Eastern Asia, to improve digestion and blood circulation (Feng et al., 2012, Zhu et al., 2012). RYR was found to contain a family of important secondary metabolites, such as monacolins, that are widely used in pharmaceutical and food industries (Hong, Seeram, Zhang, & Heber, 2008). Monacolins are known to be responsible for inhibiting hydroxymethylglutaryl-coenzyme A (HMG-CoA) reductase, and thus regulate the cholesterol synthesis in liver. In clinic, monacolins have remarkable therapeutic effects on lipid profiles of hypercholesterolemic patients (Gordon, Cooperman, Obermeyer, & Becker, 2010). Monacolin K (a.k.a. lovastatin), one of the first commercially available HMG-CoA reductase inhibitors, was the major monacolin member in RYR. In addition, monacolins were also evaluated for their potential anticancer activities in colorectal cancer (Poynter et al., 2005) and breast cancer (Campbell et al., 2006) etc.

Monacolins have been of great interest to the food and pharmaceutical industries due to their inimitable benefits, an efficient process is therefore required to achieve maximal recovery of natural monacolins from RYR. However, due to the ubiquitous nature of several fungal species, citrinin was also formed during the fermentation process. Citrinin is a mycotoxin with hepatotoxicity and nephrotoxicity and is one of the most prevalent human contaminants in the food chain (Lee, Hung, et al., 2007), and must be limited to no-more-than 200 ppb in a daily administration (Nigovic, Sertic, & Mornar, 2013). Several clinical trials have found that 5 mg/day of monacolins in RYR has a comparable efficacy to 20 mg/day of pure monacolin K in lowering blood cholesterol (Chen, Yang, Uang, & Lin, 2013). Hence, the acceptable content ratio of monacolins and citrinin (M/C) in a final product must be great than 25 (5000/200), in order to be considered as effective and safe.

To achieve that ratio, some researches focused on general culture conditions and substrate evaluation for reducing citrinin while retaining monacolins formations in RYR (Lee et al., 2007, Lee et al., 2007). However, the selective culture conditions were not always suitable for every Monascus species, even though altering culture condition was able to reduce the citrinin formation. On the other hand, the use of the optimal culture condition on an industrial scale proves to be expensive and hard to manipulate for monacolins formation in RYR (Sun, Zou, Liu, & Xiao, 2011).

The development of a postprocess to selectively remove citrinin while retain monacolins from RYR product was proposed as an alternative solution to the above problem. Extraction is often utilised as an efficient and important technique in the recovery and purification of target substances from plant materials. Recently, various extraction techniques, such as heating, boiling, Soxhlet, accelerated solvent and ultrasound extraction, were developed for the recovery of monacolins from RYR (Liu et al., 2010, Nigovic et al., 2013) but the content of citrinin by-product was always neglected in these practices.

Ultrasound-assisted extraction (UAE) exhibited the best mass transfer, cell disruption, solvent penetration and capillary effect (Da Porto, Porretto, & Decorti, 2013) and it did not damage the temperature sensitive botanical materials (Peralta-Jiménez & Cañizares-Macías, 2013). Moreover, UAE was the simplest and most economical technique, and easy to scale up for industrial production (Delgado-Povedano and Luque de Castro, 2013, Wang et al., 2013).

Many factors, including ultrasound power, extraction time and temperature, and solvent-to-material ratio etc., can influence the UAE process, individually and collectively (Xu et al., 2013), and it is difficult to single out main independent variables to optimise (Meziane, 2013). The conventional multivariable optimisation is usually based on the “one-factor-at-a-time” approach, which is unable to detect interactions among independent variables, and lack of complete information on effects of all determinants (Meziane, 2013, Xu et al., 2013). Response surface methodology (RSM) is a useful tool for evaluating multiple parameters and their interactions based on quantitative data (Li et al., 2011), and may effectively overcome the drawback of classic “one-factor-at-a-time” or “full-factors” approach.

The objectives of this study were (1) to establish an optimised simple, safe and low cost process of UAE for the food and pharmaceutical industries to extract maximum yield of monacolins with minimum citrinin content from RYR, and (2) to develop a rapid and accurate quality control method for the determination of monacolins contents using UHPLC-DAD–Q-TOF/MS. To the best of our knowledge, there has been no report on RSM application to the optimisation of UAE conditions for obtaining the high yield of monacolins with low content of citrinin in RYR.