What Is the Active Ingredient in Luo Han Fruit Extract?

Siraitia grosvenorii Swingle is the fruit of Siraitia grosvenorii Swingle, a perennial woody vine in the gourd family. It is an economically and medicinally important plant that is endemic to China. China has a long history of using plants to treat diseases. Traditional Chinese medicine believes that Luo Han Fruit can refresh the mind and generate saliva, clear away heat and moisten the lungs, dispel fire and relieve coughing, and smooth the intestines and promote bowel movements [1]. Luo Han Fruit has the reputation of “Oriental Elixir” and “Fruit of Longevity” in Southeast Asia and Western countries [2]. Monkshood glycosides, flavonoids, fatty acids, etc. have all been found in monkshood. Monkshood glycosides are the most studied compound in monkshood. The physiological effects of monkshood or its extracts, such as anti-fatigue, anti-inflammatory, anti-cancer, and lipid-lowering and weight-loss effects[3], are closely related to the chemical composition of monkshood and its metabolism and mode of action in the body.

In recent years, with the development of chemical analysis technology, some new compounds in Luo Han Guo have been identified. The inhibitory effect of the chemical components in Luo Han Guo on the growth of some fungi is closely related to the improvement of anti-inflammatory and immunity; the antioxidant effect is inseparable from fatigue resistance and anticancer; the regulation of sugar metabolism in the body and the protective effect on pancreatic beta cells can have a positive effect on the symptoms of diabetes. This article reviews the research on the chemical constituents of Luohanguo, including Mogroside, flavonoids, and lipid compounds, as well as the mechanism of action of Luohanguo extract in three aspects: antibacterial and anti-inflammatory, inhibition of diabetes, and scavenging free radicals. It also reviews the application of  Luo Han Fruit Extract in commercial products.

1 Chemical composition research

1.1 Mogrosides

Mogrosides are the most researched chemical components unique to Luo Han Guo, with mogroside V being the most abundant. Mogrosides contain unsaturated C=C bonds and hydroxyl groups, which form the basis of their chemical properties. The mogrosides have a common structure, namely a four-ring triterpene (mogroside), and the sugar moiety is always glucose. The way in which the sugar moiety is bonded differs, and the structural formula is shown in Figure 1 [4].

Because Luohanguo has long been used as a prescription medicine, its active ingredients have not been given much attention. Mogroside is a unique component of Luohanguo and was the first of its chemical components to be studied and utilized. In 1974, Lee reported that Luohanguo contains the di-terpene tianweiside. In 1983, the Japanese Takemoto was the first to use spectroscopy and chemical analysis to identify the structure of mogroside and its glycosides [5]. A total of 12 mogrosides have been isolated and identified, namely mogroside IV, mogroside V, mogroside III, mogroside II E, mogroside III E, mogroside VI, mogroside A, neomogroside, siamenside I, mogroester, grosmomoside I, III, etc.[6-7]. The hydroxyl group on the 11th position of mogroside V is oxidized to a ketone, and the resulting 11-oxo-mogroside V is also an important form of mogroside.

As research continues, new Luo Han Guo saponins are being identified all the time. For example, Luo Han Guo III A1 has been identified by 1H-NMR and 13C-NMR to have a structure of the structure was identified as Luohangol-24-O-β-D-glucopyranosyl (1→2)-[β-D-glucopyranosyl (1→6)]-β-D-glucopyranoside [8-9]. Various glycosides with the 11-position hydroxyl group oxidized, such as 11-oxo-morindin II E and 11-oxo-morindin II E[10], have been isolated and identified in immature Luohanguo.

Mogrosides are all derivatives of the tetracyclic triterpene class of cucurbitane. The difference lies in the different glucose moieties linked at R1 and R2, so the structural similarity causes some interference in quantitative analysis. The analysis methods for Mogrosides have undergone developments in ultraviolet spectrophotometry, thin-layer scanning [11], high-performance liquid chromatography (HPLC) [12-14], and high-performance liquid chromatography-electrospray ionization mass spectrometry [15], making the analysis of Mogroside composition more accurate and precise. High-performance liquid chromatography is the most widely used method for the detection of loganin because it is convenient, fast and accurate. The analytical methods for the common types of loganin are also relatively mature. Chang Qi et al. [16] used vanillin-perchloric acid method to measure the content of total triterpene saponins in Luo Han Guo for the first time, using self-made Mogrosides  as the reference substance, and the content was 3.75% to 3.85%.

Due to its high content, Mogroside V is an indicator component for evaluating the quality of Luohanguo. Zhou Jing et al. [17] used an HPLC method with an acetonitrile-water system as the mobile phase for elution, and measured Mogroside V and 11-oxo-Mogroside V at a detection wavelength of 210 nm in different cultivation systems. with contents ranging from 0.346% to 1.267% and 0.103% to 0.332%, respectively. Lu Fenglai et al. [18] used an acetonitrile-water system as the mobile phase for gradient elution, within 10 min, achieved good separation and detection of six Mogrosides: Mogroside V, Mogroside IVA, Mogroside III, 11-oxo-loganin III, Mogroside IIE and 11-oxo-Mogroside IIE.

The application of nuclear magnetic resonance and mass spectrometry techniques has played an important role in the identification of new Mogroside in Luo Han Guo. However, the newly identified Mogroside still have the basic structure of triterpene saponins, and their growth period is different from that of Luo Han Guo [19]. They are present in small amounts, are easily degraded and converted by enzymes, and are difficult to purify. However, they have laid a foundation for studying the pharmacological effects of these compounds. High-performance liquid chromatography has become a mature technique for detecting six types of Mogroside, and the preparative chromatography developed from this has been able to better separate Mogroside monomers with higher purity [7].

1.2 Flavonoids

From the existing research data, it can be seen that the analysis and research of flavonoids in Luohanguo has not yet attracted much attention, as most of the flavonoids exist in the form of glycosides. The basic units or main components are mainly quercetin and kaempferol, which have some significant physiological effects, such as antibacterial, antioxidant and smoothing blood vessels. Si Jianyong et al. [20] analyzed the components of fresh Luohanguo fruit and isolated two flavonoid glycosides for the first time: kaempferol-3-O-α-L-rhamnopyranoside-7-O-[β-D-glucopyranosyl-(1-2)-α-L-rhamnopyranoside] and kaempferol-3,7-α-L-dirhamnoside, and named the former grosvenorine. Chen Quanbin et al. [21] found the flavonoid aglycone quercetin in the hydrolysates of fresh Luo Han Guo fruit and its Mogroside, and determined the total flavonoid content to be 5–10 mg/fruit using quercetin and kaempferol as external standards by RP-HPLC. Flavonoids were also found in the hydrolysates of the Mogroside, accounting for about 1.42% of the total Mogroside.

The identification of new substances in Luohanguo is mainly due to the difference in the glycoside bound to the quercetin or kaempferol structure. The identification of the structure and composition lays the foundation for the isolation and purification of the flavonoids in Luohanguo. Tang Chunli [22] used high-speed countercurrent chromatography (HSCCC) with ethyl acetate, n-butanol and water as the stationary phase to isolate the flavonoid glycoside KR with a purity of 94.6%, and found that it can form a stable complex with Cu and has a variety of physiological effects. The research on the process of extracting Luo Han Guo flavonoids with different concentrations of ethanol, enzymatic hydrolysis and separation and purification by macroporous resin has also developed rapidly [23].

1.3 Fats and oils

Plant seeds generally have a high content of fats and oils, and Luo Han Guo is no exception. Chen Quanbin et al. [24] used Soxhlet extraction, supercritical, ultrasound and reflux extraction to extract the fats and oils from Luo Han Guo seeds, with a yield of 6.40% to 11.46%. Infrared spectroscopy and GC-MS analysis identified the main components as squalene (docosahexaenoic acid, accounting for 51.52% of the oil content, [z,z]-9.12-octadecadienoic acid (23.89%), and 3-hydroxy-1,6,10,14,18,22-docosapentaenoic acid (9.58%).

Li et al. [25] found that the seed oil of Luohanguo is rich in Farnesol (3,7,11-trimethyl-2,6,10-dodecatrien-1-ol, 52.14%), which is commonly used as a thickener in high-end perfumes.

As a food, Luo Han Guo is also relatively well-balanced nutritionally, especially rich in proteins and amino acids, trace elements, and carbohydrates. It contains all eight essential amino acids[26]; 26 inorganic trace elements; a large amount of monosaccharides and polysaccharides, and a high content of fructose among the reducing sugars.

2 Pharmacological effects

Luohanguo has been reported to have a variety of physiological activities, such as lowering blood sugar, lowering blood lipids, antibacterial and anti-inflammatory, and anti-cancer. Mechanistically, these effects are attributed to the antibacterial, anti-diabetic and free radical scavenging properties of Luohanguo and its extracts.

2.1 Antibacterial and anti-inflammatory

Due to its unique chemical composition, Luohanguo exhibits a significant inhibitory effect on the main cariogenic bacteria and oral bacteria, such as streptococci. It also has an anti-inflammatory effect on bacteria that invade the body through its effect on the lymphatic system.

Mu Jing et al. [27] used a basic medium prepared with sucrose, with 1:10 SG extract added as the experimental group, and with stevia, glucose, sucrose and fructose added as the control group. The medium was mixed with sucrose to prepare a basic medium, and this medium was used to culture Streptococcus mutans. The growth rate of the bacteria was measured by turbidity, and the metabolism of the microorganisms was investigated by measuring the viscosity and acid production. The results showed that the experimental group had a significant inhibitory effect on the growth and metabolism of Streptococcus mutans compared to all the control groups, and that SG was not easy to be mixed with sucrose as a mixed sweetener.

However, Zhou Ying et al. [28] believed that these studies lacked scientific biological assay methods and purified monk fruit extract. They used HPLC to isolate the SG extract and perform high-throughput screening of the antibacterial activities of different isolated components. using high-throughput screening of the antibacterial activity of different isolated components, it was found that the isolated fragments #18-19 and #34-35 had strong antibacterial activity against Streptococcus mutans cultured on academic agar, while loganin V did not show strong antibacterial activity. The antibacterial biological activity of loganin I-IV and other components in SG still needs to be studied.

The invasion of gram-negative bacteria into the body induces the synthesis of iNOS and COX-2 by lipopolysaccharide (LPS) on the cell wall, and both are closely related to inflammation [29-30]. Monk fruit extract (25.9% mogroside V) can inhibit the activation of the MAPK signaling pathway, thereby inhibiting the translocation of NF-κB caused by exogenous LPS, and reducing the protein levels of iNOS and COX-2 [31]. Mogroside V can coordinate the function of the immune system by blocking the active nutrient absorption pathway in the body when foreign microorganisms invade [32]. Due to its ability to inhibit oral bacteria and prevent inflammation by regulating the immune system, Luo Han Guo has the potential to become a component of chemical drugs for removing dental caries, cleaning the mouth and treating inflammation caused by tumours.

2.2 Diabetes resistance

Diabetes is a common and frequently occurring disease. It is the third leading cause of death after tumors and cardiovascular disease, and a major threat to human health. Among the three major principles of diet, exercise and drug therapy, diet therapy is the basic measure for the treatment of diabetes [33]. Luohanguo glycoside has a triterpene saponin structure, and studies have shown that triterpene saponins have hypoglycemic effects [34]. In addition, the physiological effect of Luo Han Guo in lowering blood sugar and blood lipids has also been reported. Luo Han Guo glycoside 11-oxo-luo han guo dry V can effectively inhibit the oxidation of LDL, reducing the risk of atherosclerosis caused by elevated LDL [35].

Alloxan has a selective damaging effect on pancreatic β cells, causing a drop in insulin levels in the blood of mice and resulting in insulin-dependent diabetes [36]. When mice treated with this model were given an extract of SG by gavage, the results showed that a certain concentration of the SG extract could significantly reduce the serum cholesterol (TC) and triglycerides (TG) of mice, and increase high-density lipoprotein cholesterol (HDL-C) [37]. SG extract can significantly reduce blood glucose levels in mice overloaded with glucose, apolypoprotein A-I, and starch, but increase insulin levels and decrease α-glucosidase levels [38]. GK (Goto-Kakizaki) mice have a congenital non-insulin-dependent diabetes that does not lead to obesity or hyperlipidemia, which is very similar to the type of diabetes in the Asian population. Studies have shown that monk fruit extract has a regulatory effect on the α-glucosidase inhibitor voglibose; improves renal function and has a certain preventive effect on type 1 diabetes [34]. However, the composition and ratio of loganin in the SG extract differ in terms of the level of insulin induction [36], and the metabolic and physiological mechanisms of action of the individual components after ingestion are not yet clear.

2.3 Antioxidant

Free radicals with unpaired electrons in living organisms are closely linked to the occurrence of many diseases. Luo Han Guo extract can exhibit antioxidant capacity by directly scavenging free radicals, regulating the activity of free radical scavenging enzymes, and inducing the gene expression of substances that can synthesize free radical scavenging capacity in living organisms. Glutathione peroxidase (GSH-Px) and superoxide dismutase (SOD) are typical representatives of the endogenous free radical scavenging enzyme system in animals [39]. Mangosteen extract and mangostin have a good effect on maintaining the stability and increasing the levels of glutathione peroxidase (GSH-Px) and superoxide dismutase (SOD) in mice that have consumed high-fat diets [40]. Heme oxygenase (HO) is a rate-limiting enzyme in heme metabolism with certain physiological activities. HO-1 is a heme oxygenase (HO) that can be induced by heme, ultraviolet light, hydrogen peroxide and some cytokines. HO-1 has been widely studied for its antioxidant, anti-inflammatory and anti-apoptotic effects [41-42]. SG can regulate the synthesis and activity of HO-1 through mRNA, while inhibiting the expression of Mn-SOD mRNA. However, more research is needed on the dosage and toxicity of SG in achieving its physiological effects by participating in enzyme systems with antioxidant activity [43].

Hironobu Yasuno et al. [44] A pesticide synergist, piperonyl butoxide (PBO), is a hepatocarcinogen in mice. It was found that SG extract, although it cannot inhibit the early response of hepatocarcinoma through its own ability to scavenge free radicals, can activate the expression of free radical scavenging enzymes through the transcription factor Nrf 2-Keap-ARE, thereby reducing hepatocarcinoma in mice caused by PBO intake.

Although SG extract does not have strong free radical scavenging ability, it can achieve a certain scavenging effect by regulating the transcription and expression of enzymes in the endogenous free radical scavenging system. Therefore, Luo Han Guo extract can prevent and repair early lesions caused by free radicals, especially cancer, such as skin cancer [41] and liver cancer [45]. These studies have laid the foundation for the mechanism and clinical application of the antioxidant capacity of Luo Han Guo extract. In addition, excessive exercise leads to an increase in free radicals in the body, which attack mitochondrial membranes and organelles, leading to metabolic disorders and fatigue [46]. Luo Han Guo can work in synergy with exogenous free radical scavengers to reduce fatigue [47].

Microbial invasion of the body, lipid peroxidation, damage to pancreatic β cells and a decrease in insulin receptors, and an imbalance of free radicals in the body can all cause various diseases in the human body. However, as an organic whole, the effect of the intake of monk fruit extract on the organism is also comprehensive and complex. Luohan fruit extract can improve the response time of insulin and thus improve the symptoms of type II diabetes. At the same time, it can inhibit lipid peroxidation and strengthen the pressure on the kidneys, thereby having a certain effect on improving the negative effects of obesity [3]. At the same time, the metabolic pathway and metabolites of insulin after ingestion have not been detected in the liver and blood [48]. Therefore, the metabolic pathway and products of Luohan fruit in the body have yet to be studied.

3 Products and applications

Luo han guo and Luo han guo extract are the most widely produced products on the market. However, the processing technology for these products is simple, the technical indicators for the products are relatively broad, and the added value is low. The most widely used ingredient in Luohanguo is Mogroside, which is highly sweet and low in calories. The sweetness of Mogroside V, Mogroside VI and Mogroside IV is more than 300 times that of sucrose. Therefore, monacolin is used in the development of new sweeteners [49-50]. Mogroside is also used in the tobacco leaves and filters of cigarettes to increase the aroma of tobacco and reduce the dryness of the smoke [51-52]. The pharmaceutical applications of Luohanguo are mainly concentrated in traditional Chinese medicine or over-the-counter drugs. In traditional Chinese medicine, Luohanguo is often used together with other ingredients such as Baibu, Baiqian, and Sangbai Pi to prepare the raw materials for ointments, which have the effect of relieving coughs. Luohanguo V and alkaloids are the main active ingredients in this process [53-54]. Such drugs are available in syrup, products in various dosage forms such as syrup, paste, capsules and tablets, such as Luo Han Guo cough syrup [55], Luo Han Guo cough capsules [56–57], and Luo Han Guo cough tablets [58–59].

In addition, the extract of Luo Han Guo, which is obtained by the process of blending it with traditional Chinese medicines such as chrysanthemum, wolfberry and cassia seed, and then extracting and drying it, can also be used to prepare functional health drinks [60].

4 Outlook

In-depth research and utilization of Luo Han Guo has only recently begun in China. At present, research on Luo Han Guo mainly focuses on monacolin K, while further research is needed on the purification, identification and analysis of other valuable ingredients such as flavonoids and lipids. In recent years, as the public has become more health-conscious, Luo Han Guo, as a resource for both food and medicine and a traditional Chinese medicinal ingredient, is bound to attract further attention in the development of auxiliary materials and products. However, from the perspective of the chemical composition, pharmacological effects, products and applications of Luo Han Guo, breakthroughs are still needed in the following areas: a. the extraction and isolation of Mogroside ingredients at the food and pharmaceutical grade, which mainly involves the stability and change law of Mogroside, structural identification of the components, etc.; b. the types of functional components in Luo Han Guo extract and toxicological research; c. the mechanism research of the components and their functions in Luo Han Guo extract; d. the relationship between the function of Luo Han Guo and the structure of its functional components. With the development of science and technology, further progress will definitely be made in the research of the above aspects. Luo Han Guo-related products will appear more and more in people's lives, bringing greater economic benefits.

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