Discovery of the chemical constituents, structural characteristics, and pharmacological functions of Chinese caterpillar fungus

: Caterpillar fungus ( Cordyceps sinensis ) has been widely used as a traditional Chinese medicine for several decades. It is essential to clarify the product composition, structural characteristics, and pharmacological functions of caterpillar fungus. In this review, we comprehensively and systematically summarized the various bioactive components isolated from caterpillar fungus, including nucleosides, D - mannitol, sterols, ﬂ avonoids, fatty acids, amino acids, vitamins, peptides, amides, proximate, and mineral composition. Meanwhile, peptides, nucleo - sides, and polysaccharides serve as the main active com - ponents in this genus, which possess immunomodulatory, antioxidant, anti - allergic, anti - tumor, anti - in ﬂ ammatory, antibacterial, anti - malaria, and antifungal activities. Consequently, the active components of caterpillar fungus demonstrate a vital source of treatment for various dis - eases and can be used as possible leads for drug discovery. This article reviews the composition and pharmacological action of caterpillar fungus, which is the key to ensuring the safety and e ﬀ ectiveness of caterpillar fungus, and will be of interest for future research.


Introduction
Caterpillar fungus, named after a fungus that lives on insects, has been used medicinally in China for more than 300 years.The fungus is commonly referred to as "Dong Chong Xia Cao," which means "winter worm and summer grass," whose name normally refers to Cordyceps sinensis (C.sinensis).C. sinensis is a fungus that lives on the larvae of Lepidoptera (Figure 1) [1].In the late autumn, the fungus infects the caterpillar and consumes its host, and in early summer the following year, the grass-like fruiting body protrudes from the dead host's head.Its habitat is mainly distributed in Qinghai, Tibet, Sichuan, Yunnan, and Gansu [1], where there are many grasslands and shrubs of 3,000-5,000 m.The caterpillar fungus is honored as one of the three greatest invigorants in China together with Pilose antlers and Panax.In Tibet, caterpillar fungus has probably been used for at least a thousand years.
Nevertheless, caterpillar fungus caught the attention of the world when it was discovered that several Chinese athletes had made it an important part of their training programs and had broken world records in 1993 [2].The caterpillar fungus is an aggregation of larva and parasitic fungus relating to Lepidopteran host does not just represent a monophyletic population.Over 400 kinds of Cordyceps spp.are already described worldwide, and about 90 kinds of Cordyceps spp.Moreover, based on morphological and molecular evidence, Hirsutella sinensis is now considered to be the correct clone of caterpillar fungus [4].Recent research shows that caterpillar fungus and its asexual forms have many biological effects, such as polysaccharides, cordycepin, D-mannitol, ergosterol, and so on [5].Other bioactive compounds, including proteases, cyclosporin-like metabolites, and nucleosides, are also produced by this fungus and related species.
The results of some modern studies have shown that many species of Cordyceps have a very wide range of biological activities and applications.Cordyceps is widely used in China to tonify the kidney, moisten the lung, boost and maintain overall body health.Other treatments such as for nighttime sweating, fatigue, weakness after a serious illness, high blood sugar, respiratory disorders, arrhythmias, and renal insufficiency have also been claimed [3].Due to its unique medicinal value and the growing demand and shortage of natural resources, the development of the fermentation of Cordyceps fungal mycelium for medicinal production has proven to be a viable and sustainable means.Some other Cordyceps species also have a high utilization value, and there are some commercialized products on the market, such as ditanganic acid of C. militaris [6].Consequently, fermentation of mycelium isolated from Cordyceps to achieve large-scale production of Cordyceps is a research trends for many scientists.Meanwhile, with the increasing interest in the mycology and pharmacology of Cordyceps, an overview of its potential has become more and more important and urgent.
2 Chemical constituents and active ingredients analysis

Nucleosides
The concentration of nucleosides is an important standard for quality control of caterpillar fungus and its substitutes.To date, over ten nucleosides, bases, and their related ingredients have been isolated and identified from caterpillar fungus, including adenosine, adenine, cytosine, cytidine, uridine, guanine, inosine, hypoxanthine, thymine, thymidine, and 2′-deoxyuridine [7].
Several methods have been used to analyze the nucleosides in natural and artificial caterpillar fungus, including capillary electrophoresis, thin layer chromatography, and high performance liquid chromatography (HPLC).Highly sensitive, selective, and accurate mass spectrometry combined with HPLC photodiode array detection could perform simultaneous separation, identification, and quantification of nucleosides in Cordyceps mycelium [8].In addition, a method for the determination of nucleosides, nucleotides, and their transformation products in caterpillar fungus by ion-pair reversed-phase liquid chromatography-mass spectrometry was developed [9].The concentrations of nucleosides in caterpillar fungus mycelia are shown in Table 1.
Generally, the nucleosides displayed potent antiviral, anti-cancer, anti-inflammatory, anti-tumor, neuroprotective, and antioxidant activities [10].Nucleosides are the main bioactive components and basic materials of caterpillar fungus and play a key role in regulating various physiological processes in the body [11].In addition, recent studies have shown that adenosine and cordycepin are important nucleosides in caterpillar fungus, which play an important role in protecting organs and tissues from damage, immunomodulation as well as anticancer [12].Nucleosides have been widely accepted as one of the biologically active principles of caterpillar fungus.However, they are not the sole cause of all biological activity [13].

Cordycepin
Cordycepin (also known as 3′-deoxyadenosine) is a kind of purine alkaloid, a derivative of adenosine (Figure 2).It was first extracted from Cordyceps in 1951 [14], and then found to be present in caterpillar fungus [15].Various samples were added into methanol-water (50/50, V/V) for ultrasonic extraction of cordycepin.In the fruiting bodies of caterpillar fungus, the content of cordycepin was 2.654 ± 0.02 mg/g, which was higher than that in natural caterpillar fungus (0.9801 ± 0.01 mg/g), while the content of cordycepin in the mycelia of fermented caterpillar fungus was 0.9040 ± 0.02 mg/g, which was basically consistent with that of natural caterpillar fungus [16].
In order to achieve industrial mass production of cordycepin, a lot of research has been done on efficient cultivation methods for caterpillar fungus [17,18].Investigations showed that adenosine is directly converted to cordycepin, and the related enzymes have been identified [17,19].Adenosine concentrations in Cordyceps mutans in the range 4.7-6.7 g/L can be converted to cordycepin.The maximum content of cordycepin in the medium with and without adenosine was 8.6 and 6.7 g/L, respectively [20].The effect of the addition of ferrous sulfate on cordycepin content in submerged cultures of caterpillar fungus was investigated in shaking flasks.The results showed that   the maximum amount of cordycepin was close to 596.59 ± 85.5 mg/L under the condition of optimal addition of 1 g/L ferrous sulfate, which was about 70% higher than the condition of no ferrous sulfate addition [21].
To date, cordycepin has been shown to have a variety of pharmacological effects, such as anti-inflammatory, immunological, antioxidant, anti-viral, anti-cancer, antimicrobial, and hypoglycemic properties [22].Cordycepin inhibits the expression of diabetes-regulated genes by inactivating NF-κb-dependent inflammatory responses and has also been reported to affect diabetic nephropathy by inhibiting apoptosis in a rat model of diabetic nephropathy, suggesting that cordycepin has satisfying potential as a safe anti-diabetic drug [23,24].Furthermore, cordycepin has been shown to significantly increase the levels of antioxidant enzymes and it can be considered a potential antioxidant [25].Meanwhile, cordycepin has the potential to be a fully effective bioactive anti-inflammatory, immunomodulatory, and anti-tumor component in the future [16,26,27].

D-Mannitol (cordycepic acid)
D-Mannitol as one of the quality control indicators of cultivated caterpillar fungus is the main active component of caterpillar fungus [28].Cordycepic acid was first extracted from caterpillar fungus in 1957 and identified as 1,3,4,5tetrahydroxycyclohexanoic acid, an isomer of quinic acid.However, further investigation revealed that the report about structural assignment was completely erroneous."Cordycepic acid" is neither a new compound nor an acid, but in fact D-mannitol (Figure 2) [29].
The content of D-mannitol in natural caterpillar fungus ranges from 35.42 to 38.64 mg/g, which was much higher than that in cultivated Cordyceps (10.24-13.41mg/g).A method for the determination of D-mannitol in caterpillar fungus by enzyme-label-assisted 412 nm ultraviolet spectrometry was developed based on the high specificity and high throughput of the enzyme-labeled instrument [30].
Investigation showed selenium-enriched cultures of Cordyceps can increase D-mannitol content in fruiting bodies in the concentration range of 0-6.0 ppm sodium selenite, but at selenium concentrations above 6.0 ppm, D-mannitol content decreases slightly.Nevertheless, D-mannitol content from both the non-selenium Cordyceps fruit bodies (3.12%) and the selenium-enriched Cordyceps fruit bodies (3.87%) was lower than that in wild caterpillar fungus (5.24%) [31].
Cordyceps acid is often used as one of the indicators to evaluate the quality of artificially fermented Cordyceps.
Studies have shown that D-mannitol has the ability to promote human metabolism, improve the body's microcirculatory system, significantly lower blood lipids, inhibit bacteria, increase resistance to disease, as well as cough suppressant, expectorant, and asthma calming effects [32].
Ergosterol is one of the main sterols found in caterpillar fungus [35], which can be converted to vitamin D2 by UV exposure and is also a nutrient factor that promotes normal bone development in humans and other mammals (Figure 4).Ergosterol is generally present in two forms, free ergosterol and esterified ergosterol, and the relative content of different types of ergosterol varies.Currently, a gradient reversed-phase high performance liquid chromatographic method for the simultaneous determination of free ergosterol and esterified ergosterol were carried out in the fruiting body and host caterpillar of Cordyceps.Investigations showed that the two ergosterols were similar in composition (ergosteryl esters, free ergosterol, and ergosterol analogues), and the total ergosterol content was 2.110 and 2.456 mg/g, respectively.However, host caterpillar ergosterol ester content (0.255 mg/g) was much higher than that of the cysts [36].Determination of four free sterols, including rape sterol, cholesterol, ergosterol, and β-sitosterol in natural and cultured caterpillar fungus using pressurized liquid extraction, one-step trimethylsilane derivatization and GC-MS analysis was developed [37].
At the same time, these sterols exhibit a variety of pharmacological properties such as antiviral, cytotoxic, antiarrhythmic, alleviating immunoglobulin A nephropathy, and inhibiting activated human mesenchymal cells [38,39].

Saccharides and polysaccharides
The phenol-sulfuric acid reaction is the primary method for the determination of submicron sugars and related substances.Monosaccharides, oligosaccharides, polysaccharides, and their derivatives, including methyl ethers with free or potentially free reducing groups, appear orange-yellow in the presence of phenol and concentrated sulfuric acid treatment.The phenol-sulfuric acid reaction is sensitive and the color is stable [40].Slight differences exist in the total sugar content of caterpillar fungus between the corpus and fruiting body, but they are conspicuously lower than that in mycelium [41].
Cordyceps polysaccharides (CPs), the richest group, possess enormous potential.The total polysaccharide content of natural Cordyceps is similar to the fruiting body (fungi) and worms (caterpillars), but the total polysaccharide content of Qinghai Cordyceps is slightly lower than that of Tibetan Cordyceps [42].The polysaccharide contents in cultured Cordyceps are in the range of 2.6-7.0% of the total weight [43].The contents of polysaccharides in Cordyceps are generally detected by colorimetric assay using sulfuric acid-anthrone [43].Commonly, CP includes neutral and acidic polysaccharide, the latter is the largest group of CP and comprises one or more carboxyl, phosphate, or sulfate groups in the sugar repeating unit.
Optimization of the operating conditions for polysaccharide extraction can also affect the extraction results.The Box-Behnken design was used to examine the three key variables of extraction time, extraction temperature, and number of extractions of cultured Cordyceps mycelium.When it is extracted for three times with 110 min per time at 88.9°C, the highest extraction rate of 16.10% was obtained [44].
The extraction of water-soluble crude polysaccharides from Cordyceps is usually carried out by hot water extraction and ethanol precipitation.Ion exchange and gel chromatography can also be used for the purification of CP.Natural CP has similar chemical properties to artificial CP as analyzed by the sugar labeling method [42].These polysaccharides contain mainly (1 → 4)-α-glycosidic bonds, (1 → 4)-β-D-glycosidic bonds, 1,4-β-D-mannopyranose bonds, and (1 → 6)-α-glycosidic bonds, with (1 → 4)-α-D-galacto-aldonic bonds also present in some polysaccharides.A larger number of CP and their components have been reported to be isolated and identified from natural and cultured Cordyceps species (Table 2).

Fatty acids
Fatty acids are classified into saturated and unsaturated fatty acids.Subsequently, unsaturated fatty acids are considered to be a potent physiologically active component with unique functions in lowering blood lipids and preventing cardiovascular disease.The fatty acid methyl esters in oils and fats were analyzed by gas chromatography.The main fatty acids in the natural fruiting bodies of Cordyceps are palmitic acid (C 16:0 ), linoleic (C 18:2 ), and oleic acid (C 18:1 ), accounting for approximately 97% of the total fatty acids [47].The environment in which an organism lives is closely related to the fatty acid composition of its lipids, for example, temperature significantly affects the composition of membrane lipids [48].Seventeen fatty acids were identified from different wild Cordyceps by GC-MS, and their absolute and relative contents were determined.The samples were collected from Tibet, Qinghai, and Yunnan provinces, and the main components were C 16:0 , C 18:0 , C 18:1 , and C 18:2 , with linolenic acid (C 18:3 ) (1.02-4.83%).The total unsaturated fatty acid content (75.74-90.87%) is significantly higher than that of other common fungi.The abundant C 18:3 content and high unsaturated lipids of wild Cordyceps may be inherited from the abundant C 18:3 in the host Hepialus larvae [49].

Crude proteins and proximate composition
Amino acids are the main active ingredients in Cordyceps and could be hydrolyzed from crude proteins.In general, the contents of crude protein from Cordyceps are in the range of 14.8-37.2%,as well as the proximate composition of caterpillar fungus is summarized in Table 3 [50].
The contents and compositions of amino acids are determined after hydrolysis.There are no obvious differences in the types of amino acids between different Cordyceps.
From the results we can conclude that the content of amino acids is mostly 20-25%, and contents of glutamate, serine, arginine, and aspartic acid are the highest.The levels of amino acids in the fruiting body are similar  to those in the fermented mycelium of Cordyceps species, although their total levels of amino acids are different.
The crude proteins and total amino acids in mycelia from submerged culture and mycelia from shake culture are higher than those in natural Cordyceps.
Comparing with the natural Cordyceps collected from Qinghai, Yunnan, Sichuan, and Tibet, the contents of amino acids in Cordyceps samples collected from Qinghai province are lower than those from Yunnan and Sichuan [51].

Vitamins and mineral compositions
Vitamin B1 and niacin contents of the freeze-dried mycelium from Cordyceps are 0.35 and 1.19 mg/g, respectively, while the content of niacin in natural fruiting bodies of Cordyceps is much higher (3.06 mg/g).Vitamin B6 is also detected in natural fruiting bodies of Cordyceps (1.36 mg/g) [52].
Many studies have confirmed the presence of a wide range of mineral compositions in Cordyceps.The mineral compositions include macronutrients (Ca, Na, Mg, K) and trace elements (Ni, Zn, Fe, Mo, Cu, Mn, Cr, V, Co, Se), as well as non-essential and toxic elements (As, Ba, Sn, Pb, Cd, Hg).By using inductively coupled plasma mass spectrometry to determine the concentrations of various elements in the biological samples, the researchers found that all elements in the Cordyceps matrix were higher than those in natural Cordyceps, except Zn, Mg, and Cu [53].Consequently, the contents significantly depend on different species, and are shown in Table 4 [54].

Nitrogenous compounds-peptides and amides
In recent years, many researchers have begun to focus on the ability of antifungal proteins to stop pathogenic fungi from invading crops and causing disease in animals.
Given the relatively little information available on the protein composition of artificial Cordyceps and its much lower cost compared to natural Cordyceps, researchers have conducted studies to determine if antifungal proteins can be isolated from Cordyceps.
The cordymin from the extract of Cordyceps was obtained by cation exchange chromatography, which was an antifungal protein and different from other edible/non-medicinal fungi and refer to the N-terminal sequence.The N-terminal sequence of cordymin showed no significant similarity to other known proteins/peptides in the PubMed database as AMAPPYGYRTPDAAQ.The researchers determined the purity and molecular mass of cordycepin using mass spectrometry and found that there was only one peak in cordycepin with a molecular mass of 10906.62Da [55].
Small amounts of polyamines have also been isolated and identified, such as cordyceamides A and B. Their structures are shown in Figure 5. Though the studies of peptides and amides in caterpillar fungus are not enough, peptides are certainly as potential markers for quality control of caterpillar fungus.

Flavonoids
Generally, studies on the active components of caterpillar fungus mainly focus on cordycepin, CP, cordycepic acid, etc., while studies on flavonoids in caterpillar fungus are few and lack systematic sorting and induction.Flavonoids are widely distributed in nature and have a variety of pharmacological effects.They have certain therapeutic and rehabilitative effects on the immune system, the nervous system, and the cardiovascular system.Flavonoids not only have antioxidant activity, but also anti-tumor and immune-enhancing effects.
It has been shown that the genistein flavonoids and their methyl glycoside derivatives contained in the ethyl acetate extract of caterpillar fungus can inhibit the activation of AKT and ERK1/2 proteins and exert immunomodulatory effects [57].Furthermore, the high inhibitory effect of flavonoids on HIV-1 protease could be used as a potential HIV-1 virus drug [58].In addition, it has been found that the antioxidant capacity of caterpillar fungus may be related to its ability to scavenge free radicals by flavonoids and other substances [59].

Immunomodulating function
Cordyceps extract may eliminate the inhibition of phagocytosis by the virulence factor streptococcal pyrogenic exotoxin B of group A streptococci, which causes a wide range of diseases in humans, including impetigo, pharyngitis, streptococcal toxic shock syndrome, scarlet fever, and necrotizing fasciitis.Thus, Cordyceps fungi may help to modulate the immune response and help provide resistance to bacterial infections [3].
As antecedently mentioned, several main substances with immunomodulatory activity had been isolated from caterpillar fungus, and polysaccharides were the major part [60].In the immune responses, an exopolysaccharide named cordysinocan was isolated from Cordyceps UST 2000 strain.Cordysinocan induces proliferation of cultured t-lymphocytes and secretion of interleukin-2, interleukin-8, and interleukin-6.In addition, cordysinocan temporarily induced phosphorylation of extracellular signal-regulated kinases.The application of cordysinocan in cultured macrophages enhanced phagocytosis and acid phosphatase activity of macrophages.Thus, these results demonstrate the important role of CPs in triggering this immune response [43].H1-A, a pure compound isolated from Cordyceps, inhibits the progression of autoimmune diseases in MRL Ipr/Ipr mice [61].Furthermore, immunosuppressive therapy can suppress acute cardiac rejection [62].
Currently, only a relatively small number of studies have reported on the mechanisms by which the host immune system recognizes non-pathogenic fungi.Therefore, the researchers sought to find out whether DNA from caterpillar fungus could have the same activating effect on mouse bone marrow-derived dendritic cells (BM-DCs) as DNA from other pathogenic fungi [3].Then results demonstrated that DNA extracted from caterpillar fungus could lead to activation of BM-DCs in a TLR9-dependent manner [63].Caterpillar fungus is an important source for the treatment of various diseases  9

Anti-tumor activity
Previous studies have shown that both tumor-specific and non-specific immunity is suppressed in tumor-bearing animals, so boosting immunity can be of great help in tumor treatment.Lung cancer is the most common cause of cancerrelated deaths.Aqueous extract of Cordyceps induces apoptosis and affects telomerase activity in human lung cancer A549 cells [64].Ethyl acetate from the mycelium of Cordyceps fungi is a potential natural anti-tumor product with strong anti-tumor activity [65].
Cordycepin and polysaccharides are widely determined cytotoxic and anti-tumor components of Cordyceps species [3].Artificially cultured CPs have been shown to have the same potent pharmacological effects as natural Cordyceps.
Researchers studied the effects of the extracellular polysaccharide fraction (EPSF) of a cultured Cordyceps fungus on the expression of c-Myc, c-Fos, and vascular endothelial growth factor (VEGF) in tumor-bearing mice.Three different doses of EPSF were administered intraperitoneally to mice every 2 days, and the expression of tumor-associated genes in lung and liver tissues was subsequently studied.The results showed that EPSF had an inhibitory effect on tumor growth in mice lung and liver, and may have a role in tumor control [61].Consequently, EPSF are extensively investigated for their potential for elevating immune cell activity in H22-bearing mice [66].

Antioxidation activity
Reactive oxygen species-induced oxidative stress may be a direct or indirect cause of tissue damage and many human diseases such as ageing, cancer, atherosclerosis, and inflammation.Therefore, a number of antioxidants have been reported for the prevention of cancer and coronary heart disease, protection of the myocardium against experimental myocardial infarction, and prevention of some neuronal signs of ageing [67].
Recent studies have confirmed the significant antioxidant activity of extracts of Cordyceps fungi and their polysaccharide fractions [68].The antioxidant activity of aqueous extracts of natural and artificial Cordyceps from different sources is usually analyzed by three different assays, including xanthine oxidase, induced hemolysis, and lipid peroxidation.In general, Cordyceps possesses potent antioxidation activity in all analysis tests.Artificial Cordyceps has equally strong activity compared to natural Cordyceps.The antioxidant activity of Cordyceps partially purified in vitro culture was increased by 10-30 times, suggesting that its antioxidant activity may be partly derived from CPs [69].
Therefore, recently, more and more studies have been reported on the fermentation of Cordyceps fungal mycelium for exopolysaccharide production.EPSF isolated from mycelial liquid or submerged culture of Cordyceps also shows moderate antioxidant activities [67].For instance, CP can increase the antioxidant activity of immunosuppressed mice, effectively increase the levels of catalase, superoxide dismutase, glutathione peroxidase and total antioxidant capacity in mice, and reduce the level of malondialdehyde in mice [70].Other compounds isolated from Cordyceps were also investigated on antioxidation activity.Increased glutathione reductase, superoxide dismutase, glutathione peroxidase, catalase, and glutathione-s-transferase activities and increased reduced glutathione levels in aged rats are confirmed after cordycepin treatment.Consequently, cordycepin is effective for recovering antioxidant status and decreasing lipid peroxidation in elderly rats [71].
The significant evidence for the role of altered antioxidant defenses and free radicals in the etiology of diabetes has been shown [70].Cordyceps and taurine significantly increase glucose uptake in the diaphragm of normal and diabetic rats.Moreover, Cordyceps increased HDL-cholesterol levels, serum insulin, percentage of the beta-cell function, total antioxidant capacity levels, and pancreatic reduced glutathione levels [67].

Protective effects on human organs 3.4.1 Hepatoprotective activity
The liver consists mainly of hepatocytes, which are active in the metabolism of exogenous chemicals, making the liver a target for toxic substances [72].Therefore, research on the hepatoprotective activity of biological and natural sources has attracted a lot of attention in recent years.The aqueous extract of Cordyceps has now been found to prevent oxidative damage to biomolecules.It is possible that water extracts of Cordyceps possess hepatoprotective properties.The cultured Cordyceps compared with the natural Cordyceps on the protective effects of hepatotoxicity, the results showed that the aqueous extract of Cordyceps had a protective effect against t-butyl hydroperoxide (t-BHP)induced oxidative damage in HepG2 cells [73].

Kidney protective activity
Recently, there has been a lot of research focused on clearing up the mechanisms of progression of interstitial fibrosis and identifying potential novel anti-fibrotic treatments to slow its progression [74].It is thought that the traditional Chinese medicinal preparation Cordyceps may have beneficial effects on kidney disease.
The effects of Cordyceps were observed in a model of renal fibrosis ureteral obstruction.The active component of Cordyceps is present in the water-soluble polysaccharide component of the crude extract (Cp-F1).Cp-F1 antagonizes the effect of TGF-b1 on HK-2 cells, blocks the induction of fibronectin and a-SMA, and inhibits the epithelial cell marker E-cadherin.These data further demonstrate the anti-fibrotic effect of Cordyceps in renal disease, as evidenced by the reduction in the extent of interstitial fibrosis in the renal cortex [74].A soluble polysaccharide (CPS-2), purified from the cultured Cordyceps, could efficaciously alleviate renal failure caused by pulverizing kidney [75].

Lung protective activity
Asthma is a complex disease that affects millions of people and its incidence has increased dramatically.Studies on the pharmacological properties of Cordyceps have now confirmed the beneficial effects of Cordyceps on a variety of lung diseases such as asthma, non-productive cough, chronic obstructive pulmonary disease, bronchitis, and pulmonary fibrosis.Cordyceps extract has a dose-dependent effect on ion transport in the Calu-3 human airway epithelial monolayer model.Cordyceps extract affected the anion movement from the basolateral to apical interstitial compartment of the airway epithelium, indicating that Cordyceps extract can improve lung function and can be used in the treatment of respiratory diseases [76].Meanwhile, idiopathic pulmonary fibrosis is an irreversible, chronic, and debilitating lung disease.Cordyceps showed potential to prevent and treat pulmonary fibrosis in a rat model of bleomycin-induced pulmonary fibrosis [77].

Anti-fatigue activity
The caterpillar fungus is considered to be a rare tonic because of its life-supplementing properties.Residents of high-altitude mountains such as Tibet and Nepal claim that Cordyceps gives them energy and reduces the symptoms of altitude sickness.In the West, athletes and the elderly also enjoy coffee.Compared to a placebo control group, healthy young volunteers supplemented with cultured Cordyceps powder while running with poor energy significantly enhanced energy production and resistance to fatigue [78].The current study confirmed that oral administration of in vitro cultured Cordyceps mycelium significantly improved swimming endurance in rats.Some researchers have investigated the expression levels of endurance-responsive skeletal muscle metabolic regulators GC-1α, AMPK, PPPAR-δ, as well as endurance-promoting and antioxidant genes GLUT4, MCT1, VEGF, MCT4, NRF-2, SOD1, TRX in red gastrocnemius muscle, and these results provide sufficient molecular evidence [79].
Paecilomyces japonica is a new type of Cordyceps that promises to have tonic effects [80].In a study, the effect of its extract on forced swimming ability in mice enhanced forced swimming ability by increasing fat utilization and delaying the accumulation of plasma lactate and ammonia.Cordyceps extracts also could prolong the mice swimming time [81].

Anti-hyperglycemic activity
Type 2 diabetes mellitus (DM), caused by a combination of defective insulin secretion and insulin resistance, and is a metabolic disease characterized by hyperglycemia and dyslipidemia.So, the thorny issue at hand is how to prevent diabetes and its complications.Several traditional Chinese medicines have been used to address the hyperglycemic condition of diabetics [66].
Presently, many polysaccharides, isolated from the cultural mycelia of Cordyceps, are proved to have the hypoglycemic activity [82].Natural Cordyceps fruiting bodies have hypoglycemic activity when taken orally and can reduce weight loss and excessive drinking caused by diabetes [83].The significant reduction in the overall incidence of diabetes through oral administration of Cordyceps is found.Moreover, Cordyceps could also help to prevent diabetic endothelial dysfunction and related complications [84].

Other activities
The caterpillar fungus has a radioprotective effect, reducing the loss of blood cells after radiation [85].In addition, caterpillar fungus has been used to enhance sexual performance and restitute impairment in sexual function.
The caterpillar fungus and its fractions significantly stimulate production of testosterone in mouse.Therefore, caterpillar fungus can be used as an alternative medicine for reproductive problems [15,86].The study also confirmed the anti-inflammatory effect of the chloroform and n-butanol fractions of the methanolic extract of Cordyceps [87].

Conclusions and future prospects
Products of the caterpillar fungus are becoming important health foods for different people, different physiological states, and different nutritional health need.However, with the increase of the harvest pressure of caterpillar fungus, the supply of natural caterpillar fungus exceeds supply.So far, it has become credible on the basis of numerous studies demonstrated that the substitutes have the same efficacy as natural caterpillar fungus.In the foreseeable future, bioactive active ingredients, such as CPs, cordycepin, and Cordyceps alternative materials will become a hot research topic in the field of green pharmacology and biopharmacology.

Table 1 :
Content of nucleosides in natural and cultured caterpillar fungus (mg/g)

Table 3 :
Proximate composition of the caterpillar fungus

Table 4 :
Mineral compositions in natural stroma and worm, fruit body of the caterpillar fungus n.t.: not tested; n.d.: not detected.