








key: cord-336753-4212ftlc
authors: Luo, Lu; Jiang, Jingwen; Wang, Cheng; Fitzgerald, Martin; Hu, Weifeng; Zhou, Yumei; Zhang, Hui; Chen, Shilin
title: Analysis on herbal medicines utilized for treatment of COVID-19
date: 2020-05-27
journal: Acta Pharm Sin B
DOI: 10.1016/j.apsb.2020.05.007
sha: 
doc_id: 336753
cord_uid: 4212ftlc

As coronavirus disease 2019 (COVID-19) pandemic poses a substantial global public health threat, traditional Chinese medicine (TCM) was used in 91.50% of the COVID-19 cases in China, showing encouraging results in improving symptom management and reducing the deterioration, mortality, and recurrence rates. A total of 166 modified herbal formulae consisting of 179 single herbal medicines were collected for treating COVID-19 in China. Glycyrrhizae Radix et Rhizome, Scutellariae Radix, and Armeniacae Semen Amarum are the most frequently utilized in clinics, most of which are antipyretic (47, 26.26%), expectorant and cough-suppressing (22, 12.29%), and dampness-resolving (21, 11.73%) from traditional descriptions. A total of 1212 chemical components containing β-sitosterol, stigmasterol, and quercetin were primarily selected. Additionally, using complex system entropy and unsupervised hierarchical clustering, 8 core herbal combinations and 10 new formulae emerged as potentially useful candidates for COVID-19. Finally, following scaffold analysis, self-organizing mapping (SOM) and cluster analysis, 12 clusters of molecules yielded 8 pharmacophore families of structures that were further screened as pharmacological targets in human metabolic pathways for inhibiting coronavirus. This article aims to make more easily accessible and share historical herbal knowledge used in contemporary treatments in a modern manner to assist researchers contain the global spread of COVID-19.

While COVID-19 is now being effectively controlled within China, it is increasingly affecting other countries worldwide, most notably the U.S., Iran, and Italy since Feb 25, 2020 1 3 With a long history of combating epidemic diseases with relatively low mortality, TCM has accumulated a rich antiviral herb knowledgebase from clinical observation and pharmacological selection. This is historically based on a phenomenological approach and relies on complex mixtures of herbal medicines, as well as nonpharmacological holistic interventions, such as acupuncture and health lifestyle guidance 5 . With pandemic diseases, TCM has shown the agility to be administered more quickly and more efficiently than conventional medicine 4 . It asserts that health is a state of harmony between an individual's internal physiological network of factors and that of an external environmental 6 . Diagnosis is through syndrome differentiation called "Bian Zheng", a holistic analysis of clinically observed information that guides personalized treatment options 4 , which may be adaptably changed dynamically during disease progression 7 . From point of TCM, epidemic disease is considered caused by an external pathogenic attack (traditionally named pestilent Qi or evil Qi) due to climatic or environmental changes. In the battle between healthy Qi (immunity) and pathogenic Qi (pathogen), a weakened immune system results in disease, while exhaustion of healthy Qi leads to death 7 .

Consequently, different syndromes can manifest as disease progresses. From TCM perspective, the fundamental pathogenic factor of COVID-19 would be termed as "dampness accumulation in the lung", which invades from the exterior to the interior attacking other organs as disease develops from the early stage to mild, moderate, severe, critical, and recovery stages. According to the syndrome differentiations, the pathogenesis in the mild and moderate stages of COVID-19 is damp accumulation in the lung. During the severe and critical stages, dampness develops to "damp-heat toxin and congests in the lung". While heat toxin "burns" or rapidly depletes Qi and Yin, this subsequently would appear as what is termed "dual vacuity of Qi and Yin of lung and spleen in the recovery stage" 3 . It could be otherwise expressed somewhat as exhaustion of the lung system, its moisture, membranes and resource to recover, along with its fundamental immune supporting system. Herbal formulations are then frequently modified as signs and symptoms change along with the diagnosis and corresponding syndrome differentiations. Appropriate herbal formulae are administered, containing herbs composed in a hierarchy called "Jun Chen Zuo Shi", relating to their primary, secondary, tertiary and quaternary functional class order, respectively 8 . Through design, herbal formulae guide a combination of different herbal medicines based on the individual synergistic properties of each herb. The Jun 4 (monarch) component is the principal phyto-complex targeting the major symptom of the disease. It synergizes with the Chen (minister) herbs to support its therapeutic effects. The Zuo (assistant) medicinal reduces or eliminates possible adverse or toxic effects. Lastly, the Shi (courier) herbs facilitate the delivery of the principal components to desired sites (e.g., target organ), or facilitate the overall action of the other components. The whole formulae thus strive for a synergistic effect that targets active phytochemicals to their designated sites of action 8 . Furthermore, herbs can be administered through a diverse number of routes including oral intake, nasal inhalation, moxibustion, and herbal plasters (Fig. 1 24, 25 . The rich experience and herbal knowledgebase in fighting pandemics, are now explored here as a foundation 5 to further explore potential effective components and combinations in herbal medicines using more recent advances in data analysis.

The components were converted to Murcko scaffolds, dissecting molecules into ring systems, linkers, side chain atoms, and frameworks, by scaffold analysis 32 . SOM arranges the position of these structures in two-dimensional space, after which the dataset contains inherently similar structures and is suitable for inclusion and further cluster analysis 33 . Finally, the cluster map of the complete dataset was computer generated followed by visually assessing the largest common groupings of clusters, 6 by reducing the cluster numbers manually until all the disparate clusters without neighbors were included, but the main clusters remained 34 . (2018-007-02, "Feather Spray"©2018), the R function for Statistic Computing Platform: x86_64-pc-Linux-gnu (64-bit)] 35 and Pajek visualization 36 . Armeniacae Semen Amarum (Ku Xing Ren) has been used for the treatment of pain and inflammatory diseases such as asthma, bronchitis, emphysema, constipation, nausea, leprosy and leukoderma 77 . The herb has been used to reduce fever, relieve cough and quench thirst. Amygdalin is the major compound, which has been reported to have anti-inflammatory and analgesic effects in the inhibition of the COX-2 9 pathway 77 . Ephedrae Herba (Ma Huang) has traditionally been used in China for bronchial asthma, coughs, colds, flu, fever, edema, and arthralgias. The aerial parts of different Ephedra species contain from 0.02% to 3.4% of six optically active alkaloids. Ephedrine is the major isomer comprising 30%-90% of the total alkaloids 78 .

It enhances the release of norepinephrine from sympathetic neurons. The vasoconstrictor and bronchodilator effects explain the traditional use of Ephedra as a nasal decongestant and anti-asthmatic 78 . It was reported that the growth of H1N1 virus was inhibited when the cells were treated with its extract immediately after infection. Its inhibitory effect was completely or partially reversed by FeCl 3 , a tannin-reactive agent, suggesting that tannin is one of the active components in the extract 78 .

Citri Reticulatae Pericarpium (Chen Pi) promotes the circulation of Qi and is widely taken for coughing by drying dampness and expectorating phlegm 79 

We have explored 8 groups of core combinations by hierarchical clustering with 98

individual herbs whose modified mutual information is over 0.03, drawn from the 166 The sixth group focuses on expelling the pathogen and strengthening the spleen. The seventh group aims at tonifying Qi and nourishing Yin. The last one regulates Qi, 11 warms the interior, and relieves pain ( Fig. 4A and Supporting Information Table S6 .1).

The core combinations found here correspond with the traditional rules of herbal medicine composition and compatibility to reinforce particular functions in the treatment of COVID-19. For core combinations of chemical components, only herbs appeared twice or above and with modified mutual information over 0.1 were included, at last 12 core combinations were obtained. Some chemical components share the same category of traditional therapeutic effect (Fig. 2D) , which may signify potential synergistic effects. There are mutual groupings within these core combinations of herbal pairs and chemical components, thus supporting that the herbs and formulae containing these properties may act as potential candidates for COVID-19 treatment ( Fig. 4B and Table S6 .2).

1164 Murcko scaffolds and 40 aliphatic straight chain fragments were removed from the data and not considered further as they produced useless ring structures (Fig. 5A ).

SOM resulted in distinctly dispersed map of structural similarly components. Therefore, the dataset is diverse yet contains inherently similar structures suitable for inclusion and further cluster analysis. The clusters were self-organized (SOM) based on the ring system (Fig. 5B ). Analysis mapping of the nearest neighbours visually exhibited both diverse clusters and clearly associated families of structures (Fig. 5C) . Finally, 12 significant clusters with distinctly different structures were discovered, from which one representing each family were screened in silico for coronavirus activity, against approximately two million molecules using ChEMBL online database 82 (12%), 229E (16%) and NL63 (72%). Herbs containing the same pharmacophore clusters above, due to their chemical scaffold similarity and nearest neighbours, are more likely to behave pharmacologically similar. Therapeutically they may possess synergistic properties when used in combinations due to possessing chemical similarities, yet different when act on similar metabolic targets in slightly a different manner. This is the inverse analog of "shotgun chemistry" where similar compounds are applied en masse, often indiscriminately in either chemical, biological, DNA analysis or in drug discovery 83 . Further pharmacophore synergy investigations are encouraged and could be thought of "selective shotgun" chemistry based on these findings. Table   S7 ).

TCM asserts that pandemic diseases occur naturally and periodically as the climate or geographic situations change. TCM also emphasizes the enhancement of immune function to encourage the body to expel external pathogens. This is also done through the use of herbal formulations derived from careful analysis of syndromes likely to arise in a given year or season. This preventative strategy has also been useful in the treatment of COVID-19. In addition to assisting in the relief of symptoms for most patients in all stages, this approach is especially appealing due to the strong emphasis on both prevention and participation in the recovery of various disorders 79 . The prevention strategy appears in three parts: preventing occurrence, deterioration, and reoccurrence. TCM seems to have shown encouraging results in reducing the rate of mild illness, deterioration to severe stage and overall mortality as well as shortening total disease duration. When combined with modern biomedicine, herbal medicines could relieve hypoxemia and chronic obstructive pulmonary disease (COPD) 84 This study relies on computational analysis, which has the inherent weakness of not possessing aspects of qualitative judgment and "common sense", that human conducted analysis possesses. To offset this as much as practically possible, this 16 study's scope only focused on herbal medicines which have been developed, selected and proficiently used in practice over thousands of years. Additionally, it included herbs only which have been well studied, possessing "drug like" bioavailability, further bringing a "real-world", pharmacological information to the initial dataset.

The application of relatively novel computational, pattern recognition and in silico methodologies to a preselected knowledge base of herbal formulae most frequently used, in-part attempts to overcome the shortcomings of the network analysis and computational analysis approach. The 8 resulting pharmacophores and core combinations were based on firstly traditional use, secondly "drug like" characteristics and thirdly reported in silico bioactive pharmacophore structure.

Therefore, we propose they present the most likely candidates, taking the constraints of this study into account, to be included, in few multiples or in combination to support the composition of completely novel COVID-19 herbal formulae. The strengths of computational methods far outweigh their weaknesses. They bestow the ability to recognize and extrapolate useful patterns from herbal datasets which would otherwise take many years, if at all possible for human researchers to explore and uncover. The combination of this blended approach, as presented here, utilizes the combined advantages of both, to enable new useful knowledge obtained from a meaningful practical foundation.

The pharmacologically active ingredients of a phyto-complex are not always the original molecules in its natural state, but maybe their host-specific metabolites or molecular complexes formed following co-administration with other herbs. The multicomponent nature of traditional medicines leads to multiple potential molecular interactions, multiple targets, and a myriad of metabolic byproducts, which requires a more network-oriented and holistic approach 94 . The approach could be further refined by the application of omics technologies to optimize the synergistic effect of herbal remedies. With clinicians and basic researchers creating a database of personal therapeutic responses, continual improvements to the evolution and creation of herbal formulations would become possible 5 . Integration of Western and Chinese medical conceptual perspectives has enormous potential for constructing modern technological and social innovations. They are poised to merge within the arena of personalized medicine systems, wherein patients can take a greater role in managing their health and wellness 95 . In a time of increased international awareness of the environment, health and personal responsibility for mankind, an integration of East 17 and West should most likely be mutually beneficial 91 . As the global spread of COVID-19 continues to worsen, it is necessary to disseminate the many advantages of traditional medicines for the prevention and treatment of COVID-19. Bridging the gap between traditional and conventional medicine during this global COVID-19 spread is both timely and crucial. We sincerely hope that this study will serve as a reference to the global COVID-19 research community. 

Herbal formulae were collected from 26 protocols for treating COVID-19. Using complex system entropy and unsupervised hierarchical clustering, 8 core combinations and 10 formulae emerged as potential candidates. Following scaffold analysis and self organizing mapping, 12 clusters of molecules yielded 8 pharmacophore structures screened as pharmacological targets for inhibiting corona virus.

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The authors thank Dr. Zheng Yu 

All authors declare no conflict of interests.

