以本產芸香為基礎的天然物組合來同時緩解抗藥性細菌及禽類冠狀病毒

Combination of Natural Ingredients Based on Indigenous rue Simultaneously Alleviates Antimicrobial Resistant (AMR) Bacteria and Avian Coronavirus

10.6763/IJTCMHP.202211_6(2).0001

劉全喜(Liu, Chuan-Shi)；孫立庭(Sun, Li-Ting)；賴怡汝(Lai, Yi-Ru)；金立德(Chin, Li-Te)

本產芸香 ； 肉桂芸香鹼 ； 抗微生物 ； Indigenous rue ； harmine ； antimicrobial

中醫本草健康促進期刊

6卷2期（2022 / 11 / 15）

5 - 23

繁體中文

抗微生物製劑的抗藥性問題，一直都是公共衛生上被關注的議題，甚至於COVID-19（Coronavirus Disease-2019）疫情爆發更是如此。面對抗藥性問題，可從藥物開發以提供解方。本研究從本土芸香預防保健的豐富性來擴展實際應用，以指標成分肉桂芸香鹼（harmine）為基礎，搭配卵轉鐵蛋白（ovotransferrin，OTRF）作為加成劑，測試其複方的抗微生物活性。除收集臨床抗藥性菌株進行複方抑菌測試外，復以有禽類冠狀病毒（Avian coronavirus, AvCoV）之稱的傳染性支氣管炎病毒（infectious bronchitis virus, IBV）作為標的，觀測其改善病毒感染後胚蛋病變及存活情形。本研究使用傳染性支氣管炎病毒減毒疫苗IB 4-91作為起始病毒株，接種於胚蛋二輪後收集尿囊（allantoic）液以取得活化病毒。本產芸香萃取之肉桂芸香鹼在搭配卵轉鐵蛋白之下，對呼吸道分離的臨床菌株，不論是對格蘭氏陰性或是陽性菌皆具協同性的抑菌活性。對於病毒而言，活化後的傳染性支氣管炎病毒，較原始疫苗病毒株有更強之感染性與毒性，縮短了胚胎死亡時間；以複方處理後，可減少雞胚的病毒異常毒化情形，並且將存活率由33%提升至66～100%。以本產芸香萃取出的肉桂芸香鹼為主體，搭配卵轉鐵蛋白設計成複方，不僅具協同性作用，可抑制抗藥性臨床菌株，同時亦可抑制傳染性支氣管病毒，此廣效之抗微生物的效果，具有開發成外用消毒或治療藥物的潛力。

The problem of drug resistance arising from the use of antimicrobial agents has long been a public health issue and concern, especially in the era of the current outbreak of COVID-19 (Coronavirus Disease-2019), are obvious and profound. Faced the challenge of antimicrobial resistance (AMR) head-on, new drug development may provide reasonable solutions. The success of our previous works suggested that the indigenous rue may further provide the tool to conquer pathogen-bone diseases. This study was performed based on the combination of the index component - harmine, of the indigenous rue, together with ovotransferrin (OTRF) as an additive; to evaluate the antimicrobial activity. In addition to use clinical AMR strains for antibacterial testing, the infectious bronchitis virus (IBV), also known as avian coronavirus (AvCoV), was used to test the same combination for possible improvement of embryonic viral infection. Firstly, we used the attenuated vaccine strain IB 4-91 as the starting inoculated virus, and subsequently collected the allantoic fluid to obtain the activated virus. The harmine preparation extracted from the indigenous rue, when combined with ovotransferrin, had a synergistic antibacterial activity against clinical AMR bacteria isolated from the respiratory tract, irrespective the Gram stain patterns. Furthermore, the activated AvCoV had enhanced infectivity and toxicity in chicken embryos, and consequently shortens the lifespan of embryos. However, the harmine / ovotransferrin increased the survival rate from 33% to 66~100%. The harmine preparation extracted from the rue, compounded with ovotransferrin, demonstrated not only a synergistic effect, but also simultaneously alleviates AMR bacteria and AvCoV. Thus such an effect may have a potential to be further developed into a topical disinfectant or therapeutic drugs.

1. Alderton G, Ward WH, Fevold HL. 1946. Identification of the bacteria-inhibiting iron-binding protein of egg white as conalbumin. Arch Biochem 11:9-13.
2. Anonymous. 2022. COVID-19: U.S. Impact on Antimicrobial Resistance, Special Report 2022. https://dx.doi.org/10.15620/cdc:117915, Hyattsville, MD.
3. Bullen, JJ,Rogers, HJ,Griffiths, E(1978).Role of iron in bacterial infection.Curr Top Microbiol Immunol,80,1-35.
4. Casais, R,Dove, B,Cavanagh, D,Britton, P(2003).Recombinant Avian Infectious Bronchitis Virus Expressing a Heterologous Spike Gene Demonstrates that the Spike Protein Is a Determinant of Cell Tropism.Journal of Virology,77,9084-9089.
5. Cavanagh, D(2007).Coronavirus avian infectious bronchitis virus.Vet Res,38,281-297.
6. Chen, D,Su, A,Fu, Y,Wang, X,Lv, X,Xu, W,Xu, S,Wang, H,Wu, Z(2015).Harmine blocks herpes simplex virus infection through downregulating cellular NF-κB and MAPK pathways induced by oxidative stress.Antiviral Res,123,27-38.
7. Chen, D,Tian, X,Zou, X,Xu, S,Wang, H,Zheng, N,Wu, Z(2018).Harmine, a small molecule derived from natural sources, inhibits enterovirus 71 replication by targeting NF-κB pathway.Int Immunopharmacol,60,111-120.
8. Chin, L-T,Liu, K-W,Chen, Y-H,Hsu, S-C,Huang, L(2021).Cell-based assays and molecular simulation reveal that the anti-cancer harmine is a specific matrix metalloproteinase-3 (MMP-3) inhibitor.Comput Biol Chem,94,8.
9. El Ahmer, OR,Raza, MW,Ogilvie, MM,Weir, DM,Blackwell, CC(1999).Binding of bacteria to HEp-2 cells infected with influenza A virus.FEMS Immunol Med Microbiol,23,331-341.
10. Ellison, RT, 3rd,Giehl, TJ,LaForce, FM(1988).Damage of the outer membrane of enteric gram-negative bacteria by lactoferrin and transferrin.Infect Immun,56,2774-2781.
11. Engelich, G,White, M,Hartshorn, KL(2002).Role of the respiratory burst in co-operative reduction in neutrophil survival by influenza A virus and Escherichia coli.J Med Microbiol,51,484-490.
12. Gandhi, S,Klein, J,Robertson, AJ,Peña-Hernández, MA,Lin, MJ,Roychoudhury, P,Lu, P,Fournier, J,Ferguson, D,Mohamed Bakhash, SAK,Catherine Muenker, M,Srivathsan, A,Wunder, EA,Kerantzas, N,Wang, W,Lindenbach, B,Pyle, A,Wilen, CB,Ogbuagu, O,Greninger, AL,Iwasaki, A,Schulz, WL,Ko, AI(2022).De novo emergence of a remdesivir resistance mutation during treatment of persistent SARS-CoV-2 infection in an immunocompromised patient: a case report.Nature Communications,13,1547.
13. Giansanti, F,Massucci, MT,Giardi, MF,Nozza, F,Pulsinelli, E,Nicolini, C,Botti, D,Antonini, G(2005).Antiviral activity of ovotransferrin derived peptides.Biochem Biophys Res Commun,331,69-73.
14. Giansanti, F,Rossi, P,Massucci, MT,Botti, D,Antonini, G,Valenti, P,Seganti, L(2002).Antiviral activity of ovotransferrin discloses an evolutionary strategy for the defensive activities of lactoferrin.Biochem Cell Biol,80,125-130.
15. Guy, JS(2008).Isolation and propagation of coronaviruses in embryonated eggs.Methods Mol Biol,454,109-117.
16. Hiramatsu, K,Hanaki, H,Ino, T,Yabuta, K,Oguri, T,Tenover, FC(1997).Methicillin-resistant Staphylococcus aureus clinical strain with reduced vancomycin susceptibility.J Antimicrob Chemother,40,135-136.
17. Hutterer, C,Milbradt, J,Hamilton, S,Zaja, M,Leban, J,Henry, C,Vitt, D,Steingruber, M,Sonntag, E,Zeitträger, I,Bahsi, H,Stamminger, T,Rawlinson, W,Strobl, S,Marschall, M(2017).Inhibitors of dual-specificity tyrosine phosphorylation-regulated kinases (DYRK) exert a strong anti-herpesviral activity.Antiviral Research,143,113-121.
18. Maranan, MC,Moreira, B,Boyle-Vavra, S,Daum, RS(1997).Antimicrobial resistance in staphylococci. Epidemiology, molecular mechanisms, and clinical relevance.Infect Dis Clin North Am,11,813-849.
19. McGuinness, WA,Malachowa, N,DeLeo, FR(2017).Vancomycin Resistance in Staphylococcus aureus.Yale J Biol Med,90,269-281.
20. Mohammad Reza, VR,Hadjiakhoondi, A(2007).Cytotoxicity and antimicrobial activity of Harman alkaloids.J Pharmacol Toxicol,2,677-680.
21. Moloudizargari, M,Mikaili, P,Aghajanshakeri, S,Asghari, MH,Shayegh, J(2013).Pharmacological and therapeutic effects of Peganum harmala and its main alkaloids.Pharmacogn Rev,7,199-212.
22. Mota, NSRS,Kviecinski, MR,Felipe, KB,Grinevicius, VMAS,Siminski, T,Almeida, GM,Zeferino, RC,Pich, CT,Filho, DW,Pedrosa, RC(2020).β-carboline alkaloid harmine induces DNA damage and triggers apoptosis by a mitochondrial pathway: study in silico, in vitro and in vivo.Int J Funct Nutr,1,1.
23. Moura, DJ,Richter, MF,Boeira, JM,Pegas Henriques, JA,Saffi, J(2007).Antioxidant properties of beta-carboline alkaloids are related to their antimutagenic and antigenotoxic activities.Mutagenesis,22,293-302.
24. Navarini, AA,Recher, M,Lang, KS,Georgiev, P,Meury, S,Bergthaler, A,Flatz, L,Bille, J,Landmann, R,Odermatt, B,Hengartner, H,Zinkernagel, RM(2006).Increased susceptibility to bacterial superinfection as a consequence of innate antiviral responses.Proc Natl Acad Sci U S A,103,15535-15539.
25. Nenaah, G(2010).Antibacterial and antifungal activities of (beta)-carboline alkaloids of Peganum harmala (L) seeds and their combination effects.Fitoterapia,81,779-782.
26. Pechère, JC(2001).Macrolide resistance mechanisms in Gram-positive cocci.Int J Antimicrob Agents,18(Suppl 1),S25-S28.
27. Quinteros, JA,Markham, PF,Lee, S-W,Hewson, KA,Hartley, CA,Legione, AR,Coppo, MJC,Vaz, PK,Browning, GF(2015).Analysis of the complete genomic sequences of two virus subpopulations of the Australian infectious bronchitis virus vaccine VicS.Avian Pathology,44,182-191.
28. Ramakrishnan, S,Kappala, D(2019).Avian Infectious Bronchitis Virus.Recent Advances in Animal Virology,Singapore:
29. Sevoian, M,Levine, PP(1957).Effects of Infectious Bronchitis on the Reproductive Tracts, Egg Production, and Egg Quality of Laying Chickens.Avian Diseases,1,136-164.
30. Sjaak de Wit, JJ,Cook, JK,van der Heijden, HM(2011).Infectious bronchitis virus variants: a review of the history, current situation and control measures.Avian Pathol,40,223-235.
31. Sylvester, S,Kataria, JM,Dhama, K,Rahul, S,Bhardwaj, N,Tomar, S(2003).Purification of infectious bronchitis virus propagated in embryonated chicken eggs and its confirmation by RT-PCR.Indian J Comp Microbiol Immunol Infect Dis,24,143-147.
32. Wilson, R,Dowling, RB,Jackson, AD(1996).The biology of bacterial colonization and invasion of the respiratory mucosa.Eur Respir J,9,1523-1530.
33. Wu, J,Acero-Lopez, A(2012).Ovotransferrin: Structure, bioactivities, and preparation.Food Research International,46,480-487.
34. Zhou, F,Yu, T,Du, R,Fan, G,Liu, Y,Liu, Z,Xiang, J,Wang, Y,Song, B,Gu, X,Guan, L,Wei, Y,Li, H,Wu, X,Xu, J,Tu, S,Zhang, Y,Chen, H,Cao, B(2020).Clinical course and risk factors for mortality of adult inpatients with COVID-19 in Wuhan, China: a retrospective cohort study.Lancet,395,1054-1062.
35. 劉克威,孫立庭,金立德(2020)。芸香種子中肉桂芸香鹼成分分析與微體繁殖系統之建立。台灣農業研究，69，326-334。

1. 盧世乾,劉全喜,黃琳,金立德,周淑玲,李明駿(2023)。以本產芸香萃取之肉桂芸香鹼作為緩解e抗原陰性B型肝炎病毒材料之研究。中醫本草健康促進期刊，7(2)，5-27。