Abstract

The recent years there is a spurt in the interest regarding survival of Ayurvedic forms of medication. In the global perspective, there is a shift towards the use of medicine of herbal origin, as the dangers and the shortcoming of modern medicine have started getting more apparent, majority of Ayurvedic formulation are prepared from herbs. Syrup is very popular dosage form of cough and cold medications, ease of patient compliance. The objective of this study is to develop a herbal cough syrup and to check the antibacterial activity of the extract of seeds of Hedge mustard used for the formulation of herbal cough syrup against different bacteria (Staphylococcus aureus, E.coli, Salmonella sp., Pseudomonas aeruginosa , B.subtilis), and to evaluate the physicochemical parameter of cough syrup as well. The cough syrup formulated with simple syrup 66.67% w/v as sugar base. Quality of final herbal syrup was evaluated with different parameters: physical appearance (colour, odour, taste), pH and turbidity. The formulated syrup under gone stability studies and microbial test for 72 hours, no turbidity was observed and no microbial growth was seen.

Keywords

Introduction

Sisymbrium   officinale   (L.)   Scop.,   synonym Erysimum   officinale,    commonly   known    as hedge mustard in English, erísimo in Spanish, erisimo or erba cornacchia in Italian, and velar in French, is a medicinal plant that belongs to the  Brassicaceae  family.  This  species  could have  potential  for introduction  into  the  leafy vegetable     production    for    the     minimally processed or fresh-cut industry. S. officinale is a terophyte scapose plant with a reddish-violet erect trunk, that present a lot of trichomes and many  branches.  Basal  leaves  are different from  the  upper  ones  with  a  dentate  shape. Hedge    mustard    has    a    linear    racemose inflorescence; each flower has four small (1–2 mm)  yellow  petals;  the fruit  is  a  tiny  siliqua, close-fitting to the trunk.  Flowering  occurs  in Spring–Summer,  from   May  to  July–August, depending on the climate. Siliqua pods usually are pubescent, once they reach maturity they release  seeds.  Seeds  are  very  small,  each siliqua  can  contain  from  10  to  20  seeds.  S. officinale is endemic in the Eurasian continent and widespread in all Italian regions from 0 to 1000 m. above sea level ,and rarely up to 2400 m above sea level.¹

S. officinale is largely known as “singer’s plant” and   is   used   among   singers,   actors,   and  professionals who  use the voice for working.  The   therapeutic   activity   of   this   plant   is  attributed to  its sulfurated components.  Dried  flowering      aerial      parts      contain:      total glucosinolates  (0.63–0.94%), mucilage (13.5– 10.9%),   total  thiols   (8.9–10.2%),   and   total flavonoids        (0.50–0.56%).       The        main glucosinolate        in        S.        officinale        is  glucoputranjivine.²    historically,  the  sulphated  compounds   are    reputed   to    stimulate   the  mucosal  secretion   in  the  upper  respirator tract,  so  increasing  expectoration.  It  is  used mainly against the inflammations and catarrhs of the larynx, especially to combat hoarseness, as well as against cough,  pulmonary catarrh, etc.,   and   scurvy   too.   The   fresh   plant   is preferably used. The  pharmacological activity of     Sisymbrium     shows     anti-inflammatory, analgesic, antitussive, myorelaxant and broad spectrum antimicrobial properties.3

It has also been found that the seeds contain small quantities of cardenolides. It is good for all    diseases    of    the    chest    and    lungs, hoarseness  of  voice.  The juice  made  into  a syrup with honey or sugar, is no less effectual for all other coughs, wheezing and shortness of  breath,  the  seed  is  held  to  be  a  special remedy  against   poison  and  venom.   It  was formerly used for hoarseness, weak lungs and to help the voice. Herbalists use the juice and flowers      forbronchitisandstomachailments, among   other   uses,   and   as   a   revitalizer. InTibetan  medicineit  is  used  to  repress  the symptoms of food poisoning.

In  recent  years,   plant  derived   products  are increasingly  being  sought  out  as  medicinal products,  nutraceuticals  and  cosmetics  and are   available   in   health   food   shops   and pharmacies    over    the    counter    as    self- medication or also as drugs prescribed in the non-allopathic    systems.    Herbal    medicines widely used  in  health-care  in  both developed and    developing    countries     are     complex chemical  mixtures  prepared  from  plants  and are limited in their effectiveness because they are poorly absorbed when taken orally. Cough Syrup  is  liquid  dosage  form;  the  oral  use  of liquid   pharmaceutical   has   generally   been justified on the basis of ease of administration to  those  individuals  who  have  difficulties  in swallowing  solid  dosage  forms.   Syrup  is  a concentrated  mixture  of  sugar  and  purified water.  The  high  sugar  content  distinguishes syrups from  other types  of solutions.  Syrups may or may not contain  medication or added flavouring      agents.      Syrups      without      a medication,  but  with  a  flavouring  agent,  are called   non-medicated   or   flavoured   syrups. Flavoured syrups  are  often  used as vehicles for unpleasant tasting medications: the result is medicated  syrup.  The  high  amount  of  sugar present    in    syrups    predisposes    them    to bacterial contamination, so they often contain a  preservative.4   According  to  an  estimate  of the World  Health Organization  (WHO); about 80%  of the world  population  still  uses  herbs and   other   traditional   medicines   for   their primary health care needs. Herbal formulations have   reached   widespread   acceptability   as therapeutic agents for diabetics, arthritics, liver diseases, cough remedies, memory enhancers and adoptogens.5 As per WHO definition, there are three kinds of herbal medicines: raw plant material,    processed    plant    material    and medicinal  herbal  products.  Herbal  drugs  are finished  labelled  products  that  contain  active ingredients  such  as   aerial  or   underground parts   of   plant   or   other   plant   material   or combination thereof, whether in the crude state or as plant preparations.⁶

MATERIAL AND METHODS

Plant material

The seeds  of  Hedge  mustard were collected from    local    market    of    Lucknow.    It    was authenticated      by      Acube       Lifesciences, Lucknow.

Method of preparation of extract

The  seeds  were  cleaned,  shade  dried  and powdered mechanically and stored in air tight containers. The extraction was carried out by maceration.   About   5   gm   of   powder   was extracted with 80% methanol. The extract was kept for 48 -  72  hours  and after that  it was filtered.    The    extract    was    preserved    in refrigerator at 4°C.

Antimicrobial activity

In vitro antibacterial activity of the methanolic extract was studied against gram +ve and -ve bacterial  strains  by  the  agar  well  diffusion method.   Nutrient   Agar   was   used   as   the bacteriological    medium.The    Nutrient    agar media was melted and cooled to 48-50 ºC was then  poured  into  sterile  petridishes to  give  a solid plate. Wells were prepared in the seeded agar  plates. The test  compound  (50µl,  75  µl and  100  µl)  was  introduced  in  the  well.  The plates were incubated overnight at 37oC. The antimicrobial   spectrum   of   the   extract   was determined for the bacterial species in terms of zone sizes around each well. The diameters of zone of inhibition produced by the agent were compared    with    those    produced    by    the commercial control antibiotic ciprofloxacin.

Phytochemical screening

Qualitative      phytochemical      analysis      for secondary metabolites was carried out for the crude extracts as per standard methods.

a) Saponin

5ml  distilled  water  was  added  to   1ml  plant extract and then shaken well, froth formation took   place.   Stability   of   froth   confirms   the presence of saponin in plant extract.

b) Tannin

1ml 5?Cl3 was added to 1ml plant extract. Appearance of dark blue, black or dark green confirms presence of tannin in plant extract.

c) Flavonoid

2ml 1% NaOH was added to 1ml plant extract, presence of yellow colour indicates the in plant extract.

d) Carbohydrate

1ml Fehling A and 1ml Fehling B was added to 2ml  plant  extract  and  then  test  tube  was heated in water bath for 20 min. Appearance of red   precipitate   confirms   the    presence   of carbohydrate in plant extract.

e) Protein

1ml of 1% CuSo4  and  1ml  of  1%  NaOH was added  to  2ml   plant  extract.  Appearance   of purple color confirms the presence of protein in plant extract.

f) Alkaloid

1ml  iodine  was  added  to   1ml  plant  extract. Appearance   of   reddish    brown    precipitate confirms  the   presence  of  alkaloid   in   plant extract.

g) Starch

1ml  iodine  was  added  to   1ml  plant  extract. Appearance of blue or black color confirms the presence of starch in plant extract.

h) Fat Test

1mL of distilled water and few drops of ethanol were added to 1mL of plant extract.The white color precipitate formed showed the presence of fat in the plant extract.

 i) Terpenoid Test

250µl  chloroform  was  added  to  500  µl  plant extract then 625 µl Conc. H2SO4 was added to the solution. Reddish brown ppt, of the solution confirms presence of terpenoids.

Method   of   preparation   of   simple   syrup (usp)

666.7 g of Sucrose was weighed and added to purified water and heated until it dissolved with occasional stirring.

Evaluation of herbal syrup Physicochemical parameters

The  herbal  syrup  was  evaluated  for  various physicochemical parameters such as physical appearance (colour, odour, taste), pH.

       a) Color examination

Five   ml  final  syrup  was  taken   into  watch glasses and placed against white back groundinwhite tube light. It was observed for itscolor by naked eye.

        b) Odour examination

Two ml of final syrup was smelled individually. The time interval among two smelling was kept 2  minutes  to  nullify  the  effect  of  previous smelling.

        c) Taste examination

A pinch of final syrup was taken and examined for its taste on taste buds of the tongue.

        d) Determination of pH

Placed an accurately measured amount 10 ml of the final syrup in a 100 ml volumetric flask and  made  up  the  volume  up  to  100  ml  with distilled water. The solution was sonicated for about 10 minutes. pH was measured with the help of digital pH meter.

       Stability testing

Stability testing  of the  prepared  herbal syrup was  performed  on   keeping  the  samples  at accelerated temperature conditions.  The final syrup was taken in culture tubes and were kept at   accelerated   temperature   at   4°C,   Room temperature    and    47°C    respectively.    The samples      were      tested      for      all      the physicochemical    parameters,   turbidity   and homogeneity at the interval of 24 hr, 36hr and 72 hr to observe any change.

       RESULTS AND DISCUSSION

The basic objective of this work was to develop herbal  cough  syrup  from   seeds  of   Hedge mustard. It has also been found that the seeds contain  small  quantities  of cardenolides.  The development of such herbal syrup will mark an important advancement in the area of Coughs, Use   as   a   gargle   or   mouthwash,   Chronic bronchitis,    Urinary tract diseases,    Swelling (inflammation)   of   the    gallbladder   and   Other conditions. The       present       investigation       examines development and evaluation of herbal syrup. Phytochemical screening investigations on the phytochemical  screening  of  Hedge  mustard extracts       revealed      the       presence      of carbohydrates, alkaloids, tannin, saponin, fat, terpenoid and flavonoids, which are known to be  biologically active. These  metabolites  can exert  antimicrobial  activity  through  different mechanisms. The   antimicrobial   activity   of methanolic  extract  of  hedge  mustard  seeds were     tested     against     bacterial     strains (Staphylococcus   aureus,   E.coli,   Salmonella sp.,  Pseudomonas  aeruginosa  ,  B.subtilis  ), the extract showed zone of inhibition.

The formulated herbal syrup was found to be clear without particles and sweet in taste. The developed   herbal  syrup  was  evaluated  for stability studies for 24, 36 and 72 hours with varying temperature of 4°C, room temperature and 47°C. There was no change observed in physical appearance (colour, odour, taste), pH of  the  formulated  syrup.   It  did  not  produce turbidity  at  lower  temperature  of 4°C.  It  was clear  homogenous  liquid  without  turbidity  at higher storage temperature of 47°C too. Thus it can be concluded that the herbal syrup was in suitable form which was developed.

CONCLUSION

In recent years there is a spurt in the interest regarding   survival   of   Ayurvedic   forms    of medication. In the global perspective, there is a shift towards the use of medicine of herbal origin, as the dangers and the shortcoming of modern  medicine  have  started  getting  more apparent, majority of Ayurvedic formulation are prepared  from  herbs.  Syrup  is  very  popular dosage form  of cough and cold  medications, ease of patient compliance.

Sisymbrium     officinale,     known     as Hedge mustard, (formerly Erysimum   officinale)   is   a plant    in    the    familyBrassicaceae.    Hedge mustard   contains   an   essential   oil,   rich   in sweet-smelling sulphur compounds, consisting mainly  of  glucosinolates.   It  has  also  been found that the seeds contain small quantities of cardenolides. The  Phytochemical  properties of  the  extract were    evaluated.    The    methanolic    extract showed       the  presence       of       various phytochemicals  such  as,  Flavonoid,  Alkaloid, Saponin,  Protein,  Carbohydrate,  Starch,  Fat, Terpenoid.  The  extract  was  tested  for  anti- microbial  activity.  Methanolic  extract  showed good  activity  at  different  concentration.  The methanolic extract were formulated as a herbal cough  syrup  of  one   part  of  decoction  was mixed  with  five  parts  of  simple  syrup  (1:5), peppermint oil (0.02%) and required quantity of Sodium  benzoate  (0.2%)was     added  to  the above  mixture  (Sodium  benzoate)  act  as  a preservative  to  the  above  mixture.  Solubility was   checked   by   observing   the   clarity   of solution  visually.  The  final  herbal  syrup  was then subjected for evaluation.

ACKNOWLEDGEMENT

Authors  are  grateful  to  Acube  Lifesciences, Lucknow for the support and providing facilities to carry out this work.

REFERENCE

1. Guarise  M,  Borgonovo  G,  Bassoli  A and  Ferrante  A.  Evaluation  of  Two Wild  Populations  of  Hedge  Mustard (Sisymbrium officinale (L.) Scop.) as a Potential           Leafy           Vegetable. Horticulturae. 2019;5:13.
2. Carnat   A,   Fraisse   D,   Carnat   AP, Groubert     A     and     Lamaison     JL. Normalization    of    hedge    mustard, Sisymbrium    officinale    L.    Annales Pharmaceutiques                Françaises. 1998;56: 36–39.
3. Calcinoni    O    Sisymbrium.    Singers’ Plant  Efficacy  in  Reducing  Perceived Vocal   Tract    Disability.    Journal    of Otolaryngology-ENT             Research. 2017;8(2):00243.
4. Kaushik A, Chauhan V  and Dr Sudha. Formulation And Evaluation Of Herbal Cough  Syrup.   European  Journal  of Pharmaceutical         And          Medical Research. 2016;3(5):517-522.
5. Michael M. Herbal Formulas For Clinic and Home. Bisbee, AZ 85603. 1995;1- 34.
6. Choudhary    N  and  Sekhon  BS.  An overview     of     advances     in     the standardization   of   herbal   drugs.   J Pharm Educ Res. 2011;2(2):55-70.
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Table 1: Result of Phytochemical Tests

Table 2: Result of Antibacterial Tests

Table 3: Result of Chromatography

Table 4: Formulation of hand sanitizer 
       
       
           
       
    
    

Table 6: MIC against Palm bacteria,S aureus, B. Subtilis and E. coli

Table 7: Comparison of Different Hand Sanitize