An experimental study to evaluate galactagogue activity of Bidali (Dioscorea pentaphylla Linn.) with special reference to Stanya Janana

Akshatha Somayaji K; Giri Prashanth KG; Suresh Janadri; Shiva Manjunath MP; Seema Pradeep; Manjunatha P Mudagal

doi:10.21760/jaims.9.1.4

E-ISSN:2456-3110

Research Article

Dioscorea pentaphylla

Journal of Ayurveda and Integrated Medical Sciences

20249 Volume 9 Number 1 January

Publisher

www.maharshicharaka.in

An experimental study to evaluate galactagogue activity of Bidali (Dioscorea pentaphylla Linn.) with special reference to Stanya Janana

Somayaji K A^1*, Prashanth KG G², Janadri S³, Manjunath MP S⁴, Pradeep S⁵, Mudagal MP⁶

DOI:10.21760/jaims.9.1.4

^1* Akshatha Somayaji K, Final Year Post Graduate Scholar, Department of Pg Studies in Dravyaguna, Sri Sri College of Ayurvedic Science and Research, Bengaluru, Karnataka, India.

² Giri Prashanth KG, Professor, Department of PG Studies in Dravyaguna, Sri Sri College of Ayurvedic Science and Research, Bengaluru, Karnataka, India.

³ Suresh Janadri, Assistant Professor, Department of Pharmacology, Acharya BM Reddy College of Pharmacy, Bengaluru, Karnataka, India.

⁴ Shiva Manjunath MP, Senior Scientist, Department of PG Studies in Dravyaguna, Sri Sri College of Ayurvedic Science and Research, Bengaluru, Karnataka, India.

⁵ Seema Pradeep, Professor and HOD, Department of PG Studies in Dravyaguna, Sri Sri College of Ayurvedic Science and Research, Bengaluru, Karnataka, India.

⁶ Manjunatha P Mudagal, Principal, Acharya BM Reddy College of Pharmacy, Bengaluru, Karnataka, India.

More than 8 Lakh infantile deaths in India are attributed to inadequate breastfeeding, which is correlated with a prevalence of 45% to 53.6% cases of oligogalactia. Bidali (Dioscorea pentaphylla Linn.) is one among the two varieties of Vidari as described by Acharya Dalhana, with Stanya Vardhaka Karma. Hence the experimental evaluation of galactagogue activity of Bidali (Dioscorea pentaphylla Linn.) was carried out using milk estimation by the means of pup weight and weight gain along with estimation of serum prolactin and histology of breast tissue. Assessment was done by dam weight, daily milk yield by the means of pup weight and weight gain, serum prolactin estimation and histology of breast issue. The treatment groups have shown positive result in dose dependent manner in the parameters; Weight of Dams, and Weight of Pups. Histology reports re-emphasize the same; the development of Tubulo-alveolar unit and intraluminal secretions were more in group treated with higher dose than that of group treated with lower dose of Bidali (Dioscorea pentaphylla Linn.).

Keywords: Bidali, Dioscorea pentaphylla Linn., Galactagogue, Stanya Janana

Corresponding Author	How to Cite this Article	To Browse
Akshatha Somayaji K, Final Year Post Graduate Scholar, Department of Pg Studies in Dravyaguna, Sri Sri College of Ayurvedic Science and Research, Bengaluru, Karnataka, India. Email:	Somayaji K A, Prashanth KG G, Janadri S, Manjunath MP S, Pradeep S, Mudagal MP, An experimental study to evaluate galactagogue activity of Bidali (Dioscorea pentaphylla Linn.) with special reference to Stanya Janana. J Ayu Int Med Sci. 20249;9(1):32-45. Available From https://jaims.in/jaims/article/view/2906

Manuscript Received	Review Round 1	Review Round 2	Review Round 3	Accepted
2023-11-14	2023-11-20	2023-11-24	2023-11-30	2023-12-24
Conflict of Interest	Funding	Ethical Approval	Plagiarism X-checker	Note
None declared	Nil	Yes	21.98%

© 20249by Somayaji K A, Prashanth KG G, Janadri S, Manjunath MP S, Pradeep S, Mudagal MPand Published by Maharshi Charaka Ayurveda Organization. This is an Open Access article licensed under a Creative Commons Attribution 4.0 International License https://creativecommons.org/licenses/by/4.0/ unported [CC BY 4.0].

J Ayu Int Med Sci 20249;9(1)32

Download PDF Back To Article Introduction Materials and
Methods Observations and
Result Discussion Conclusion References

Akshatha Somayaji K. et al. Galactagogue activity of Bidali (Dioscorea pentaphylla Linn.)

Introduction

Inadequate breastfeeding contributes to a prevalence of 45% to 53.6% incidences of Oligogalactia, which is responsible for about 8 Lakhs deaths in India.^[1] Despite many medications used in contemporary medicine claim to be beneficial, but they may also have long-term negative effects. For instance: Several gastrointestinal issues, insomnia, severe depression, and seizures are all brought on by the Metoclopramide (Dopamine antagonist), which also causes seizures in infants who consume breast milk from a mother who has been treated for them.^[2] Ayurveda explains many drugs that enhance the production of Stanya as Stanya Janana/Stanya Vardhaka.^[3] One such drug is Vidarikanda (Pueraria tuberosa D.C).^[4]Acharya Dalhana in Sushruta Samhita has described two varieties of Vidarikanda,^[5]one with the elongated tubers-Dirghakanda and the other with tuber resembling the foot of an Elephant-Hastipadavat, Bidali (Dioscorea pentaphylla Linn.), has the elongated tubers is the variety of Vidarikanda.^[4,6] Vidarikanda (Pueraria tuberosa D.C) is listed under the nearly-threatened red listed important medicinal plant species of India,^[7] whereas Bidali is listed under the least concerned plants, which is available throughout India,^[8] is also consumed as both staple food^[9]and as medicine to treat many health ailments by the Tribals as per the folklore claim.^[10] Hence, the galactagogue activity of Bidali - Dioscorea pentaphylla Linn. through experimental evaluation was intended to be studied.

Materials and Methods

Collection and Authentication

Bidali Kanda (Dioscorea pentaphylla Linn.) fresh samples were collected from Perdur, Udupi &, Badamanavarathekaval, Bengaluru and were authenticated by Dr. Shiva Manjunath M.P, Senior Scientist, Dept. of Dravya Guna at SSCASR, Bangalore.

Preparation of Kashaya

The authenticated Bidali Kanda (Dioscorea pentaphylla Linn) fresh sample was cleaned with water to remove dust and foreign particles, cut into small pieces and was sun dried. Dry sample was checked for any impurities, foreign matter and preserved in air tight container. Dried sample of Bidali

(Dioscorea pentaphylla Linn.) was made into a coarse powder in Laboratory, Department of PG Studies in Dravyaguna, Sri Sri College of Ayurvedic Science and Research, Bengaluru. The obtained coarse powder was collected and stored in zip lock cover and kept inside an air tight container.

Kashaya was prepared daily and administered in fresh form orally. General method of Kashaya preparation according to Acharya Sharangadhara^[11] was followed i.e., 1 part of drug, 8 parts of water reduced to 1/4^th (1:8-1/4) and was then filtered using a Khora cloth.

Experimental Study

The experimental procedure was carried out in accordance with the ethical guidelines for animals proposed by CPCSEA, Government of India. The Animal ethics clearance was obtained from Department of Pharmacology, Acharya & B.M Reddy College of Pharmacy, Bengaluru as per the protocol outlined in publication of the Committee for the Purpose of Control and Supervision of Experiments on Animals Standards Guidelines (CPCSEA) and approval was obtained from Institutional Animal Ethics Committee (IAEC) with reference no: IAEC/ABMRCP/2023-24/2

Source of Animals

From Biogen Laboratory Animal Facility, Bengaluru, twenty-four healthy mature female Wistar albino rats required for the study were procured.

Acute Toxicity Study^[12]

Acute oral toxicity - Limit test of Bidali Kanda (Dioscorea pentaphylla Linn.) Kashaya was conducted according to OECD guidelines 425. Nulli-parous, non-pregnant, healthy Wistar rats of 10-12 week-old, weighing about 180-200g were selected.

A single dose of Bidali (Dioscorea pentaphylla Linn.) was given in the dose of 2000mg/Kg p.o., was given and observed for 14 days.

For 14 days, a single dosage of Bidali (Dioscorea pentaphylla Linn.) at a dose of 5000 mg/Kg p.o. was administered and monitored.

Animals were monitored every 30 minutes for the first 4 hours after dosing, then on occasion for the next 24 hours, and then every day for the next 14 days.

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Akshatha Somayaji K. et al. Galactagogue activity of Bidali (Dioscorea pentaphylla Linn.)

Table 1: The table denotes the observation particulars during the Acute Toxicity Study.

Observed for
Behavioural Profile	Spontaneous, restlessness, irritability, and fearfulness
Neurological Profile	Spontaneous activity, reactivity, touch response, pain response and gait
Autonomic Profile	Defecation and urination
Other Changes	Change in skin or fur, eyes, mucosa and respiratory, Somatomotor and tremors, convulsion, diarrhoea, lethargy, sleep, coma, changes in the weight and any mortality.

Galactagogue Activity^[13]

Housing and feeding

The standard guidelines for the housing of the animals mentioned in the OECD (Organization for Economic Co-operation and Development) 425 Guidelines were followed- Animals were maintained in cages with paddy husk as bedding and housed at an ambient temperature 24 0C and humidity with 12-hours light and 12-hours dark cycles. Animals had free access to food and portable water.

Preparation of animals and Grouping

Randomly selected rats were divided into 4 groups consisting of 6 rats in each group. Each rat was marked for their individual identification as C (C1, C2, up to C6), S (S1, S2, up to S6), TL (TL1, TL2, up to TL6) & TH (TH1, TH2, up to TH6). The cages were labeled with the experiment name, dosage group and number of rats in that group. (Figure 5)

Table 2: The table represents the grouping of animals

SN	Group	Drug used	No of rats
1.	Normal control (C)	(normal diet and portable water)	6
2.	Standard control (S)	Vidari (Pueraria tuberosa D.C-1000mg/kg)	6
3.	Trial drug in Lower Dose (TL)	Bidali (Dioscorea pentaphylla Linn. - 500mg/Kg)	6
4.	Trial drug in Higher Dose (TH)	Bidali (Dioscorea pentaphylla Linn. - 1000mg/Kg)	6

Study Protocol

For weight gain of pups

24 lactating dams weighing 200 – 250 g at the beginning of lactation and suckling their pups were used for this experiment. Females were divided into 4 experimental groups and received normal diet and portable water (n=6), the standard group were administered with Kashaya of Pueraria tuberosa D.C (1000mg/Kg) (n=6) and 2 trial groups were administered with the Kashaya of Dioscorea pentaphylla Linn. In D1 and D2 doses (D1 and D2 were calculated after Acute Toxicity Study:

D1-Lower dose and D2-Higher dose). All animals were treated daily, starting on the evening of day 1 of lactation. The Kashaya was administered orally with a gavage syringe each day at 10AM. Milk production was measured from 1 day to day 15 of lactation. Milk yield and body weight of dams, and weight gain of pups were measured each day with an electronic balance. Every day during the study period, the pups were weighed (Figure 8) at 9AM (w1) and subsequently isolated from their mother for 4 h (Figure 7) (Sampson & Jansen 1984). At 12PM, the pups were weighed (w2), and returned to their mother and were allowed to feed for 1 h. At 1PM, they were weighed (w3). Milk yield was estimated as w3 – w2. Daily milk yield was corrected for weight loss due to metabolic processes in the pup (respiration, urination, and defecation) during suckling. The value used was (w2 – w1)/4. This value was then be multiplied by the number of suckling hours per day and added to the daily suckling gain (Sampson & Jansen 1984). Daily weight gain of pups was calculated from the pup weight at w2.

Finally, the milk yield was calculated by the following formula;

Milk yield (g) = (W3-W2) + [(W2-W1)/4]

Where; (W3-W2)-weight gain of pups after lactating/feeding

(W2-W1)/4-weight loss due to metabolic processes in the pups

For Serum Prolactin Concentration

The samples were collected on 1^st, 14^th and 21^st day of the study and were estimated by ELISA method from two rats from each group.

For Histopathology of breast tissue

At the end of study, the breast tissue was dissected under local anaesthesia from three rats from each group.

Parameters of the study

Physical parameters; weight of the dams and weight of the pups. Biochemical parameters; Serum prolactin concentration- was done on 1^st, 14th and 21^st day on two rats from each group. Histopathological assessment: Histopathological study of breast tissue-Was done at the end of study; the samples were being collected from three rats from each group.

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Akshatha Somayaji K. et al. Galactagogue activity of Bidali (Dioscorea pentaphylla Linn.)

Statistical analysis

The collected Data was analysed by repeated measures analysis of variance, Friedman repeated measures analysis of variance and Friedman Repeated measures analysis of variance on ranks followed by Holm Sidak method and Tukey’s test. P value < 0.05 was being considered as statistically significant and Sigma Stat 3.1 was used for statistical analysis.

Observations and Result

1. Acute Toxicity Study (Table 3-6)

There were no changes observed either physically or behaviorally from 0^th day to 14^th day at the dose of 2000mg/Kg p.o. of Bidali (Dioscorea pentaphylla Linn.). Further, on the dose of 5000mg/Kg p.o., of Bidali (Dioscorea pentaphylla Linn.) no significant physical and behavioral changes were noted.

Table 3: Changes recorded in animals during Acute Toxicity Study of Bidali (Dioscorea pentaphylla Linn.) at the dose of 2000mg/Kg p.o., orally on 0^th day, 7^th, and 14^th day.

SN	Signs & Symptoms		0th day				7th day	14th day
SN	Signs & Symptoms		Basal	2h	4h	24h	7th day	14th day
1.	General impression		N	Active	Active	N	Active	Active
2.	Increased motor activity		-	-	-	-	-	-
3.	Convulsion	Tonic	-	-	-	-	-	-
3.	Convulsion	Clonic	-	-	-	-	-	-
4.	Straub’s reaction		-	-	-	-	-	-
5.	Muscle spasm		-	-	-	-	-	-
6.	Catatonia		-	-	-	-	-	-
7.	Opisthotonos		-	-	-	-	-	-
8.	Hyperesthesia		-	-	-	-	-	-
9.	Decreased motor activity		-	-	-	-	-	-
10.	Muscle relaxation		-	-	-	-	-	-
11.	Anesthesia		-	-	-	-	-	-
12.	Arching and rolling		-	-	-	-	-	-
13.	Lacrimation		-	-	-	-	-	-
14.	Diarrhea		-	-	-	-	-	-
15.	Writhing		-	-	-	-	-	-
16.	Salivation	Viscid	-	-	-	-	-	-
16.	Salivation	Watery	-	-	-	-	-	-
17.	Respiration	Depression	-	-	-	-	-	-
17.	Respiration	Failure	-	-	-	-	-	-
18.	Skin color	Blanching	-	-	-	-	-	-
		Cyanosis	-	-	-	-	-	-
		Vasodilatation	-	-	-	-	-	-
19.	Grip strength		N	N	N	N	N	N
20.	Visual placing response		N	N	N	N	N	N
21.	Tail pinch response		N	N	N	N	N	N
22.	Auditory response		N	N	N	N	N	N
23.	Mucus membrane		N	N	N	N	N	N
24.	Piloerection		N	N	N	N	N	N

Table 4: CNS & ANS changes recorded during the acute toxicity study of Bidali (Dioscorea pentaphylla Linn.) at the dose of 2000mg/Kg p.o. on 0^th, 7^th and 14^th day.

SN	Particulars		0th day				7th day	14th day
SN	Particulars		Basal	2h	4h	24h	7th day	14th day
1.	Exitus		-	-	-	-	-	-
2.	CNS depression	Hypo activity	-	-	-	-	-	-
		Passivity	-	-	-	-	-	-
		Relaxation	-	-	-	-	-	-
		Ataxia	-	-	-	-	-	-
		Narcosis	-	-	-	-	-	-
3.	ANS	Ptosis	-	-	-	-	-	-
3.	ANS	Exophthalmos	-	-	-	-	-	-
4.	CNS stimulation	Hyperactivity	-	-	-	-	-	-
		Irritability	-	-	-	-	-	-
		Stereotypy	-	-	-	-	-	-
		Tremors	-	-	-	-	-	-
		Convulsion	-	-	-	-	-	-
		Straub tail	-	-	-	-	-	-
5.	Others		N	N	N	N	N	N

Table 5: Changes recorded in animals during 0^th,7^th, 14^th day of Acute Toxicity Study of Bidali (Dioscorea pentaphylla Linn.) at the dose of 5000mg/Kg p.o., orally.

SN	Signs & Symptoms		0th day				7th day	14th day
SN	Signs & Symptoms		Basal	2h	4h	24h	7th day	14th day
1.	General impression		N	Active	Active	N	Active	Active
2.	Increased motor activity		-	-	-	-	-	-
3.	Convulsion	Tonic	-	-	-	-	-	-
3.	Convulsion	Clonic	-	-	-	-	-	-
4.	Straub’s reaction		-	-	-	-	-	-
5.	Muscle spasm		-	-	-	-	-	-
6.	Catatonia		-	-	-	-	-	-
7.	Opisthotonos		-	-	-	-	-	-
8.	Hyperesthesia		-	-	-	-	-	-
9.	Decreased motor activity		-	-	-	-	-	-
10.	Muscle relaxation		-	-	-	-	-	-
11.	Anesthesia		-	-	-	-	-	-
12.	Arching and rolling		-	-	-	-	-	-
13.	Lacrimation		-	-	-	-	-	-
14.	Diarrhea		-	-	-	-	-	-
15.	Writhing		-	-	-	-	-	-
16.	Salivation	Viscid	-	-	-	-	-	-
16.	Salivation	Watery	-	-	-	-	-	-
17.	Respiration	Depression	-	-	-	-	-	-
17.	Respiration	Failure	-	-	-	-	-	-
18.	Skin color	Blanching	-	-	-	-	-	-
		Cyanosis	-	-	-	-	-	-
		Vasodilatation	-	-	-	-	-	-
19.	Grip strength		N	N	N	N	N	N
20.	Visual placing response		N	N	N	N	N	N
21.	Tail pinch response		N	N	N	N	N	N
22.	Auditory response		N	N	N	N	N	N
23.	Mucus membrane		N	N	N	N	N	N
24.	Piloerection		N	N	N	N	N	N

Table 6: CNS & ANS changes recorded during the 0^th, 7^th, 14^th day of acute toxicity study of Bidali (Dioscorea pentaphylla Linn.) at the dose of 5000mg/Kg p.o.

SN	Particulars		0th day				7th day	14th day
SN	Particulars		Basal	2h	4h	24h	7th day	14th day
1.	Exitus		-	-	-	-	-	-
2.	CNS depression	Hypo activity	-	-	-	-	-	-
		Passivity	-	-	-	-	-	-
		Relaxation	-	-	-	-	-	-
		Ataxia	-	-	-	-	-	-
		Narcosis	-	-	-	-	-	-
3.	ANS	Ptosis	-	-	-	-	-	-
3.	ANS	Exophthalmos	-	-	-	-	-	-
4.	CNS stimulation	Hyperactivity	-	-	-	-	-	-
		Irritability	-	-	-	-	-	-
		Stereotypy	-	-	-	-	-	-
		Tremors	-	-	-	-	-	-
		Convulsion	-	-	-	-	-	-
		Straub tail	-	-	-	-	-	-
5.	Others		N	N	N	N	N	N

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Akshatha Somayaji K. et al. Galactagogue activity of Bidali (Dioscorea pentaphylla Linn.)

Dose fixation:

As there was no toxicity observed at the dose of 5000mg/Kg p.o., the Lower dose was fixed as 500mg/Kg p.o., and Higher dose as 1000mg/Kg p.o..

2. Estimation of Milk Yield by the means of Pup Weight and Weight Gain (Galactagogue activity):

Assessment criteria and results:

1. Weekly average of weight of Dams

Table 7: Table showing the changes in weekly weight of Dams in different groups.

Groups	Control	Standard(Pueraria tuberosa D.C-1000mg/kg)	TL(Dioscorea pentaphylla Linn. -500mg/kg)	TH(Dioscorea pentaphylla Linn-1000mg/kg)	Significance
Week 1	181.263 ±7.326	180.792 ±8.275	194.667 ±4.763	181.155 ±9.577	NS
Week 2	189.175 ±6.760	188.015 ±8.959	225.517 ±17.693	186.318 ±4.947	NS
Week 3	197.023 ±4.739	198.825 ±9.736	241.682 ±18.003	205.752 ±8.709	NS
Significance	HS (P <0.001)	HS (P <0.001)	S (P <0.006)	HS (P <0.001)

The above table describes the weekly change in the body weight of Dams, expressed as mean ±SEM, (n=6), where, TH-- Trial drug (Dioscorea pentaphylla Linn.) in Higher dose (1000mg/Kg p.o.) and TL- Trial drug (Dioscorea pentaphylla Linn.) in Lower dose (500mg/Kg p.o.).

The statistical analysis was done by using one-way repeated measure analysis of variance, between the groups there was no Statistical significance seen even though there was apparent difference were observed (NS-Not Significant).
Within the groups, the statistical analysis was done using one-way repeated measures analysis of variance, significant after the analysis of variance followed by Holm-Sidak method, where highly significant difference was seen in Control, Standard and Trial Drug with Higher Dose group with P<0.001 and Significant difference was observed with P<0.006.

1. Milk Estimation by the means of Pup Weight and Weight gain
2. Milk Estimation by means of Pup weight:

Table 8A: Daily milk production by the means of weight gain of Pups from day 1 to day 7 expressed as mean ±SEM (n=6) (between).

Groups	Daily milk production by the means of weight gain of Pups from day 1 to day 7 (g)
Groups	D1	D2	D3	D4	D5	D6	D7
Control	0.206 ±0.08	0.0821 ±0.0168	0.105 ±0.0511	0.0392 ±0.00701	0.137 ±0.0294	0.173 ±0.0674	0.116 ±0.00835
Standard (Pueraria tuberosa-1000mg/kg)	0.0465 ±0.016	0.0608 ±0.008	0.0770 ±0.015	0.144 ±0.031	0.057 ±0.012	0.186 ±0.061	0.168 ± 0.03
TL (Dioscorea pentaphylla Linn. -500mg/kg)	0.161 ±0.06	0.161 ±0.06	0.245 ±0.063	0.183 ±0.05	0.107 ±0.04	0.185 ±0.06	0.224 ±0.13
TH (Dioscorea pentaphylla Linn-1000mg/kg)	0.0675 ±0.028	0.119 ±0.021	0.104 ±0.038	0.0802 ±0.021	0.0673 ±0.021	0.137 ±0.054	0.0830 ±0.022

Table 8B: Table showing daily milk production by the means of weight gain in Pups (g) the from day 8 to day 15 expressed as mean expressed as mean ±SEM (n=6).

Groups	Daily milk production by the means of weight gain in Pups (g) from day 8 to day 15
Groups	D8	D9	D10	D11	D12	D13	D14	D15
Control	0.0921 ±0.00645	0.185 ±0.0114	0.0839 ±0.00377	0.150 ±0.0353	0.124 ±0.0124	0.127 ±0.0198	0.127 ±0.0181	0.0810 ±0.00733
Standard (Pueraria tuberosa-1000mg/kg)	0.0953 ±0.0270	0.0922 ±0.0250	0.0741 ±0.0205	0.203 ±0.0424	0.130 ±0.0334	0.0935 ±0.0141	0.119 ±0.0247	0.136 ±0.0565
TL (Dioscorea pentaphylla Linn. -500mg/kg)	0.264 ±0.0775	0.191 ±0.0558	0.190 ±0.0504	0.205 ±0.0488	0.107 ±0.0482	0.264 ±0.0826	0.131 ±0.0363	0.393 ±0.132
TH (Dioscorea pentaphylla Linn-1000mg/kg)	0.111 ± 0.0241	0.146 ±0.0341	0.170 ±0.0487	0.231 ±0.0803	0.165 ±0.0674	0.128 ±0.0299	0.169 ±0.0584	0.209 ± 0.0610

The data related to the daily milk production by the means of weight gain in pups (g) on different days have been depicted in Table 9A & 9B. Comparison of D1 milk production by the means of weight gain in pups (g) was done by the milk production by the means of weight gain in pups (g) recorded on D2 to D15.
Between the groups, no significant difference could be observed in milk production by the means of weight gain in pups (g) as measured on D1 to D15 even though there was apparent increase in the same during the treatment course.
Within the group; Control group, Statistical analysis was made using repeated measures ANOVA, followed by Tukey’s

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Akshatha Somayaji K. et al. Galactagogue activity of Bidali (Dioscorea pentaphylla Linn.)

test, where statistical significance was found with a p<0.007. In Standard Group, Statistical analysis was made using repeated measures of ANOVA, where statistical difference was not found. In TL Group, Statistical analysis was carried out by using Friedman repeated measures ANOVA, where no statistical difference was noted. In TH Group: Statistical analysis was carried out by using Friedman repeated measures of ANOVA, where no statistical difference was noted.

b.Pup Weight Gain

Table 9A: The table describes the data related to daily pup weight gain (g) from day 1 to day 7 expressed in Mean and ±SEM (n=6) (between the group)

Groups	Daily Pup weight gain from day 1 to day 7 (g)
Groups	D1	D2	D3	D4	D5	D6	D7
Control	6.116 ±0.377	6.209 ±0.472	6.847 ±0.697	7.929 ±0.784	9.151 ±0.980	10.251 ±1.242	11.782 ±1.459
Standard (Pueraria tuberosa-1000mg/kg)	6.390 ±0.124	7.046 ±0.223	7.866 ±0.391	8.789 ±0.567	10.101 ±0.584	11.234 ±0.705	12.524 ±0.845
TL (Dioscorea pentaphylla Linn. -500mg/kg)	6.420 ±0.269	7.383 ±0.351	8.438 ±0.467	9.726 ±0.622	10.912 ±0.807	12.017 ±0.964	13.116 ±1.124
TH (Dioscorea pentaphylla Linn. -1000mg/kg)	6.067 ±0.195	6.349 ±0.212	7.402 ±0.288	8.496 ±0.459	9.526 ±0.462	10.680 ±0.507	12.064 ±0.493

Table 9B: The table describes the data related to daily pup weight gain(g) from day 8 to day 15 expressed in Mean and ±SEM (n=6) (between the group)

Groups	Daily Pup weight gain from day 8 to day 15 (g)
Groups	D8	D9	D10	D11	D12	D13	D14	D15
Control	13.385 ±1.760	14.951 ±1.957	16.618 ±2.128	17.740 ±2.306	18.830 ±2.553	19.840 ±2.511	21.645 ±3.051	23.502 ±3.457
Standard (Pueraria tuberosa-1000mg/kg)	13.962 ±0.853	15.404 ±0.922	16.926 ±1.021	18.329 ±1.015	19.601 ±1.145	20.670 ±1.138	22.911 ±1.516	24.239 ±1.511
TL (Dioscorea pentaphylla Linn. -500mg/kg)	14.074 ±1.300	15.140 ±1.424	16.926 ±1.557	18.109 ±1.683	19.058 ±1.797	20.873 ±1.894	22.276 ±2.001	23.6 ±2.162
TH (Dioscorea pentaphylla Linn. -1000mg/kg)	13.275 ±0.499	14.659 ±0.498	15.490 ±0.360	16.867 ±0.309	18.347 ±0.240	20.086 ±0.285	21.359 ±0.325	22.815 ±0.547

The above table 15A &B depicts the daily weight gain of Pups (g) from day 8 to day 15 daily weight of Pups (g), expressed in Mean and ±SEM (n=6). where TH- Trial drug (Dioscorea pentaphylla) in Higher dose (1000mg/Kg p.o.) and TL- Trial drug (Dioscorea pentaphylla Linn.) in Lower dose (500mg/Kg p.o.)
Between the Group; the statistical analysis

was carried out using repeated measures of one-way analysis of variance, where there was no statistical significance was elucidated.
Within the group; In Control Group: Statistical analysis carried out by using Freidman repeated measures one way analysis of variance on rank, followed by Tukey Test; where the results are Highly significant; p<0.001 from Day 1 to Day 15. In Standard Group: Statistical analysis carried out by using repeated measures of one-way analysis of variance, followed by holm Sidak method: where the results are highly significant; p<0.001 from Day 1 to Day 15. In TL Group: Statistical analysis carried out by using repeated measures of one-way analysis of variance, followed by Tukey Test: where the results are highly significant; p<0.001 from day 1 to day 15. In TH Group: Statistical analysis carried out by using repeated measures of one-way analysis of variance, followed by Holm Sidak method: where the results are highly significant; p<0.001.
Serum prolactin concentration

Table 10: Table showing the serum prolactin concentration of different groups.

SN	Serum prolactin concentration(ng/dl)
SN	Groups		Day1	Day14	Day 21
1.	Control	C1	<0.47	1.29	<0.47
1.	Control	C2	<0.47	<0.47	<0.47
2.	Standard (Pueraria tuberosa-1000mg/kg)	S1	<0.47	<0.47	<0.47
2.	Standard (Pueraria tuberosa-1000mg/kg)	S2	<0.47	<0.47	<0.47
3.	TL (Dioscorea pentaphylla Linn. -500mg/kg)	TL1	<0.47	<0.47	<0.47
3.	TL (Dioscorea pentaphylla Linn. -500mg/kg)	TL2	0.68	<0.47	<0.47
4.	TH (Dioscorea pentaphylla Linn-1000mg/kg)	TH1	<0.47	<0.47	<0.47
4.	TH (Dioscorea pentaphylla Linn-1000mg/kg)	TH2	4	<0.47	<0.47

1. Histology of Breast Tissue

Table 11: The table showing findings of Histology of Breast Tissue in all four groups.

Figure 1: Control group
Section studied shows increase in tubule-alveolar unit with focal distorted alveoli [Fig 1A, arrow] and devoid of secretions. At focal areas hyperplasia of alveoli with distinct luminal lined by cuboidal to columnar epithelium, few containing intra-luminal secretions [Fig.1B, Arrow] and surrounded by basement membrane are seen. The intervening fibro-collagenous stroma appears variably.
Figure 1A	Figure 1B

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Akshatha Somayaji K. et al. Galactagogue activity of Bidali (Dioscorea pentaphylla Linn.)

Figure 2: Standard Group
Section studied shows increase in tubulo-alveolar unit with hyperplasia of alveoli in the hypodermis. The alveoli have distinct lumina lined by columnar epithelium with intraluminal secretions and surrounded by basement membrane [Fig.2A, Arrow]. The epithelium appears hypertrophied at some places. The intervening fibro-collagenous stroma appears reduced [Fig.2B, Arrow]
Figure 2A	Figure 2B
Figure 3: TL Group
Section studies showed decreased Tubulo- alveolar unit with alveoli in the hypodermis. The alveoli have distinct lumina lined by single layer of cuboidal epithelium surrounded by thick basement membrane [Fig.3A, Arrow] and interstitium. The alveoli are spaced with intervening dense fibro collagenous stroma [Fig.3B, Arrow].
Figure 3A	Figure 3B
Figure 4: TH Group
The section studied showed increase in tubule-alveolar unit with hyperplasia of alveoli in the hypodermis. The alveoli have distinct lumina lined by columnar epithelium with intra-luminal secretions and surrounded by basement membrane [Fig.4A, Arrow]. The epithelium appears hypertrophied and hyperplastic at some places. The intervening fibro-collagenous stroma appears reduced [Fig.4B, Arrow].
Figure 4A	Figure 4B

Discussion

Acharya Dalhana in Nibandhasangraha has described two varieties of Vidarikanda while commenting on the same in Sushruta Samhita, Viz., One with tubers that resemble the foot of an Elephant- Hastipadavat, and the other with tubers that are elongated- Dirghakanda.^[4]

Bidali (Dioscorea pentaphylla Linn.), which is abundantly found throughout India, has the elongated tubers-Dirghakanda, is well accepted by K C Chunekar & Thakur Balwant Singh as the variety of Vidarikanda.^[4,6] It also is used as the vegetable/staple food in the Western Ghats and North-Eastern parts of India.^[9]Hence, Bidali was chosen for study its Galactagogue action by experimental evaluation.

Weight of Dams

Earlier, in the study related to assessment of milk production, the breast of the dams under the study were dissected and weighed to infer the increase in milk production, Later the procedure was modified, and the body weight of dams (as whole) was considered to infer the same.^[14]

In the present study, increase in body weight of dams were noted in all the four groups, although there was no statistical significance between the groups, Statistical significance was observed within the groups (Control- P <0.001, Standard- P <0.001, Trial Drug- Dioscorea pentaphylla Linn. in lower dose: 500mg/Kg- P <0.006 and Trial Drug-Dioscorea pentaphylla Linn. In higher dose:1000mg/Kg-<0.001).

Milk Estimation by the means of Pup Weight and Weight gain

1. Milk Estimation by means of Pup weight:

Weight increase in the litter/ pups is frequently employed as a measure of milk production since it provides an estimate of milk productivity which is corrected by the weight loss due to metabolic activities like excretion and respiration in pups.^[13,^14] Although there was increased milk estimation by the means of weight but was not up to the statistical significance in all the groups.

1. Pup weight Gain

The Pups/ Litter growth is an indicative of adequate milk production by their dams for pup nourishment.^[13,14] Here was statistical significance within the group with P <0.001 in all the four groups, but between the groups there was no statistical significance.

Prolactin

The initiation of lactation, the composition of milk's macronutrients, and milk production all depend

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on the main lactogenic hormone, Prolactin, which is secreted by the anterior pituitary. Its levels in the blood rise throughout pregnancy to a level that is 10 to 20 times higher than usual at the end of gestation.^[15]

Rat prolactin (rPRL), a single-chain polypeptide hormone produced by the rat anterior pituitary, has a mass of about 23,000 molecules. The amino acid sequence of prolactin varies significantly between species. About half of all residues in rat prolactin are different from those in human prolactin.^[16]

In an experiment, Rats were used to evaluate serum prolactin levels, and the results revealed that the serum prolactin level increased on the first postpartum day and persisted high for at least 8 days due to the suckling stimulus. Serum prolactin levels decreased to about half of what they were on the first post-partum day on the 15^th and 23^rd days after delivery. When litters were removed from mother rats on the fourth postpartum day, serum prolactin dropped quickly and returned to pregnancy levels three hours later.^[17]

In the current study, the serum prolactin levels were <0.47 ng/dl, except on 14th day in Control group (1.29ng/dl), 1st day of TH group (Dioscorea pentaphylla Linn-1000mg/Kg) (4ng/dl), and 1st day of TL group (Dioscorea pentaphylla Linn. -500mg/Kg) (0.68ng/dl).

The result- (uniform, unchanged, undetectable) might be due to lack of suckling reflex (nipple stimulation), which is also a cause for the secretion of Prolactin ^[18] or due to the insensitivity of the ELISA kit used for the estimation (below the detection level).

Histology of Breast tissue

Parenchyma and interstitial tissue were seen in the normal breast tissue samples taken from non-inbred adult rats. The terminal ducts and acini that made up the parenchyma were homologous to the terminal duct-lobule in the human breast. Fatty tissues made up most of the interstitial tissues. Ductal epithelial cells, myoepithelial cells, and basement membrane made up the three layers of normal ducts. The ductal epithelial cells were constituted of one layer of slender, club-shaped cells made up of two types of cells: dark cells and intermediate cells. A single layer of cells, mostly dark cells with sporadic myoepithelial cells, lined the acini.^[19]

In the present study, the animal receiving the Bidali (Dioscorea pentaphylla Linn.) in higher dose (1000mg/Kg) breast tissue histology revealed increased tubule-alveolar unit with hyperplasia of alveoli in the hypodermis, The alveoli have distinct lumina lined by columnar epithelium with intra-luminal secretions and surrounded by basement membrane and in the group receiving the lower dose (500mg/Kg) of trial drug Bidali (Dioscorea pentaphylla Linn.) breast tissue showed very few tubulo-alveolar unit with fewer alveoli in the hypodermis and thick basement membrane. This is suggestive of that the Bidali (Dioscorea pentaphylla Linn.) in the higher dose (1000mg/kg), promoted the growth of tubule-alveolar unit, and intra-luminal secretion.

Mode of action of Bidali on Stanya Janana

Mode of action

Shleshma-Vardhaka Dravya Prayoga is advocated in Stanya Janana, due to the Samanya Guna between Shleshma and Stanya^[20]

Table 12: The Table showing the Mode of Action of Bidali through Rasapanchaka and its Karma

Rasapanchaka		Karma
Rasa	Madhura	Dhatu Bala Pradhana Stanya-Shleshma Vardhana Brihmana[21]
Guna	Guru	Shleshma Vardhana Bala Vardhana Brihmana[22]
	Shita	Shleshma Vardhana[22]
	Snigdha	Shleshma Vardhana[22]
Virya	Shita	Shleshma Vardhana[23]
Vipaka	Madhura	Shleshma Vardhana[24]
Karma		Rasayana, Brihmana[25] Stnaya Vardhana [26]

Therefore, because of the above Guna-Karma, its advocation in Stanya Kshaya is well justified where it counters the Samprapti of Stanya Kshaya and increases the Stanya to adequate quantity.

Mode of action based on Phytochemical constituents:

Qualitative Phytochemical analysis showed the presence of Alkaloids, Flavonoids, Tannins, Saponins, Steroids, Carbohydrates, Phenols. On quantification of Alkaloids, Tannins, Flavones, and Saponins, Flavones was found in more quantity among them.^[27]
Diosgenin (steroidal sapogenin), A phytoestrogen which exerts agonist effect

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Akshatha Somayaji K. et al. Galactagogue activity of Bidali (Dioscorea pentaphylla Linn.)

and effects on the anterior pituitary to produce Prolactin secretion in Lactotrophs.^[28]
Flavonoid, is known to have effect on ERα (Alpha Estrogen Receptor), which promotes growth of Mammary gland and Production of Prolactin Hormone.^[29]
The alkaloids in the drug function as a protein synthesis reservoir and accelerate metabolism, which may also explain why both Dams and Pups have gained weight.^[30]
The presence of steroids in the drug aids in the stimulation of growth factor and its impact on bones, muscles, and adipose tissue, which may have contributed to weight gain in dams and pups as well as the development of the mammary gland.^[30]
The influence of steroids on the growth of adipose tissue aids in the production of prolactin hormone since adipose tissue also produces prolactin hormone.^[18,30]

Conclusion

Bidali (Dioscorea pentaphylla Linn.) is one among the two varieties of Vidari. The model estimation of milk yield by the means of pup weight and weight gain on Wistar albino rats was done. Outcome of the present study showed better results in the group treated with the Trial drug in dose dependent manner, Bidali (Dioscorea pentaphylla Linn.) in terms of Dam weight, Daily Milk Yield in means of pup weight and pup weight gain and Histology. The statistical comparison revealed that the difference was insufficient to reject the null hypothesis. Thus, based on this experimental study, it can be concluded that Bidali acts as both Stanya Janana and Stanya Vradhaka.

Acknowledgement:

The author is grateful to valuable guidance, support and constant encouragement provided by Dr Ravishankar B, in completion this research work.

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Figure 5: Grouping and housing of Animals