Published in the Oct.1998 issue (Vol.90 # 10, p.614-618) of the Journal
of the National Medical Association, Washington,D.C. USA

Anti-diabetic Effects
of Dietary Supplement
"Pancreas Tonic"

by

Ramachandra M. Rao, Ph.D.,
Department of Internal Medicine
Charles R.Drew University of Medicine and Science
Los Angeles, California 90059
and
Fathi a. Salem, MD
Department of Pathology
Charles R.Drew University of Medicine and Science
Los Angeles, California 90059
and
Irene Gleason-Jordon, MD
Department of Pathology
Charles R. Drew University of Medicine and Science
Los Angeles, California 90059

Abstract 

Pancreas Tonic contains plant products shown to possess hypoglycemic activity. In thisstudy, we investigated the effect of Pancreas Tonic on serum glucose, glycosylatedhemoglobin and pancreatic islet cell regeneration. Our results showed that body weights ofthree groups were not significantly different from each other before study period, after12 week study, weights increased with non-significant difference among the groups.

Serum glucose concentration in the control group was 133 mg ± 12..3, the diabeticgroup was 182 mg ± 16.4; the diet-group was 95 mg ±8.9. The diabetic group hadsignificantly higher serum glucose compared to controls (P<0.01) and the diet-treatedgroup had significantly lower serum glucose concentration (P<0.0001) compared to thediabetic group. Percentage of glycosylated hemoglobin (GHB) in the control group was 8.1± 0.27; the diabetic group was 9.1 ± 0.23, the diet group was 7.0 ± 0.29. The diabeticgroup's GHB was significantly higher compared to control (P<0.01) and the diet treatedgroup had significantly lower GHB compared to the diabetic group (P<0.0001) and thecontrol group.

Histological analysis of the pancreas showed a generalized reduction in size and numberof islets in the diabetic group and regeneration of islet cells in the diet-treated groupcompared to the diabetic group. The number of cells per islet were: controls: 111 ± 8.9;diabetic: 40 ± 14; and diet group: 79 ±8.8. The diabetic group had a significantreduction in the number of cells compared to the controls (P<0.02). The diet-treatedgroup contained a significantly increased number of cells (P<0.02) compared to thediabetic group. Our data suggests that Pancreas Tonic induced an anti-diabetic effectthrough pancreatic islet cell regeneration in experimental rats.

Pancreas Tonic (PT) a dietary supplement, is composed of several plant products andextracts of the Indian sub-continent. The majority of these ingredients either alone or incombination have been shown to have anti-diabetic effects in diabetic animal models. Forcenturies, the Indian people have been using these plant products in their diets as cookedor steamed vegetables, apparently deriving the benefits of regulating their blood sugarconcentration, as the predominant ingredient in their diet is carbohydrate.The extracts of seeds and leaves of the following plants are in the PT: Cinnamomum tamala,Pterocarpus marsupeum, Momardica charantia, Azardichta indica, Tinospora cardifolia, Aeglemarelose, Gymnema sylvestre, Syzigium cumini, Trigonella foenum graecum and Ficusracemosa. The flavonoidal component epicatechin from Pterocarpus marsupeum has been shownto have anti-diabetic action (¹). Several types of extracts of Trigonellafoenum graceum when administered to orally to rabbits have been shown to have hypoglycemicactivity (²). Momordica chirantia which is commonly known as "karela"in northern India is being consumed in the daily diet (in fruits). The hypoglycemicactivity of its seeds was demonstrated in rats (³) and its active principle wasshown to possess anti-diabetic activity (⁴). Trigonella foenum graecum anotheringredient in PT, known as fenugreek, shown anti-diabetic activity in humans with diabetes(⁵). Another independent study with Trigonella foenum graecum showed a doserelated hypoglycemic effect in alloxan induced diabetic rats (⁶). Gymnemasylvestre another component of PT, when administered orally to diabetic rats, has loweredblood sugar level (⁷) but later investigation in Japan (⁸) usingGymnema sylvestre has been unable to show any improvement in insulin resistance instreptozotocin induced diabetic rats. The information available so far pertaining to theseplant products is either scanty and/or conflicting. In the present study, we investigatedthe effect of oral administration of PT which contains known or reported hypoglycemicplant products on the biochemical and histological parameters in alloxan-induceddiabetes. 

RESEARCH DESIGN AND METHODS

Materials: PT was prepared from the following plant products: Pterocarpus marsupeum(heartwood); Momardica charantia (seeds); Gymnema sylvestre (leaves); Cinnamomum tamala(leaves); ; Aegle marelose (leaves); Azardichta indica (leaves); Tinospora cardifolia(stem); Trigonella foenum graceum (seeds) ; Ficus racemosa (leaves) and Syzigium cumini(fruit).

Animals: Animals in this experiment were comprised of three groups (group n=10) ofSprague-Dawley (SD) male rats (6 weeks of age). Animals were randomly assigned to one ofthe following groups: (a) control group - no treatment placed on normal rat chow (b)diabetic group - intraperitoneal injection of alloxan (80 mg/Kg body weight) after 4-6hours food withdrawal and placed on normal rat chow (c) PT group - intraperitonealinjection of alloxan (80 mg/Kg body weight) given after 4-6 hours food withdrawal andplaced on normal diet for five days (to have alloxan induced damage on pancreas) andshifted to PT-supplemented (2 % W/W) rat food. PT-supplemented food has been prepared byHarlan Teklad, Madison, WI (diet formulation # TD 96313). All of the animals were placedon these diets for 12 weeks. Body weights and feed consumption were recorded weekly. Allanimals were observed daily for general health and normal movements in the cages. Nosignificant changes were observed in the overall health of these animals during the periodof study.

Method of collection and processing of blood samples: The access to food waswithdrawn for approximately four hours and animals were anesthetized by ketamine-xylazine(80 mg-8 mg/Kg intraperitoneal injection) and blood samples were collected by 10 mlsyringe with an 18 gauge needle. Approximately 8-10 ml blood was placed in a tube forserum and 3-4 ml was placed in a lavender top tube for glycosylated hemoglobin analysis.

Serum biochemistry: All samples were coded with numbers and the technician whoperformed the analysis was blinded to the group information. Serum samples were analyzedfor chem.-20 panel (serum glucose) by standard autoanalyzer made by Beckman Synchron CX-7.Glycosylated hemoglobin concentration were determined by routine gel electrophoreticmethod by DiaSTAT system made by BioRAD (San Francisco, CA).

Preparation of histological slides: The pancreas was excised from the euthanizedrats and fixed in 10% buffered formalin. Paraffin blocks were prepared and 5 m sectionswere cut with a microtome (American Optical Corporation, "820" Spencer)and routine microscopic slides were prepared. Hematoxylin - Eosin staining was performedand all slides were histologically examined for number of islets and total number of cellsper pancreatic islet.

Statistical Analysis: All observations were first recorded in a note book andentered into Macintosh LCII computer and verified by another person for accuracy of dataentry. The statistical analysis was performed using StatView 4.5 (Abacus Concepts, CA)software program. Each value contained at least nine or ten observations and expressed asmean ± SEM. The significance of differences among groups was determined by ANOVA andFisher's PLSD test with P values.

RESULTS

Body weights: The beginning body weights of these three groups, presented in Table1, were not significantly different. At the conclusion of 12 weeks of the study period,the final body weights of three groups uniformly increased, but without any significantdifferences among the groups (Table 1).

Serum glucose and Glycosylated Hemoglobin: The serum glucose concentration and GHBvalues are presented in Table 2. The diabetic group had significantly higher serum glucosecompared to controls (P<0.01) and the diet-treated group had significantly lower serumglucose concentration (P<0.0001) compared to the diabetic group. The diabetic group GHBwas significantly higher compared to control (P<0.01) and diet-treated group hassignificantly lower GHB compared to the diabetic group (P<0.0001) and the controlgroup.

Number of cells per Islet: The histological analysis of the pancreases showed ageneralized reduction in size and number of islets in the diabetic group and regenerationof islets in the diet - treated group compared to the diabetic group (Fig.1A, 1B, 1C andFig.2A, 2B, 2C). The total number of cells per islet were counted and presented in Table3. The diabetic group had significant reduction in the number of cells compared to thecontrol group (P<0.02), the diet-treated group contained significantly higher cells(P<0.02) compared to the diabetic group.

DISCUSSION

Our data demonstrate that the overall body weights of the three groups of animals aresimilar (Table 1). This observation could mean that overall feed consumption and bodymetabolism were not been altered by diet-treatment. In addition, the daily feedconsumption, general locomotion and absence of any adverse symptoms among the diet-groupclearly suggest that PT-treatment did not produce adverse consequences in overall weightand health of the animals. This observation of no difference in body weights and normalfeed consumption by the diet-group is suggesting that mixing of PT in the rat feed did notworsen the taste of manufactured rat feed.

The serum glucose data suggest that intraperitoneal alloxan injections did elevateglucose concentration (Table 2) and the GHB support the fact the glucose elevations inserum were chronic in the diabetic group of rats. PT-treatment significantly decreased(P<0.0001) the serum glucose concentration along with a significant reduction (P<0.0001) in GHB in the diet-treated group. This is a significant new finding of ourstudy demonstrating not only blood glucose lowering effect of PT but also a reduction inGHB. Independent studies by other investigators (⁹,¹⁰) previouslyshowed an anti-diabetic effect of one or more plant products which are components of thePT, but in the present study we are presenting new evidence for a chronic reduction inblood glucose by PT-treatment.

The histological evidence provided in the present study (Fig. 1-3 and Table 3) clearlydemonstrate that alloxan injections destroyed the pancreatic b-cells in the diabetic groupof rats. We observed a very significant reduction in total number of cells per pancreaticislet in the diabetic group with a generalized shrinkage in size of islets. Thisobservation supports the fact that an increase in serum glucose of diabetic rats was dueto the damage done to pancreatic islets. The PT-treatment group had significantly highernumber of cells per pancreatic islet which suggests that PT-treatment regenerated thepancreatic islet cells. The histological observation of regeneration of islet cells in thePT-treated group correlates with a significant reduction of serum glucose and GHB at thesystemic level.

In conclusion, our present study suggests that the PT-treatment induced a chronicreduction in serum glucose due to the regeneration of pancreatic islet cells. Theunderlying cellular and molecular mechanisms for these observed beneficial effects of PTare to be investigated.

ACKNOWLEDGMENTS

Authors are thankful for Dr. Paul Meehan, Ph.D., Associate Professor of Physiology atUniversity of Southern California, School of Medicine for reviewing this manuscript andoffering his comments. We are also thank vivarium staff who cared for our experimentalanimals with special consideration. We thank US Botanicals, Bell Gardens, California for aresearch grant in partial support of this research project.

Table 1

Effect of Pancreas Tonic-treatment on the Body weights of Experimental Rats before andafter 12 week study period

No significance of differences among groups

Table2

Effect of Pancreas Tonic-treatment on the Serum glucose and glycosylated hemoglobin inexperimental rats

* significantly higher compared to control group (P<0.01)

** significantly lower compared to diabetic group (P<0.0001)

Table 3

Effect of Pancreas Tonic-treatment on the number of cells per Pancreatic islet inexperimental rats

* significantly lower compared to control group (P<0.02)

** significantly higher compared to diabetic group (P<0.02)

LEGENDS

Fig. 1 - Hematoxylin - eosin section showing morphology and number of cells in apancreatic islet of (A) control rat (B) diabetic rat (C) PT-diet treated rat. A, B, C x400

Fig. 2 - Hematoxylin - eosin section showing morphology and number of cells in apancreatic islet of (A) control rat (B) diabetic rat (C) PT-diet treated rat. A, B, C x1000