Indian Journal of Experimental Biology


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VOLUME 44

NUMBER 6

JUNE 2006

CODEN : IJEB (A6) 44(6) 425-514(2006)

ISSN : 0019-5189

 

CONTENTS

 

Review Articles

 

Cerebellar control of visceral responses – possible mechanisms involved

429

      O P Tandon, Varun Malhotra, VijayaBhaskar P & Shankar P R

 

 

 

Peroxynitrite: A potent oxidizing and nitrating agent

436

      J P Kamat

 

 

 

Papers

 

Histological evidences of reparative and regenerative effects of ß-adrenoceptor agonists, clenbuterol and isoproterenol, in denervated rat skeletal muscle

448

      Surender S Katoch, Asha Garg & Sushma Sharma

 

 

 

Oral poly-(lactide-co-glycolide) nanoparticle based antituberculosis drug delivery: Toxicological and chemotherapeutic implications

459

      Rajesh Pandey, Sadhna Sharma & G K Khuller

 

 

 

Antioxidant and anti-inflammatory activity of extract obtained from Aspergillus candidus MTCC 2202 broth filtrate

468

      Pankaj P Malpure, Abhishek S Shah & Archana R Juvekar

 

 

 

Antioxidant and antiulcer activity of aqueous extract of a polyherbal formulation

474

      Annie Shirwaikar, H N Aswatha Ram & P Mohapatra

 

 

 

Positive inotropic effect of Murraya koenigii (Linn.) Spreng extract on an isolated perfused frog heart

481

      Kunal J Shah & Archana R Juvekar

 

 

 

Alkaline lipase production by Citrobacter freundii IIT-BT L139

485

    Vijay Gunasekaran, Shireen Meher Kotay & Debabrata Das

 

 

 

Optimization of RQRT-PCR protocols to measure b-1,3-glucanase mRNA levels in infected tissues of rubber tree (Hevea brasiliensis)

492

      I Thanseem & A Thulaseedharan

 

 

 

Reduction of vitrification in in vitro raised shoots of Chlorophytum borivilianum Sant. & Fernand., a rare potent medicinal herb

499

      Urvashi Sharma & J S S Mohan

 

 

 

Influence of cytokinins, auxins and polyamines on in vitro mass multiplication of cotton (Gossypium hirsutum L. cv. SVPR2)

506

      Ganesan M & Jayabalan N

 

 

Author Index

Aswatha Ram H N

474

Das Debabrata

485

Ganesan M

506

Garg Asha

448

Gunasekaran Vijay

485

Jayabalan N

506

Juvekar Archana R

468, 481

Kamat J P

436

Katoch Surender S

448

Khuller G K

459

Kotay Shireen Meher

485

Malhotra Varun

429

Malpure Pankaj P

468

Mohan J S S

499

Mohapatra P

474

Pandey Rajesh

459

Shah Abhishek S

468

Shah Kunal J

481

Shankar P R

429

Sharma Sadhna

459

Sharma Sushma

448

Sharma Urvashi

499

Shirwaikar Annie

474

Tandon O P

429

Thanseem I

492

Thulaseedharan A

492

VijayaBhaskar P

429

 

 

Keyword Index

Acclimatization

506

Actin

492

Alcohol

474

Anti-inflammatory activity

468

Antioxidant activity

468

Antioxidant

474

Anti-ulcer

474

Aphrodisiac

499

Aspergillus candidus

468

Butoxamine

448

Chemotherapy

459

Citrobacter freundii

485

Clenbuterol

448

Crebellar control

429

Denervation

448

Enzyme production

485

Fermenter studies

485

Frog heart

481

b-1, 3-Glucanase

492

Gossypium hirsutum

506

Growth retardant

499

Hardening

506

Hevea brasiliensis

492

Hingwashtak churna

474

Ibuprofen

474

Isoproterenol hydrochloride

448

Lipase

485

Micropropagation

499

MS medium

506

Murraya koenigii

481

Nanoparticles

459

Neuronal architecture

429

Nitrating agent

436

Nuclei

448

Optimization

485

Organogenesis

506

Oxidant

436

Peroxynitrite

436

Phenolic compound

468

Phytophthora

492

Poly (lactide-co-glycolide)

459

Polyamines

506

Positive inotropic effect

481

Rat gastrocnemius

448

Reducing power

468

Relative RT-PCR

492

Rheology

485

Safed musli

499

Shoot multiplication

499

Somatomotor

429

Toxicity

459

Tuberculosis

459

Visceral response

429

 

Announcement

Call for BRSI Annual Awards Nominations-2005

 

The Biotech Research Society of India [BRSI] invites nominations for the following BRSI Annual Awards–2005—1. Young Scientist Medal: Any researcher/academician of 35 years or below as on 31st December 2005 involved in Biotech R&D for his/her outstanding contribution; 2. Woman Scientist Medal: Any woman researcher/academician below the age of 45 years as on 31st December 2005 involved in Biotech R&D for her outstanding contribution carried out in India; 3. BRSI Life Time Medal: Any researcher/academician above the age of 55 years as on 31st December 2005 involved in Biotech R&D for his/her life time contribution to the field of Biotechnology; 4. BRSI Industrial Medal: Any researcher involved in Biotech R&D for his/her outstanding contribution which has resulted in commercialisation of a product/process; 5. Fellow of BRSI: Members of BRSI having long-standing and significant contribution to the field as well as eminent scientists/technologists working in various areas of biotechnology shall be considered to be elected as Fellow of BRSI (FBRS).

Further details and application form can be obtained from Prof. Ashok Pandey, President-BRSI, Regional Research Laboratory, CSIR, Trivandrum-695 019, India; Phone: 0471-2515279/2495949; Fax 0471-2491712; E-mail: brsihq@yahoo.co.in, ashokpandey56@ yahoo.co.in

 

———————————

 

Review Article

Indian Journal of Experimental Biology

Vol. 44, June 2006, pp. 429-435

 

 

  

Cerebellar control of visceral responses–possible mechanisms involved

O P Tandon, Varun Malhotra, VijayaBhaskar P & Shankar P R

 

 

It seems reasonable to assume that cerebellar autonomic control operates according to similar principles as those utilized in the somatomotor coordination. The unique and very uniform neuronal architecture throughout the cerebellum speaks in favour of such a view.

 

 

Review Article

 

Indian Journal of Experimental Biology

Vol. 44, June 2006, pp. 436-447

 

 

 Peroxynitrite: A potent oxidizing and nitrating agent

 

J P Kamat

 

Nitric oxide (NO.) reacts with superoxide (O2-.) forming peroxynitrite (PXN) (ONOO-), a strong oxidant which reacts with several biomolecules leading to enormous implications in biological process1-3, holds enormous implications for the understanding of free radicals. The ONOO- formation in vivo has significant implications in free radical biology. It exerts a defensive role in large number of pathophysiological reactions2,3 and also acts as signaling molecule in activation of several protooncogenes4,5. It decomposes rapidly to an intermediate and reacts with several biomolecules4. Evidence for PXN formation in vivo has been obtained immunohistochemically through detection of a characteristic reaction product with protein tyrosine residues and 3-nitrotyrosine6. This “biomarker” of PXN formation has now been identified in various pathologies such as Lou Gehrig’s disease, Parkinson’s disease, cancer, atherosclerosis as well as in biological aging.
3-nitrotyrosine formation has been documented in various tissues, e.g. even in non-diseased embryonic heart during normal development. Therefore, there is a great opportunity in the postgenomic period to understand the interplay of these molecular interactions with biological events such as apoptosis, gene regulation etc. This review deals with biological significance of peroxynitrite, its precursors, reactions with large range of biomolecules, including aminoacids, proteins, lipids, nucleic acids, antioxidants as well as cytotoxic aspects.

 

 

Indian Journal of Experimental Biology

Vol. 44, June 2006, pp. 448-458

 

 

Histological evidences of reparative and regenerative effects of ß-adrenoceptor agonists, clenbuterol and isoproterenol, in denervated rat skeletal muscle

Surender S Katoch, Asha Garg & Sushma Sharma

 

Received 22 February 2005; revised 23 March 2006

The aim of this study was to determine the contribution of ß-adrenoceptor activation in the reconstruction of the structural and functional organization of denervated skeletal muscle. ß-agonists, clenbuterol (1.2 mg/kg body weight) and isoproterenol (2 mg/kg body weight), administration (daily oral administration; maximum 7 days) to normal innervated rats as well as denervated animals caused muscle hypertrophy. An increase in mean fiber diameter confirmed this stimulated growth both in normal innervated and denervated rat gastrocnemius muscle. Examination of muscle nuclei from treated but normal innervated rat gastrocnemius exhibited features like large size, active nucleoplasm and an increase in their number per fiber cross section and per mm mean fiber length indicating towards an elevated biosynthetic activity in tissue in the presence of ß adrenoceptor agonists. Administration of drugs to normal innervated animals resulted in an emergence of central muscle nuclei. The hyperactive and enlarged muscle nuclei ultimately organized themselves into unusually elongated nuclear streaks. ß agonist treatment to denervated rats resulted in amelioration of atrophic state of tissue characterized by hypertrophy of muscle fibers thus lending to a restoration of structural organization of tissue. Bizarre shapes of nuclei in denervated muscle tend to recover to that characteristic to normal innervated muscle in presence of clenbuterol and isoproterenol hydrochloride. All observations were confirmed by administering butoxamine, a ß-adrenoceptor antagonist along with ß-agonists. The results suggests that both clenbuterol and isoproterenol hydrochloride are capable of mimicking normal innervation functions in skeletal muscle and thus play important role in the structural and functional reorganization of tissue. Amelioration of denervation atrophy in rat gastrocnemius in the presence of ß-agonists supports this.

 

  

Indian Journal of Experimental Biology

Vol. 44, June 2006, pp. 459-467

 

 

Oral poly(lactide-co-glycolide) nanoparticle based antituberculosis drug delivery: Toxicological and chemotherapeutic implications

Rajesh Pandey, Sadhna Sharma & G K Khuller

 

Received 21 October 2005; revised 17 March 2006

The present study reports on the detailed toxicological and chemotherapeutic evaluation of antituberculosis drug loaded nanoparticles in mice. A single oral dose administration of poly(lactide-co-glycolide) (PLG, a synthetic polymer) nanoparticles containing rifampicin+isoniazid+pyrazinamide+ethambutol could maintain drug levels in various tissues for 9-10 days and did not elicit any adverse response even when administered at several fold higher than the recommended therapeutic dose. However, dosing with conventional free drugs at the equivalent higher doses was lethal. Despite multiple oral dosing with the formulation at every 10th day, no toxicity was observed on the completion of subacute (28 days) or chronic (90 days) toxicity studies based on survival, gross pathology, histopathology, blood biochemistry and hematology. In mice harboring a high mycobacterial load (mimicking human tuberculosis), two independent chemotherapeutic regimens, i.e. 5 doses of PLG nanoparticles encapsulating (rifampicin+isoniazid+pyrazinamide+ethambutol) administered 10 days apart, or 2 doses of the 4-drug formulation followed by 3 doses of 2-drug formulation (rifampicin+isoniazid) resulted in undetectable bacilli. Further, the efficacy was comparable to 46 daily doses of oral free drugs. Therefore, the experimental evidence suggests that PLG nanoparticle-based antituberculosis drug delivery system is safe and well suited for prolonged and intermittent oral chemotherapy.

 

  

Indian Journal of Experimental Biology

Vol. 44, June 2006, pp. 468-473

  

Antioxidant and anti-inflammatory activity of extract obtained from
Aspergillus candidus MTCC 2202 broth filtrate

Pankaj P Malpure, Abhishek S Shah & Archana R Juvekar

 

Received 20 September 2005; revised 13 March 2006

Antioxidant potential of Aspergillus candidus MTCC 2202 broth filtrate extract was studied using different antioxidant models, whereas anti-inflammatory potential was studied using carrageenan-induced rat paw oedema model. The ethyl acetate extract at 1000 µg/ml showed maximum scavenging activity of the stable radical 1,1-diphenyl,2-picryl hydrazyl upto 96.65% (IC50=430.36 µg/ml) and scavenging of the radical cation, 2,2-azinobis-(3-ethylbenzothiazoline-6-sulphonate) upto 92.25% (IC50=606.29 µg/ml) at the same concentration. The extract had good reducing power, however showed moderate inhibition for conjugated dienes and thiobarbituric acid reactive acid substances (59.56 and 51.45%). The total phenolic content of various extracts of A. candidus broth filtrate was measured and a correlation between radical scavenging activities of extracts with total phenolic content was observed. The ethyl acetate extract (125 mg/kg ip) showed significant anti-inflammatory activity in carrageenan-induced rat paw oedema model. The exhibited antioxidant activity of ethyl acetate extract of A. candidus broth filtrate was comparable with BHA and ascorbic acid, while anti-inflammatory activity was comparable with standard diclofenac sodium.

 

Indian Journal of Experimental Biology

Vol. 44, June 2006, pp. 474-480

  

Antioxidant and antiulcer activity of aqueous extract of a polyherbal formulation

Annie Shirwaikar, H N Aswatha Ram, P Mohapatra

 

Received 15 July 2005; revised 14 March 2006

The aqueous extract of Hingwashtak churna was evaluated for gastroprotection in rats using the ibuprofen and ethanol induced ulcer models. Efficacy was assessed by determination of mean ulcer size, ulcer number and ulcer index. Oral administration of the aqueous extract (750 mg/kg) significantly protected against gastric lesions by 84.96% and 91.12% as compared to ranititidine (95.54 and 95.2%) in the ibuprofen and alcohol induced ulcer models respectively. The findings suggest that the significant gastroprotective activity could be mediated by its antioxidant activity which was evaluated by using different antioxidant models of screening.

 

Indian Journal of Experimental Biology

Vol. 44, June 2006, pp. 481-484

  

Positive inotropic effect of Murraya koenigii (Linn.) Spreng extract on an isolated perfused frog heart

Kunal J Shah & Archana R Juvekar

 

Received 23 September 2005; revised 17 March 2006

Ethanolic extract of fresh leaves of M. koenigii (MKEE) showed a dose dependent positive inotropic effect on isolated frog heart. The responses to MKEE (62.5–1000 mg) were not affected in either way by theophylline, imidazole, propranolol and sildenafil. The change in potassium and sodium concentration did not alter MKEE-induced positive inotropic effect. Lignocaine did not alter the responses to MKEE significantly. Responses to MKEE were significantly inhibited when calcium concentration was reduced to half (from 1.58 to 0.79 mM) and were significantly potentiated when calcium concentration was doubled (from 1.58 to 3.16 mM). Verapamil was found to inhibit the responses significantly. The results suggest that M. koenigii induced positive inotropic effect possibly by increasing availability of calcium from extra cellular sites.

 

Indian Journal of Experimental Biology

Vol. 44, June 2006, pp. 485-491

Alkaline lipase production by Citrobacter freundii IIT-BT L139

Vijay Gunasekaran, Shireen Meher Kotay & Debabrata Das

 

Received 26 December 2005; revised 22 March 2006

Around 150 lipase producing bacterial isolates were screened from the local soils enriched with oil. Citrobacter freundii IIT-BT L139, an isolated microbial strain, produced lipase that had high activity (8.8 U/ml) at pH 9.0 and 40oC. The 16S rDNA phylogenetic studies showed that Citrobacter freundii belongs to the family Enterobacteriaceae and later confirmed by the microbial identification. Suitable C and N sources for lipase production were deduced to be starch and peptone-urea, respectively. In a controlled fermenter (1 L), the lipase activity was found to increase by 36% (12 Uml-1). The variation of lipase activity, pH and dissolved oxygen (DO) during growth of the organism in the controlled batch fermenter were monitored. The rheological characteristics of the fermentation broth indicated that it behaved like a Newtonian fluid throughout the fermentation. The fermentation time was comparatively short (60 h). The lipase was also found to be substantially resistant to common detergents. This lipase was, thus, characterized as alkaline, thermostable and solvent stable, which was essentially desirable in pharmaceutical, detergent and other industrial applications or production.

 

Indian Journal of Experimental Biology

Vol. 44, June 2006, pp. 492-498

 

 

Optimization of RQRT-PCR protocols to measure b-1,3-glucanase mRNA
levels in infected tissues of rubber tree (Hevea brasiliensis)

 

I Thanseem & A Thulaseedharan

 

Received 16 December 2004; revised 21 March 2006

RQRT-PCR technique was evaluated for its validity as an alternative to Northern blotting for quantification of plant gene expression in diseased tissues of Hevea. Reliable RT-PCR results could be obtained by co-amplification of housekeeping actin gene as the internal control along with the gene of interest. The product of interest was quantified relative to that of the internal control by measuring net intensity of bands. Expression levels of defense-related b-1,3-glucanase gene was studied in the pathogen infected tissues of rubber. The b-1,3-glucanase gene was found to be induced in infected leaf tissues and reached a peak at 48 h after inoculation. The b-1,3-glucanase gene expression during pathogen infection was determined through Northern blot hybridization also, using 18S RNA as the internal control. RQRT-PCR and Northern hybridization showed almost similar results, thereby validating the use of this technique to study the gene expression in rubber.

 

 

Indian Journal of Experimental Biology

Vol. 44, June 2006, pp. 499-505

 

 

Reduction of vitrification in in vitro raised shoots of Chlorophytum borivilianum Sant. & Fernand., a rare potent medicinal herb

 

Urvashi Sharma & J S S Mohan

 

Received 2 November 2005; revised 17 March 2006

Reduction of vitrification in in vitro raised shoots derived from shoot bases and immature floral buds along with inflorescence axis used as explants of C. borivilianum, a rare medicinal herb is described. Shoot multiplication was obtained on MS medium with 2 mg l-1 benzylaminopurine (BAP) + 0.1 mg l-1 indole-3-butyric acid (IBA) and MS medium with 2 mg l-1 kinetin (Kin) + 0.1 mg l-1 2,4-dichlorophenoxy acetic acid (2,4-D) from shoot bases and inflorescence axis respectively. Best multiplication rates were obtained from both the explants on MS medium with 2 mg l-1 BAP. Vitrification of shoots in cultures appeared during the multiplication stage. Culture bottles with aerated caps reduced the vitrification to 80%. Reduction of BAP concentration from 2 mg l-1 to zero during subsequent subcultures also minimized vitrification. Use of 0.5-2 mg l-1 Kin produced healthy shoots when compared to BAP. In vitro raised shoots rooted on Knop salts containing iron and vitamins of MS medium, 2 mg l-1 IBA and 0.1% activated charcoal. About 80% plantlets survived upon soil transfer. Scanning electron microscopic and image analyzer studies reveal the morphological structural differences between the leaves of normal and vitrified plantlets.

 

 

 

Indian Journal of Experimental Biology

Vol. 44, June 2006, pp. 506-513

 

 

Influence of cytokinins, auxins and polyamines on in vitro mass multiplication of cotton (Gossypium hirsutum L. cv. SVPR2)

Ganesan M & Jayabalan N

 

Received 23 September 2005; revised 23 March 2006

In the present investigation, the influence of different forms of cytokinins, auxins and polyamines were tested for mass multiplication and regeneration of cotton. Initially, for the identification of effective concentration for multiple shoot induction, various concentrations of BAP, Kin and 2iP along with IAA and NAA were tested. Among tested concentrations, media fortified with MS salts; B5 vitamins; 30 g/l, glucose; 2.0 mg/l, 2iP; 2.0 mg/l, IAA and 0.7 % agar showed best response for multiplication of shoot tip explants (20 shoots per shoot tip explants). In nodal explants, maximum of 18.6 shoots were obtained in the media fortified with MS salts, B5 vitamins, 30 g/l, glucose, 2.0 mg/l, 2iP, 1.0 mg/l, NAA and
0.7 % agar. Effect of different concentrations of polyamines like spermidine and putrescine were also tested along with the above said multiplication media. Among the various treatments, 20 mg/l of putrescine showed best response and the multiple of shoots were increased to 26.5 shoots per shoot tip explants and 24.5 shoots per nodal explants. Elongation of shoots was achieved on multiple shoot induction medium. Significant number of roots were initiated in the medium supplemented with MS salts, vitamin B5 and 1BA (2.0 mg/1). The frequency of root induction was increased by addition of, PVP (10 mg/l) along with root induction medium and after 2 weeks, the roots reached the maximum length of 22 cm. Further, these plantlets were hardened by using sand, soil and vermiculate in 1:1:1 ratio. The hardened plants were transferred to the environmental growth chamber for proper acclimatization. The hardened plants were then transferred to field for boll yielding and they exhibited 100% survival.