sporlac monograph
DESCRIPTION
Sanzyme is the 1st ever Indo-Japanes venture in India [earlier known as Uni-Sankyo], has pioneered the fermentation techniques for manufacture of LAB, ins spore form.This booklet is a tribute to all the pioneers especially Prof.Nakayama, which has changed the course of using probiotics as bacteriotherapy not only for acute, but chronic disorders.TRANSCRIPT
C e l e b r a t i n g 4 0 Y e a r s
SPORLAC
I n d i a ' s F i r s t P r o b i o t i c
1
Prof. Mashita and Prof. Nakayama assumed that the probiotic
organism they just had isolated from green malt might
immensely benefit the injured soldiers who fought the war.
That was the time the Probiotic research began, catalyzed by the
indications that probiotics may be able to prevent or improve
diarrhoea associated with antibiotic use.
How accurate they were!
The organism has been healing millions of lives since then, as Sankyo
Company developed and commercialized the organism as a
formulation for human use.
The organism isolated was a spore-bearing lactic acid-forming
bacteria. Due to their formation of spores, these bacilli have high heat
and acid resistance.
It was in the year 1973 that Sankyo, Japan transferred this strain, and
the fermentation technology to their Indian counterpart Sanzyme
limited ( the then Uni-Sankyo ltd.) the first ever Indo-Japanese joint
venture.
1949, Just after World War II... JAPAN
Prescription Pattern of SPORLAC
other than Diarrhoea*
*(The survey still being carried out across various zones of India)
Immunity Aphthous ulcers Vaginitis
Jaundice Cholesterol
AphthousUlcers - 49%
Vaginitis 39%
Jaundice32%
Cholesterol10%Immunity
52%
3
INTRODUCTION
SPORLAC, the first probiotic in India, has been successfully
used to manage GUT problems, and as it evolved SPORLAC
has been advocated for unconventional indications such as
jaundice, immunity induction, bacterial vaginitis, etc.
A survey has been conducted to understand prescription pattern and
inclination of doctors for all the applications of SPORLAC, involving
doctors across various parts of India. The survey revealed that around
89% of the doctors prescribe SPORLAC in unconventional
applications other than diarrhoea and gut problems. Amongst these
doctors 52% prescribe for immunity, 49% for Aphthous ulcers, 39%
prescribe in cases of bacterial vaginitis, 32% prescribe in cases of
Jaundice and about 10% of doctors in cases of hypercholesterolemia.
The observation from survey that 80.5% of these doctors prescribe in
more than one unconventional application makes SPORLAC a
popular probiotic in indications other than gut problems.
Internationally about half a dozen brands of this probiotic have been
marketed by companies of distinguished repute. In India, SPORLAC
is the most prescribed probiotic and has its reach to lakhs of retailers
and doctors across the country making it one of the most popular
brands.
INDEX
5
1. Birth of SPORLAC 6
2. Lactobacillus sporogenes Past,
Present & Future 8
3. Description 10
4. Characteristics of an Ideal Probiotic 14
5. Benefits of SPORLAC as a Probiotic 15
6. SPORLAC Characterization 18
7. Fermentation Characteristics 20
8. Morphological & Biochemical
Characteristics 21
9. SPORLAC Acid & Bile resistance 22
10. SPORLAC Applications 28
a. Gastroenteritis 30
b. Aphthous Ulcers 32
c. Recurrent Bacterial Vaginosis 34
d. Hypercholesterolemia 36
e. Immunity 38
f. Urticaria 40
g. Azotemia 42
h. Necrotizing Enterocolitis 44
11. SPORLAC Mechanisms of Action 46
12. SPORLAC* metabolites and
EPIGENETIC mechanism 50
13. SPORLAC Clinical Experiences 52
a. Use of Sporlac in the Treatment of
Non-specific Vaginitis 55
b. Hypercholesterolemia 58
c. Preliminary observations on effect of
SPORLAC on serum lipid levels in
hypercholesterolemic patients 59
d. SPORLAC (L.sporogenes) in
Diarrhoea 63
14. SPORLAC in Other Clinical
Conditions 64
15. Summary of the Nutritional &
Therapeutic Benefits of SPORLAC 67
16. Emerging areas for SPORLAC
Research 68
17. Bacillus coagulans : a viable adjunct
therapy for relieving symptoms of
rheumatoid arthritis according to a
randomized, controlled trial 69
18. Clinical Studies with LACBON* 70
19. PROBITECH The Science & Technology
of Manufacturing Probiotics 73
20. Citations 76
21. International Brands 80
Birth of SPORLAC
7
1949
The health of the Japanese people, at the end of world war II was at its
lowest point. Dr.Nakayama, a Japanese physician, first isolates
Lactobacillus sporogenes (Bacillus coagulans) on green malt. He believes
that these bacteria could be the answer to the problem of dysbiosis
running rampant in the Japanese people.
1964
Japanese tested Lactobacillus sporogenes (Bacillus coagulans) on adults, as
well as infants, suffering from diarrhoea and constipation. These
clinical trials used antibiotics in conjunction with Lactobacillus
sporogenes (Bacillus coagulans).
1972
The Japanese Ministry of Health and Welfare approves and Japanese
doctors are able to prescribe Lactobacillus sporogenes (B.coagulans) for
diarrhoea and constipation.
1973
Erstwhile Sankyo ( now Daiichi Sankyo Co. ltd.) offered formulation
and fermentation technology to Sanzyme ( the then Uni-Sankyo ltd.),
that led to the first probiotic of India, by brand name SPORLAC.
Lactobacillus sporogenesPast, Present & Future
9
One country and 3 names… Hindustan, Bharat and India.
Likewise Lactobacillus sporogenes, Lactic Acid Bacillus
[LAB] and Bacillus coagulans are three references used to
identify the same organism.
The species L.sporogenes was originally isolated and described in 1933
by Horowitz-Wlassowa and Nowotel now and subsequently reclassified
as Bacillus sporogenes. It has been evidenced that B.sporogenes shares
the same characters of B.coagulans, and therefore it has been moved
into Bacillus coagulans group. Accordingly to the 8th edition of
Bergey’s Manual of Determinative Bacteriology, spore-bearing rods
producing lactic acid, facultative or aerobic and catalase positive are to
be classified within the genus Bacillus.
While in the global market it is referred to as a food or nutraceutical ,the
current status in India as per the regulatory approvals for SPORLAC
treat Lactic Acid Bacillus not as nutraceutical, but as a medicine.
For all practical purposes we shall refer Bacillus
coagulans or Lactobacillus sporogenes or Lactic Acid
Bacillus as SPORLAC in this compendium.
Description
11
SPORLAC is a spore-forming Lactobacillus. The spore is
naturally encapsulated in a protective coat consisting of calcium,
protein and carbohydrate complex. It is this encapsulation that
makes SPORLAC the ideal supplemental Probiotic, protecting the
spores from gastric acid, bile, heat, oxygen and antibiotics. SPORLAC
can be stored without refrigeration, and remains viable for extended
periods of time. After ingestion, the spore coat dissolves harmlessly
leaving the spores to multiply.
SPORLAC is composed of L-lactic acid producing spores of
lactobacilli. There are three types of lactic acid that occur in the form of
three isomers (substances with identical molecular structures that have
different shapes): L(+) lactic acid, D(-) lactic acid and DL-lactic acid.
These bacteria have antagonistic effects on any bacteria that abnormally
increase, resulting in restoration of a normal balance of the bacterial
flora in the intestine. In addition, once the intestinal bacterial flora is
restored to its normal state, the spore-bearing lactic acid forming bacilli
are quickly removed from the body without them being able to form
colonies. There have been a number of reports of their favorable effects
on patients with acute and chronic inflammation of the intestine and
also on those with constipation.
The intestinal balance of bacterial composition affects the metabolism
in the intestine, thus affecting nutrition, the efficacy of drugs,
physiological functions, aging, the occurrence of cancer and
immunological functions.
13
As dietary habits have changed dramatically since World War II and
society has become more stressful due to changes in the social
environments, probiotics were good health supplements, due to their
ability to promote the maintenance of an appropriate intestinal
environment. An increasing number of reports on the health
promoting effects of SPORLAC in yogurt and health drinks.
To enhance health, it is essential for the bacilli to reach the intestinal
tract alive and intact. Spores produced by SPORLAC presented as
tablets / powder form permit the transfer of the bacilli to the large
intestine unaffected by gastric and bile acids (unlike vegetative forms,
which are vulnerable during transit), making it the choice probiotic for
a variety of ailments either singly or in combination with other
probiotics.
Characteristics of an Ideal Probiotic
Survival
Intestinal Residence
Surface Proteins
Adherence factors
Bio-film formation
Safety
Gene transfer potential
Antibiotic resistance
Putative virulence factor homologs
Functionality
Stress tolerance
Acid tolerance
Bile tolerance
Immuno-modulation
Bacetriocin production
Carbohydrate utilization &
metabolism
15
Clinical studies have revealed that SPORLAC can be
successfully implanted in the intestine. As explained in an
earlier section, SPORLAC satisfies the essential
requirements of an efficient probiotic. Preparations of SPORLAC in
pharmaceutical dosage form such as tablets, capsules, dried granules
or powder have the following characteristics :
1. Contain a large number of viable lactobacilli that retain viability
during preparation in pharmaceutical dosage forms and during
storage before consumption. The spores are thermostable as
against viable L.acidophilus cells which may not withstand
lyophilization.
2. Survive in gastric secretions and bile of the upper digestive tract
and reach the intestine safely.
3. Settle in the digestive tract and produce enough lactic acid and
other antagonistic substances to inhibit the growth of
pathogenic bacteria, thus exhibiting antibacterial activity.
Being sporulated, they germinate under favourable conditions and
produce sufficient viable cells which proliferate and perform vital
functions as described earlier. In addition SPORLAC are semi-
resistant and are slowly excreted out of the body (7 days after
discontinuation of administration).
Benefits of SPORLAC as a Probiotic
Diagrammatic Representation of the
cytological changes that take Place
during the formation
of the bacterial endospore
Vegetative cell
Chromatinfilament
upto 1.5hours after
start of sporulation
Sporeseplum
1.5 to 2.5hours after
start of sporulation
SporeProtoplast
2.5 to 4.5hours after
start of sporulation
Cortexformation
4.5 to 6.0hours after
start of sporulation
Coatformation
6.0 to 7.0hours after
start of sporulation
Maturation
17
SPORLAC is effective in the form of dietary supplements as well as
when added to food products. Natto is a traditional fermented product
from soya bean, consumed widely in japan as a rich source of protein.
Its flavor is improved by the incorporation of lactic acid bacteria such
as SPORLAC, L.acidophilus or Pediococcus acidilactiti to the starter
culture (Bacillus natto), to yield a product called Yogurunatto having
superior flavor and storage characteristics as well as improved
nutritional and therapeutic properties. A nutritive medium
(homogenized mushroom, Lentinus edodus) is mixed with soya beans
and fermented to yield this food product. The requirement of
availability of a number of viable lactic acid bacteria is fulfilled by
using SPORLAC.
Multiplication of SPORLAC begins approximately four
hours after ingestion and colonies double every thirty
minutes, producing beneficial L(+)lactic acid in the small
and large intestine through the fermentation of glucose,
fructose, sucrose, trehalose and other sugars. SPORLAC is
considered a transient inhabitant.
SPORLAC
Characterization
19
Consequent to the joint FAO/WHO Expert Consultation on
evaluation of Health and Nutritional Properties of probiotics
in 2001, certain guidelines were framed to set out a systematic
approach for the evaluation of probiotics, which include certain in
vitro tests to determine physiologic and functional health
characteristic of the strain and in vivo trials to substantiate efficacy in
humans.
Invitro tests
Acid Bile Stability
To exert their beneficial effects probiotics must resist to the acidity of
the stomach, lysozyme and bile acid. SPORLAC survives at gastric
pH and reach the intestine, where sporulation could occur.
Adhesivity
Adhesion properties are considered an important issue, and
particularly, ability to adhere to intestinal mucosa is one of the
essential selection criteria for probiotics, since adhesion to intestinal
mucosa represents the first step in colonization process. Stable
adhesion to colonic mucosa seems associated to shortening of
diarrhoea, immunogenic effects, competitive exclusion and other
effects.
Bacteriocin Production
A plasmid-encoded bacteriocin-like inhibitory substance, named
Coagulin, is produced by SPORLAC. Because of its spectrum of
activity encompassing other SPORLAC strains, enterococci, Listeria
spp, it has been proposed as an alternative to Nisin.
SPORLAC
Fermentation Characteristics
Substrate
Glucose
Xylose
Fructose
Galactose
Acid Production
+
+
+
+
Lactose
Saccharose
Maltose
Starch
Inulin
Glycerol
Mannitol
+
+
+
+
+
+
+
1
S. No
2
3
4
5
6
7
8
9
10
11
Antibiotic Resistance Pattern
Antibiotic
Cefoxitin
Colistin(Methane Sulphonate)
Polymyxin-B
Novobiocin
Resistance pattern
+
+
+
+
1
S. No
2
3
4
+5 Metronidazole
21
Lactic Acid Production
Vegetative cells
Size
Motility
Endospore
Shape
Reaction to stain
Surface colonies onsolid medium
Opacity
Growth in simulatedintestinal fluid
Litmus milk
Production of indole
Production ofhydrogen sulphide
Voges proskauer testAcetyl methanol carbinol
Catalase
Motile
Spore former, oval and Terminal position.oPresent after 48 hrs incubation at 37 C
Ellipsoidal 0.9 to 1.2 by 1.0 to 1.7 micronsterminal,occurring single
Gram stain - Positive Spore staining - Sporesstain red and vegetative cells stains blue.
Medium: PNY, Colony shape: Circular Surface:Smooth Edge : Entire without exception
Opaque
Growth is abundant in ( PNY broth withNaHCO 0.15%, Pancreatin 0.25 % pH 7.23
oincubation 24 hours at 37 C )
pH 4.4 - 4.6
Not produced
Not produced
Not produced
Positive
Positive
Rods, occurring singly, rarely in short chains,variable in filaments by cultural conditions
0.9 by 3.0 to 5.0 Microns
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
S.No. Parameter Characterstic
Morphological & Biochemical Characteristics
SPORLACAcid & Bile resistance
23
A number of peer-reviewed publications have cited, that
spores are therapeutically beneficial as compared to
vegetative cells. Studies have been carried out on spore
forming lactic acid producing bacteria (SFLAB) as probiotics. Due to
their spore forming ability, these bacilli have high heat and acid
resistance. Most cells of ordinary Lactobacilli die at 70°C, while it has
been reported that spore-bearing lactic acid forming bacteria do not
show a decrease in viable cells even after heating in saline at 85°C for
30 min and are also stable in artificial gastric juice (pH 2.0) for a typical
time of application (3 hr).
SFLAB are a group of Gram-positive bacteria, sharing characteristics
common to the genera Bacillus (spore forming, motile) and
Lactobacillus (micro-aerophilic, lactic acid production).
Many factors make SFLAB good candidates for a probiotic use :
(I) They are easily cultured in ‘bulk’
(II) They produce organic acids
(III) Due to spore form, SFLAB are more resistant to heat, which
facilitates the pelleting process used in the mass.
(IV) SFLAB is resistant to the acidity of the stomach, lysozyme, bile
and pancreatic enzymes.
Bacterial spores are produced in nature as a means to survive extreme
environmental conditions enabling long-term survival in conditions
Outer Coat Cortex Inner Coat Exosporium
Plasmamembrane Nuclear
material
Core orprotoplast
Incorporatedmother cellcytoplasm
Corticalmembrane
of germ cellwall
The Bacillus Spore
25
that could otherwise kill vegetative bacteria. The survival of spores
depends on the sporulation during the bio-mass production, when the
vegetative forms go into the spore stage.
The decision to sporulate is very much dependent upon the decline in
nutrients in the immediate vicinity of the live cell. Sensing this, the
bacterium enters an irreversible program of development that results
in the production of a spore. About eight hours later intrinsic to
survival is the structure of the bacterial endospore, that contains, at its
core, a condensed and inactive chromosome.
Additional layers surround the spore, including a peptidoglycan rich
cortex and one or more layers of proteinaceous material referred to as
the spore coat, meant to protect the spore from UV radiation, extremes oof heat (typically up to 80-85 C in most species), exposure to solvents,
hydrogen peroxide and enzymes such as lysozyme. The spore itself, is
dehydrated and if exposed to appropriate nutrients will germinate, a
process taking just a few minutes, allowing water to enter the spore,
breakage and removal of the spore coats, and outgrowth and
resumption of vegetative cell growth. The nutrients of the GI tract in
the distal end beginning in the ileum and caecum provide ambient
environment / conditions and nutrients for the spores to germinate
and exert the beneficial effects to the host, a purposeful and healthy
symbiotic relationship.
Depending on species spores are spherical or ellipsoidal in shape,
between 0.8-1.4 mm in length, have a negative surface charge and are
moderately hydrophobic.
Under conditions of nutrient starvation the growing Vegetative Cell (VC)
will undergo a series of morphological changes that create a Forespore (F)
within the Mother Cell (MC) of the sporangium. After approximately
eight hours the Spore (S) is released by lysis of the MC.
SPORLAC - Life Cycle
MC - Mother Cell
F - Forespore
VC - Vegetative Cell
S - Spore
Sporulation Vegetative Cell Growth
MC F
MC F
VC
S
VC VC
Starvation
Germination
Binary Fission
27
The next question is how do spores germinate, & how is it defined ?
Spore germination, is defined as those events that result in the loss of
the spore-specific properties, is an essentially a controlled natural or
biophysical process during manufacture. It occurs without any need
for new macromolecular synthesis, so the apparatus required is
already present in the mature dormant spore. Germination in response
to specific chemical nutrients requires specific receptor proteins,
located at the inner membrane of the spore. The full molecular details
of the signal transduction process in spore germination are not yet
clear, but reasonable hypotheses can be constructed with the available
information, most of which is derived from studies with Bacillus
subtilis, the genetic and biochemical paradigm spore-former.
SPORLACApplications
29
SPORLAC has a wide range of applications, apart from being
conventionally a solution for gut problems. For various novel
applications, there is an etiology that is linked to gut microflora
imbalance (dysbiosis), and probiotics are quite well studied for their
proven clinical efficacy with their enteric approach. Following are the
applications for which SPORLAC is understood to be useful :
1. Gastroenteritis
2. Aphthous ulcers
3. Recurrent Bacterial Vaginosis
4. Hypercholesterolemia
5. Immunity
6. Urticaria
7. Azotemia
8. Necrotizing Enterocolitis
Inflammation
Infection
Pathogen
Gut barrierdystunction
Tolerance
Immuneresponse
Allergen
SPORLACBalanced
microflora
Aberrantmicroflora
Aetiology & Role of SPORLAC
GASTROENTERITIS
31
Gut Microflora in Inflammation
Inflammation may direct the composition and function of a balanced
normal microflora to become aberrant and immunogenic, leading to
perpetuation of the inflammation and gut barrier dysfunction.
Probiotics like SPORLAC may
1. Counteract the inflammatory process by enhancing the
degradation of enteral antigens,
2. Reduces the secretion of inflammatory mediators,
3. Promote the normalization of indigenous flora and the exclusion
of pathogens.
Source: E Isolauri, P V Kirjavainen, S Salminen, "Probiotics: a role in the treatment of
intestinal infection and inflammation" Gut 2002;50(Suppl III):iii54–iii59.
The common Aetiological factors for gastroenteritis include:
i) Infections ii) Radiation iii) Antibiotic therapy iv) Tube feeding
v) Bacterial agents vi) Viruses
These pathogens produce different types of toxins that can cause
severe or life-threatening dehydration and diarrhoea.
Microbiota
Stress
Acidic fruits
Sharp tooth surface
Vit- B12,
Zinc & Folic acid
GIT Disease
Immune deficiency
APHTHOUS
ULCERS
SPORLAC
SPORLAC
APHTHOUS ULCERS
Aetiology & Role of SPORLAC
33
Aphthous ulcers or Recurrent Aphthous Stomatitis (RAS) or
canker sores, are inflammatory lesions of the mucous lining
of the mouth which may involve the cheeks, gums, tongue,
lips, and roof or floor of the mouth. Possible aetiological factors
include :
1. Trauma & Stress
2. Systemic diseases and Nutritional deficiencies
3. Food allergies
4. Infection
5. Immune disorders
6. Drug induction – Antibiotics & Anti-inflammatory Drugs
The ability of SPORLAC to increase the activity of the phagocytes
must be the key factor in combating RAS, as Porter and Scully have
stated that in RAS, phagocytosis by PMNLs (Polymorphonuclear
leukocytes) and the chemotaxis to PMNL s are defective. SPORLAC
is very useful in Aphthous stomatitis and Glossitis, which may be a
result of Vitamin B Complex deficiency due to denudation of
intestinal saprophytic flora.
According to Dr.S.N.Mathur et.al., SPORLAC is proved most
efficacious in the treatment of Aphthous ulcers within 2-3 days.
According to Dr.Sharma et.al., SPORLAC therapy is one of the best
available methods to treat these recurrent oral ulcerations successfully.
Source: UP State Dental Journal, Vol. II 7-12, 1970.
RECURRENT
BACTERIAL VAGINOSIS
Aetiology & Role of SPORLAC
Replacement of lactobacilliVaginal or oral
Reducion of Lactobacilli Reduction in H O Production2 2
Maintains pH at4.5 by vaginalacidifying gel
Intermittenttreatment
Vaginal or oral
SPORLAC
Overgrowth of BVassociated bacteria
Raised pH
35
Wilson J, Sex Transm Infect 2004;80:8–11.
SPORLAC is administered vaginally or orally in women with
non-specific vaginitis twice daily with for 14 days. Women with
severe infections of Trichomonas and Candidias were not
included in the study. Complete relief from pruritus and discharge was
reported in 91% of the treated subjects. These results were attributed to
lactic acid produced in the vaginal milieu which inhibited the growth
of vaginal pathogens, by SPORLAC. Post menopausal women, had a
slower response to therapy but eventually had complete relief.
Deconjugationof bile salts andinterruption ofentero-hepatic circulation of bile acids
Bile Salts
Dietary Fat
Emulsification
Absorption
CholesterolSynthesis
Inhibithydroxymethyl glutarate Co Areductase, therate-limiting enzymein cholesterol synthesis
SPORLAC
SPORLAC
Aetiology & Role of SPORLAC
HYPERCHOLESTEROLEMIA
37
Hypercholesterolemia (elevated serum cholesterol levels) is
typically due to a combination of environmental and
genetic factors. Environmental factors include obesity and
dietary choices. Genetic contributions are usually due to the additive
effects of multiple genes, though occasionally may be due to a single
gene defect such as in the case of familial hypercholesterolaemia.
A number of secondary causes exist including: Diabetes mellitus type
2, Obesity, Alcohol, Monoclonal gammopathy, Dialysis, Nephrotic
syndrome, Obstructive jaundice, Hypothyroidism, Cushing's
syndrome, Anorexia nervosa, Medications (thiazide diuretics,
ciclosporin, glucocorticoids, betablockers, retinoic acid).
Probiotics like SPORLAC acts by
1. Deconjugation of bile salts and interruption of entero-hepatic
circulation of bile acids
2. Inhibit HMG CoA reductase thereby inhibiting Cholesterol
synthesis
3. Improve elimination
Source: Gilland .S.E. and Speck .M.L (1977). Deconjugation of Bile acids by Intestinal
lactobacilli, Appl.Environ. Microbiol. 33: 15.
Role of SPORLAC
Lumen
Mucins and defensins
Pathogens
Mucus
(5) Production ofanti-microorganismsubstances
(3) Inhibition ofpathogen adhesion
(4) Competitive exclusion ofpathogenic microorganisms
Lamina propria
DCs
IL-10 TGF (6) Modulation of theimmune system
Immature DC
Treg
Th1 Th2Th17
Macrophage
IECs
(1) Enhancement of the epithelial barrier
(2) Increased adhesion tointestinal mucosa
SPORLAC
SPORLAC
SPORLAC
IMMUNITY
39
The immune system can be divided between the innate and
adaptive systems. The adaptive immune response depends on
B and T lymphocytes, which are specific for particular
antigens. In contrast, the innate immune system responds to common
structures called Pathogen Associated Molecular Patterns (PAMPs)
shared by the vast majority of pathogens. The primary response to
pathogens is triggered by Pattern Recognition Receptors, which bind
PAMPs. In addition, extracellular C type Lectin receptors and
intracellular Nucleotide binding Oligomerization Domain containing
protein like receptors are known to transmit signals upon interaction
with bacteria .
Major mechanisms of action of probiotics like SPORLAC include
1. Enhancement of the epithelial barrier
2. Increased adhesion to intestinal mucosa
3. Concomitant inhibition of pathogen adhesion
4. Competitive exclusion of pathogenic microorganisms
5. Production of anti-microorganism substances
6. Modulation of the immune system
Probiotic which have the ability to interact with epithelial and
dendritic cells (DC) and with monocytes/macrophages and
lymphocytes can exert immunomodulatary effect.
Source : Miriam Bermudez-Brito, Julio Plaza-Díaz, Sergio Muñoz-Quezada, Carolina
Gómez-Llorente, Angel Gil Probiotic Mechanisms of Action , Ann Nutr Metab
2012;61:160–174.
Elevated secretion ofTNF- ECP
Increased rate oftransmural passageof enteral antigensin their undegraded forms
Allergicinflammation
in the gut
Imbalance ofi n t e s t i n a lmic ro f lo racomposition
Defective generationof mucosa l IgAresponse to centeralantigens
Oral allergen
1
2
3
4
5
6
SPORLAC
URTICARIA
Aetiology & Role of SPORLAC
41
Isolauri E et al. Am J Clin Nutr 2001;73:444s-450s
There are various suggested targets of probiotic therapy to modulate
immune responses to dietary antigens during the course of allergic
inflammation in the gut. Probiotic bacteria such as SPORLAC are
shown to modulate allergic inflammation by :
1) Altering the immunogenicity of allergens via proteolytic activity
2) Reducing the secretion of inflammatory mediators in the gut
3) Reversing the increased intestinal permeability and enhancing the
degradation of enteral antigens
4) Diverting the gut antigen uptake toward Peyer's patches
5) Normalizing the composition of intestinal microflora
6) Enhancing mucosal Immunoglobulin A (IgA) response to enteral
antigens - TNF (Tumor Necrosis Factor); ECP (Eosinophil
Cationic Protein).
Freep-cresyl-sulfatePortal circulation
HN
Proteins
Peptides
Tyrosine
Putereffective bacteria
Tryptophan Indole
p-cresylOH
CH3
NH
Gut lumen
Liver
Free indoxyl-sulfate
albumin
S
o
CH3
So
Colon
Albumin bound p-cresyl-sulfate & indoxl-sulfate not efficiently removed
by hemodialysis
Kidney
SPORLAC
Prevent the formation of p-cresyl-sulfate & indoxl-sulfate
in the gut
Aetiology & Role of SPORLAC
AZOTEMIA
Hemodialysed patients have more than 100 times higher aerobic
bacteria (putrefactive bacteria) than normal subjects. Probiotics
reduce the putrefactive bacteria by creating the acidic environment
and normalizes the microflora. Probiotics such as SPORLAC
improves nutrition and decreases the toxic substances like DMA
(Dimethyl amine), NDMA (Nitroso dimethyl amine) and toxic
amines. The intake of SPORLAC effectively restores the disturbed
microflora to a normal one by creating the acidic environment which
then prevent the carcinogenic aerobacteria like E.coli.
Haemodialysis (HD) cannot effectively eliminate protein-bound
solutes as opposed to small water-soluble solutes, and the
accumulation of protein-bound uremic toxins such as p-cresol and
indoxyl sulphate is suggested to be related to complications such as
Azotemia and mortality in HD patients. Serum concentration of
indoxylsulphate & p-cresyl sulphate is increased markedly in uremic
patients with because insufficient renal clearance.
A pilot study found that SPORLAC supplementation for 16 weeks
attenuated BUN levels & enhanced survival of 5/6th nephrectomized
rats while slowing the progress of renal injury induced by reduced
nephron mass. Supplementation of probiotics prolonged the life of
uremic rats, thereby showing significantly less mortality (p < 0.05) and
highest BW gain.
43
NECROTIZING ENTEROCOLITIS
Aetiology & Role of SPORLAC
Prematurity
Formula feeding
Intestinal ischemia
Increased PAF production anddecreased PAF degration
Excess PAF
Bacterial colonization
TLR signaling
Increased TLRExpression;
activation of TLR
Breach of mucosalbarrier integrity
Bacaterial translocationProduction ofPro-inflammatory Cytokines
SPORLAC
SPORLAC
45
Necrotizing Enterocolitis (NEC) is a medical condition primarily seen in
premature infants where portions of the bowel undergo necrosis (tissue
death). NEC has no definitive known cause. An infectious agent has been
suspected, as cluster outbreaks in neonatal intensive care units (NICUs)
have been seen, but no common organism has been identified.
Prematurity, Intestinal ischemia and Bacterial colonization leads to
increased production and decreased degradation of Platelet Activating
Factor (PAF). Excess of this PAF increases the activation and expression
of TLR's (Toll Like Receptors), increased signaling of TLR causes
Bacterial colonization, production of proinflammatory cytokines, Breach
of mucosal barrier integrity. All these leads to Necrotizing Enterocolitis in
infants.
Probiotics offered as nutritional supplements that act in the intestine of
the host organism by regulating the local bacterial flora. They act by
improving gastrointestinal permeability and increasing the resistance of
the mucosa against bacterial penetration. Regarding the protection
mechanisms, they: (i) Increase the resistance of the intestinal barrier
against the passage of bacteria and their toxins (ii) modify the host
response in relation to microbial products (iii) increase the mucosal
response to IgA (iv) produce bactericidal substances and (v)
competitively exclude potential pathogens.
SPORLAC Mechanisms of Action
47
First, it should be understood that the microflora in the adult
human body consist of an enormous biomass of more than
100 trillion bacteria of more than 400 different species, which
generate intense metabolic activity, mainly in the colon, and play an
important physiological role in the host. The metabolic activities, that
exert therapeutic benefits are Proteolysis, Lipolysis and in addition to
these activities SPORLAC induce enzymes ß-galactosidase,
Glycolase and Lacatate dehydrogenase.
Lactic Acid Production
The ability of conversion of lactose to lactic acid, has been used with
great success in treating lactose intolerance. Individuals with lactose
intolerance, are unable to metabolize lactose due to a deficiency of
essential enzyme Lactase. Lactic acid, by lowering the pH of the
intestinal environment to 4.0 or 5.0, inhibits the growth of putrefactive
organisms and E.coli which survive in optimal pH of 6.0 to 7.0.
Volatile acids produced during fermentation, also possess some
antimicrobial activity, under conditions of low oxidation-reduction,
potential.
Production of Bacetriocins
SPORLAC is generally recognized as safe (GRAS) and the
preservative effect exerted by SPORLAC is mainly due to the
production of organic acids (such as lactic acid) which result in
lowered pH. It also produces antimicrobial compounds including
hydrogen peroxide, CO , diacetyl, acetaldehyde, D-isomers of amino 2
acids, reuterin and bacteriocins.
The metabolites of SPORLAC, that exert
antagonistic action against putrefactive micro-organisms
and the action of some of the metabolites are summarized
Metabolic Product
Carbon dioxide
Diacetyl
Hydrogen peroxide /
Lactoperoxidase
Interaction with arginine
binding proteins
Inhibits
decarboxylation
Mechanism of Action
Oxidizes basic proteins
Reduction of membrane
permeability
Lactic acid
Bacteriocins
Undissociated lactic acid
penetrates the membrane,
lowering the intracellular
pH. It also interferes with
metabolic processes such as
oxidative phosphorylation.
Affects membrane, DNA -
Synthesis and protein
synthesis
49
Bacteriocins are ribosomally synthesized antimicrobial peptides that
are active against other bacteria, either of the same species (narrow
spectrum), or across genera (broad spectrum). In recent years,
bacteriocin producing probiotics have attracted significant attention
because of their GRAS status and potential use as safe additives for
food preservation. Some of the bacteriocins, produced by SPORLAC
are Acidolin, Acidophilin, Lactain B & F.
Vitamin-B Synthesis
Mammalian cells cannot synthesize folate (an important B-group
vitamin, participates in many metabolic pathways such as DNA and
RNA bio-synthesis and amino acid inter- conversions); therefore, an
exogenous supply of this vitamin is necessary to prevent nutritional
deficiency. Milk is not a rich source of dietary folate. However, many
dairy products are processed using microbial fermentations in which
folate can be synthesized, significantly increasing folate
concentrations in the final product. Numerous researchers have
reported that SPORLAC, has the ability to synthesize folate.
SPORLAC confer physiological benefits to the hosts:
1. Improving digestive capacity of the consumed milk proteins
2. Improving absorption and mineral metabolism [Calcium, Phosphorus & Iron]
3. Stimulates secretion of gastric juices
4. Improving peristaltic movements of the digestive tract
5. Serving as an energy source in cellular respiration
SPORLAC* metabolites
and EPIGENETIC mechanism
Antagonism ofpathogens and
putrefactive bacteria
Colonocytes
Reduced colon cancer riskand IBD inflammation
De novo lipogenesis
Controlled serum lipids andcholesterol
HDACinhibitor
Epigenetic regulation
Probiotics
Prebiotics
+ Trophic and anti-neoplastic
effectFermentation
SCFA
Reduced pH
Ca++Mg++Increased mineral absorption
Fewer toxic bacterialmetabolitesR
educed cancer risk
51
Probiotics when ingested orally ferment indigestible carbohydrate
from food and produce short chain fatty acids (SCFA). Anaerobic
bacteria also produced SCFA from peptide, polysaccharide, protein
and oligosaccharide and are the final product of bacteria’s activity into
the GI tract.
SCFA can lower the lipids in blood through blocking synthesis of
hepatic cholesterol and/or through redirecting plasma cholesterol
toward the liver SCFA are potential modulator of food intake and
energy sensing process into the brain, which might indirectly play an
important role in reduction of cholesterol and other metabolism
deranging lipids into the host body.
SCFA involve reduction of pH which is important phenomenon for
increased absorption of Ca, Mg minerals and reduced pH results in
fewer toxic bacterial metabolites thus reduces cancer risk.
Epigenetic Regulation:
SCFA effects on HDAC ( Histone Diacetalyse ) inhibitors, a class of
compound that interfere with the function of HDAC inhibition will
have a effect on psychiatry and neurology as mood stabilization and
epileptics. And this concept investigated for possible treatments for
cancer and inflammatory diseases like IBD.
SPORLACClinical Experiences
53
®Sharma et al., noted that SPORLAC therapy is one of the best
available methods to treat recurrent oral ulcerations. At the dosage
level of two tablets thrice daily for five days, (corresponding to 120
million spores of L.sporogenes per day), aphthous stomatitis was cured
in two to three days.
Clinical Trials In The Treatment of Aphthous Stomatitis & Glossitis
Mathur, S.N. et al. (1970). UP State Dental Journal 11:7-12
Lactic Acid Bacteria Is Beneficial For Oral
Aphthous Ulcerations - A Preliminary Report
Hashim, B.Y; Rahman, R.A; Philip, K. Annal Dent Univ Malaya 1999; 1: 43 _46
Recurrent aphthous ulcers of the mouth are difficult to treat because
of no known definite aetiology. This paper presents the use of lactic
acid bacteria thought to modulate the host immune response to affect
improvements in the disease. Twenty-five patients with the disease
were treated with six lactic acid bacteria capsules daily for a period of
six months, and their responses were evaluated. Seventeen patients
(73.9%) became free of the disease six months later, while six (26.1 %)
experienced very dramatic improvements. Two patients were lost to
follow-up. It is concluded that lactic acid bacteria is beneficial in the
treatment of recurrent aphthous ulcers of the mouth.
Acidic
Alkaline
Change in Vaginal pH with Treatment
Num
ber
of
Subje
cts
Duration of Treatment (Days)
45
40
35
30
25
20
15
10
5
00 3 5 7 14
Complete relief of pruritis and discharge was reported by
91 percent of subjects. These results were thought to be
due to a beneficial change in vaginal acidity via lactic acid
production by SPORLAC. Postmenopausal subjects had a
slower response to therapy but eventually had complete
relief as well.
55
Use of SPORLAC in the Treatment of
Non-specific Vaginitis
Shirodkar NV, Sankholkar PC, Ghosh S, Nulkar SM. Indian Pract 1980;33:207-210.
Non-specific vaginitis is caused by a variety of pathogens including
Staphylococci, Streptococci, Pneumococci and E.coli. It may also be
induced by a variety of causes including chemicals, drugs, surgical
procedures, trauma and foreign bodies. SPORLAC administration to
increase the vaginal acidity by the action of the Lactobacillus on
glycogen in the vaginal epithelial tissues was adopted in a clinical trial
on 44 patients. The patients were divided into two groups:
Group 1: Twelve patients suffering from leucorrhea (white discharge)
following cervical surgery.
Group 2 : Thirty two patients with nonspecific vaginitis without
previous therapy. Of these, 26 were in the reproductive age and 6 were
menopausal. The change in vaginal pH following treatment with
SPORLAC tablets over a period of two weeks is shown in the figure:
Most of the cases showing persistently alkaline pH were post-
menopausal, where acid could not be produced in sufficient amounts
due to low substrate glycogen levels. Glycogen levels depend upon
circulating estrogen.
The Response to Treatment in Both Groups
Type of Response
Number of Cases
Group 1 Group 2 Total
Quick Response &
Complete Relief
8
(67%)
2
(16.5%)
2
(15.5%)
12
(100%)
0 0
26
(81.25%)
4
(12.5%)
2
(6.25%)
32
(100%)
34
(77.25%)
6
(13.60%)
2
(9.15%)
44
(100%)
0
Delayed Response But
Complete Relief
Improvement But Not
Complete Relief
No Relief
Total
57
In comparision with a clinical trial using M.T.P. vaginal pessaries
containing broxyquinoline and brobenzoxeldine where only 26.67%
of cases studied were cured, it can be seen that SPORLAC therapy is
the better alternative in the treatment of non-specific vaginitis.
SPORLAC therapy provided complete relief to 91% of the patients
and partial relief to approximately 9%.
Vaginal administration of a commercial formulation of SPORLAC
tablets called MYCONIP was given to 44 women with non-specific
vaginitis twice daily for 14 days. Subjects with Trichomonas or
Candida vaginitis were excluded from the study. Complete relief of
pruritis and discharge was reported by 91 percent of subjects. These
results were thought to be due to a beneficial change in vaginal acidity
via lactic acid production by SPORLAC. Postmenopausal subjects
had a slower response to therapy but eventually had complete relief as
well.
Hypocholesterolemic Effects of SPORLAC : Clinical Studies
200
150
100
50
0Total SerumCholesterol
(p,0001)
LDLCholesterol(p<0.001)
HDLCholesterol
(p<0.05)
Ser
um
lev
els
mg/
dl
Before
After
350
300
250
Hypercholesterolemia
59
Preliminary observations on effect of SPORLAC
on serum lipid levels in hypercholesterolemic patients.
Mohan JC, Arora R, Khalilullah M.Indian J Med Res. 1990 Dec; 92:431-2.
Short term hypolipedemic effects of oral SPORLAC therapy (369
million spores per day in tablet form) were studied in 17 patients (15
men and 2 women in the 32-61 year age group) with type II
hyperlipidemia in an open label fixed dose trial. Total serum
cholesterol, LDL-Cholesterol and total cholesterol to HDL-
Cholesterol and LDL-Cholesterol to HDL-Cholesterol ratios
(p<0.001) was reduced significantly over a period of three months.
HDL-Cholesterol was marginally increased (43±7mg/dL vs 46.8±8.9
mg/dL, p<0.05), there was however no change in serum triglyceride
levels.
Atherogenic lipid ratios observed were as follows
Total/HDL-Cholesterol 24.0% decrease, LDL/HDL-Cholesterol
33.4% decrease. No adverse effects of therapy were noted, except
constipation in one patient.
Cholesterol Precursor of Bile Salts
Conjugated Bile Salts
Taurine - Conjugated Bile Glycine - Conjugated Bile Salts
Lactobacillus Bile Salt Hydrolase Lactobacillus
Hydrolysis
Amino Acid Group
Taurine Reabsorbed into GIT Glycine
De-Novo Synthesis
Free CholicAcid
Free CholicAcid
Excreted into the Faeces
The Basic Mechanism of Action of SPORLAC as hypolipidemic agent
Cholesterol-Lowering Effects of Probiotics and Prebiotics:A Review of in Vivo and in Vitro Findings
Lay-Gaik Ooi and Min-Tze Liong Int. J. Mol. Sci. 2010, 11, 2499-2522
61
The basic mechanism is to prevent the absorption of fat and
cholesterol in the form of micelle for which bile salts are essential in
the small intestine. Bile salts and cholesterol have enetro-hepatic
circulation and any method of interfering in this process will hinder
their absorption and consequently the serum levels. Bile salts are
deconjugated in the small intestine by the bacterial flora to a small
extent and these deconjugated salts can not be incorporated in the
micelle. Implantation of large quantities of Lactobacilli to facilitate
this deconjugation process is an attractive alternative method to treat
hyperlipidemias but has not been tried so far. Availability of spores of
SPORLAC which germinate the vegetative form in the gut after oral
intake has solved this problem.
Jaundice
Septicemia
Cord Infection
Vomiting
3
3
3
3
The Symptoms Experienced by Subjects in Addition to Diarrhoea
Diarrhoea
Jaundice
60
3
49
3
3
81.7%
100%
100%
Condition Cases Treated Cases Cured Success Rate
3Constipation
Results of Diarrhoeal Symptoms
63
Most of the subjects (about 80%) had a history of breast-feeding.
About 19% were both breast and bottle-fed and 1% were bottle-fed.
The average duration for recovery was 1.8 days.
As compared to the normal practice of administration of antibiotic
and anti-diarrhoeal mixtures, the complicating side effects were not
seen in the series of SPORLAC trials. The average recovery time of 1.8
days helped to reduce dehydration in the subjects to a great extent. In a
similar study in Japan a comparable success rate of 78.4% with
SPORLAC treatment for infantile diarrhoea was obtained. An earlier
study in India by Mathur et al. Found the average time for
improvement in diarrhoeal conditions to be two to three days, with
treatment.
A total of 60 cases of diarrhoea with watery stool frequency greater
than 6 were examined for efficacy of SPORLAC treatment. Based on
the suggested dosage level of SPORLAC at 5 million spores per
kilogram body weight, each neonate was given a spore level of about
15 million spores per day.
®SPORLAC (L.sporogenes) in Diarrhoea
Dhongade, R.K. Anjaneyulu .R Abstract from Maharastra Medical Journal
SPORLAC in Other Clinical Conditions
65
SPORLAC & Lactose Intolerance
Individuals with a deficiency of the enzyme β-galactosidase [lactase]
suffer from abdominal distress, such as diarrhoea, flatulence, bloating
etc, when they consume milk and milk-products. However they are
tolerant to Yogurt enriched with SPORLAC. It was found to have
considerable lactase activity when tested in vitro. Lactobacilli provide
considerable amounts of the enzyme Lactase, which hydrolyzes the
lactose. The hydrolyzed lactose is converted to physiological amounts
of lactic acid.
SPORLAC & Antibiotic Associated Diarrhoea
Treatment with antibiotics, has been associated with diarrhoea, due to
the imbalance of the GI microflora. The main causative organism is
C.difficile, which causes diarrhoea even after discontinuation of the
antibiotic. Treatment with SPORLAC, has replenished the intestinal
microbiota, and causing a reduction in the frequency and duration of
diarrhoeal episodes on administration of either powder or tablet form.
In infants and neonates, there was significant reduction in the number
of episodes of diarrhoea due to viral infections and also improved the
feeding pattern, when administered with ORS.
SPORLAC & Intestinal Infections
Dysbiosis, causes an imbalance in the gastrointestinal milieu may
occur due to antibiotic use, immune suppression, allergy insult and
stress. In order to achieve, optimal balance between beneficial bacteria
and pathogenic bacteria, administration of probiotic microorganisms
helped alleviate the abdominal distress in a variety of clinical
conditions. The Summary of studies are cited in table.
SPORLAC & Other Associated Disorders
IBS [Irritable Bowel Syndrome]
A multifactorial disorder, with varied aetiology of episodic
abdominal pain, distension, flatulence, alternating episodes
of diarrhoea and constipation, symptoms have shown
improvement on administration of SPORLAC.
Oral Conditions
Apthous ulcers and Stomatitis – SPORLAC administered in
doses of upto 120 CFU/day resolved the symptoms of
apthous ulcers and stomatitis in as little as 2-3 days
Dental caries
SPORLAC was administered to children in whom salivary
Streptococcus mutans was detected. Treated individuals were
tested for Streptococcus and there was a significant reduction,
suggesting that SPORLAC formulations in paediatric
population may be effective in preventing dental caries.
67
Summary of the Nutritional &
Therapeutic Benefits of SPORLAC
Lactobacilli
Inhibition of Growth of pathogens
Alleviation of Lactose Intolerance
Enhancement of Immunity
Therapeutic Benefits
Nutritional Benefits
Restoration of the Ecological Balance of the GI Flora
Detoxification
Elimination of end products with carcinogenic potential
1. Vitamin B & Folate Production2. Improved Digestive Capability of ingested food-stuffs3. Improved bio-availability of minerals and trace elements
Diseases influenced by gut microbial metabolism. The variety of systemic diseases that are directly
influenced by gut microbial metabolism and its influence on other mammalian pathways, such as the innate
immune system, are shown. Specifically highlighted are the metabolic pathways involved in drug
metabolism and obesity that are directly influenced by the gut microbial content. Ags, Antigens; Clostridium
bolteae; Dendritic cells; SCFA, (short-chain fatty acid) TLR, Toll-Like Receptor. Kinross et al. Genome
Medicine 2011 3:14. [Gastroenterology Research and Practice Volume 2012, Article ID 872716, 16 pages]
Emerging areas for SPORLAC Research
The importance of the microbiota organisms & the manifetstions
Hypertension/Ischemicheartdisease
Asthma/atopyhygeine hypotheticExagerated innate immune responseup regulation of regulatory T cellsafter capture of Ags by DCs Bifidobacteria, gram +ve organisms clostridia
Gut-brain hypothesis 1. Autism C. Bolteae/ clostridia sporesmechanism unknown2. Mood: depression, anxiety
Peripheral vascular diseaseresult of metabolic syndrome-altered lipid deposition/metabolism
Altered xenobiotic/ drug metabolism
e.g. paracetamol metabolism: predose urinary p-cresol sulfate leads to postdose urinaryacetaminophen sulfate: acetaminophen glucuronide.Bacterially mediatedp-cresol reduces the effective systemic capacityto sulfonate acetamtnophen.
Altered enterohepatic circulation of bileBillary disease
Diet high in red meat and animal fatLow SCFA/butyrateHigh fecal fatsLow vitamin absorption7a dehydroxylating bacteria:cholic acid deoxycholic acid (cocarcinogen)Low in H S metabolizing acteria2
Diet high in red meat and animal fatLow SCFA/butyrateHigh fecal fatsLow vitamin absorption7a dehydroxylating bacteria:cholic acid deoxycholic acid (cocarcinogen)Low in H S metabolizing acteria2
Colon cancer
Obesity/metabolic syndrome
Bacteroidetes and Actinobacteria in obeseAltered energy/lipid metabolismHigher relative abundance of glycoside hydrolases,carbohydrate-binding modules,glycosyltransferases, polysaccharide lyases, and carbohydrateesterases in the BacteroidetesTLR mediated
Inflammatory bowel disease
Hygiene hypothesisAltered immune response:TLR signalingLess microbial diversityActivation of specific species e.g., Escherichia
69
The anti-inflammatory and Immuno-modulating properties of
SPORLAC and other lactic acid bacteria, have shown an impact on
the symptoms of arthritis. In randomized studies, men and women
with an average age of 62 years, received SPORLAC of 2 Billion CFU
for 60 days, in addition to their regular arthritis medications.
Evaluations of pain and disability at 30 and 60 days of the treatment,
showed statistically significant reduction in pain scale scores and
disability as compared to placebo. A reduction in c-reactive protein
was seen, in the treatment group but not the placebo group. This
development coupled with well controlled studies could be just the
beginning of a new era, wherein probiotics could call the shots in not
just acute but chronic disorders and also the future may see the
development of probiotics as targeted carriers of vaccines, hormones
and other bio-therapeutic agents.
Bacillus coagulans : a viable adjunct therapy for
relieving symptoms of rheumatoid arthritis according
to a randomized, controlled trial
Mandel DR,Eichas K and Holmes J BMC Compl. & Alternat. Med. 2010, 10:1
Diagnosis
Number
of
subjects
Treatment:
Lactobacillus
spores (millions)
Effective
RatesRemarks
Acute and
chronic
intestinal
catarrh
Diarrhoea 15
75-600/day in
divided doses
for 3-12 days.
100.00%
38
100-600/day in
divided doses
for 2-12 days
86.80%
Constipation 10
300-750/day in
divided doses
for 2-10 days.
70.00%
Abdominal
intestinla
fermentation
9
300-600/day
in divided doses
for 3-14 days.
100.00%
Recovery from diarrhoea to
regular, normal stools
from third to fourth day.
Recovery from diarrhoea
to regular normal stools;
general symptoms
including anorexia
improved.
Recovery to normal stools &
disappearance of abdominal
distension.
Vomiting & nausea
disappeared; appetite
improved; stools became
normal & regular; diarrhoea
& stomach ache cured.
Dyspepsia
Infantum
Allergic skin
diseases
Miscellaneous
26
5
10
100-200/day in
divided doses
for 1-7 days.
200-450/day in
divided doses for
4-12 days
20-50/day in divided
doses for 4-20 days.
84.60%
80.00%
80.00%
General condition &
nature of stools improved.
Frequency of stools
decreased to half or less
than that before
medication.
Obvious eruptions of
strophulus and eczema
decreased from the third
day (topical therapy
employed concomitantly).
Response seen in anorexia
of nervous type and
malnutrition in infants.
Clinical Studies with LACBON*
* is the L.sporogenes / B.coagulans brand of erstwhile Sankyo Japan
71
Author Year ConditionStudy
Population
Daily
Dose Results
La Rosea
et al2003
Antibiotic
associated
diarrhoea
98 childrenNot
specified
71%of children in treatment
group had resolution of diarr-
hoea versus 38% in placebo
group; duration significantly
shortened in treatment group.
Dolin et al 2009
Irritable bowel
syndrome
diarrhoea
predominant
(IBS-D)
52 adults2 billion
cfu
Decreased number of bowel
movements, no statistically
significant change in other
IBS symptoms
Hun L 2009 IBS-D 44 adults
800
million
cfu
Statistically significant
improvements in abdominal
pain and bloating in
treatment group compared to
placebo.
Chandra RK 2002Acute retrovirus
diarrhoea112 newborns
100
million
cfu
Statistically significant
decrease in frequency and
duration of diarrhoea in
treatment group compared
to placebo.
Sari Fn et al. 2011
Narcotizing
enterocolitis
(NEC)
221 very-low-
birth weight
(VLBW)
neonates
(<1500 g,
<33 weeks)
350
million
cfu
No effect on rate of death or
NEC on VLBW infants;
significantly improved
feeding tolerance in
treatment group
Dutta p et al 2011
Diarrhoea with
dehydration
(diverse etiology)
148 infants
16-24 months
Not
specified
No therapeutic effect on
management of acute
dehydrating diarrhoea of
diverse etiology, including
rotavirus associated diarrhoea
in children.
Kalman
DS et al2009
Intestinal pain,
gas bloating 61 adults
2 billion
cfu
Significant reduction in
intestinal gas and pain scores;
strong trend toward improved
abdominal distention scores;
improved quality of life scores.
= X X
Shelf - Life Water Activity Temperature Oxygen
Shelf - Life Rule-of Thumb Recommendation
The Shelf-Life Equation
potencypoint
#1
potencypoint
#2
potencypoint
#3
potencypoint
#4
Powderproduction
Bulkdosageforms
Finaldosageforms
Boxing &Labeling
Distri-bution
StoreShelfing
Consum-ption
End of shelf life
Sanzyme Guarantees CFU count All the way to potency point #4 when delivering to final packaging material
Illustrative example of potency decline
Important Terms:
CFU - CFU = Colony Forming Units ,One CFU = one live bacteria cell
[“Live” defined as being able to multiple and thereby form a colony ]
Potency is minimum number of CFU in a Sachet or in one dosage form of either
Capsules or Tablets
5.5B 5B 1B6.5B
= X X24 months < 0.15a
o< 25 C < 5%
73
In the early 1973s after the transfer, of the parent culture and the
technology to manufacture, SPORLAC was handed over, then came
the role of R&D team of Sanzyme. Over the years, Sanzyme has
developed a trademarked technical process known as PROBITECH
for the manufacture of Probiotics for human application and also
those useful in veterinary, aquaculture and bio-remediation.
The process of PROBITECH has evolved over 4 decades, as the
challenges faced for developing, designing and manufacturing.
The Basic Steps in PROBITECH
1. Selection of Strains : Though the task looks seemingly easy, the
compatibility of the species if spore forms are combined with
vegetative forms or vegetative forms combined with other vegetative
forms
2. Purification & Drying : Once the strain/strains are chosen, the
fermentation process is decided upon and once the desired strains are
isolated, they are purified of impurities and are tested for bacterial
contamination and other tests for ensuring that the desired strain is
obtained. The process of drying with ambient temperature is
esssential, as spore forms withstand the process of drying, but
vegetative forms may not be able to withstand the temperatures other
than the optimal.
3. Manufacturing : Once the strain/strains are obtained, compatible
excipients are chosen to ensure that the spores or vegetative forms do
not germinate after manufacture either in the capsule, tablet or sachet
TMPROBITECH
The Science & Technology of Manufacturing Probiotics
PROBITECH
1 28
5 46
Shelf-Life
37
OtherRegulatory
Claim
Potency GI TransitSurvival
Dosage Form
Strain/Strains
Indication
Product
Summary of Probitech Technology
IngredientsCompatibility of Probiotics
Excipients & Packaging are crucial
Physiological State of Bacteria
When Prepared in the Product
OxygenIn combi-Probiotic blends the
fine art of maintaining aerationwithout altering the potency of
the blend is crucial
Temperature [Critical Factor]
Manufacture & Storage
Moisture Levels & Water Activity [a ] Crucialw
Storage of manufactured products can be increased
pHIn-vitro & In-vivo Survival
Gastric Transit Time[1-2hrs.]Quiescent Cell-Enhanced
Survival
75
form. During manufacture, correct compression of the tablet without
excess pressure and temperature must be maintained for obtaining
finished formulations. The blend in the case of combi-probiotic when
filled in the capsule must be maitained at the correct temperature while
filling and packing into either strips or blister packs.
4. Shipping of Finished Goods : The probiotic either single or the
blends are formulated in ideal conditions and ambient temperatures,
however as the climate in different geographical regions of India or the
globe are not ambient, hence a suitable packaging must be designed to
ensure that the probiotic formulations are able to withstand both hot
and cold climates. However certain formulations may need to be
stored in refrigerated conditions
5. Ensuring Shelf-Life & Potency : In order to ensure that the desired
therapeutic benefits are conferred, the desired potency of the probiotic
must be maintained to the end of shelf-life and the viability of either
the single probiotic or the blend must be maintained as there would be
some loses during the manufacturing stages and adequate overages
must be added to ensure that label claims are met as per the norms of
the drug authorities.
In addition to SPORLAC Powder and SPORLAC DS Tablets, with PROBITECH,
innovative blends have been developed in the following categories:
Life Style – Cholesterol Busters & Stress Busters
Renal Care – Prevention of Renal Calculi, Prevention & Treatment of Azotemia
Womens Health- Oestrogen recycling, Bacterial Vaginosis and UTIs
GI Care – Oro-care, Liver Disorders, IBS/IBD,Diarrhoea
ENT and Immunity
Rheumatoid arthritis – Under study
Citations
1. “Screening of probiotics for acid and bile tolerance was evaluated and B.coagulans strains
were tolerant to bile concentrations over 0.3% (w/v)” – Int J of Food Micro; 61
(2000):193–197
2. “One single Bacillus strain is often able to produce several types of molecules stable over a
wide range of pH and temperature ….these substances are usually protein- and peptide-
based compounds such as enzymes, bacteriocins and lipopeptides which are inhibitory
compounds, mainly active against food-borne and gut pathogens”- Science against
microbial pathogens: communicating current research and technological advances -
FORMATEX 2011
3. “…..The Bacillus coagulans-based product was effective in improving the quality of life and
reducing gastrointestinal symptoms in adults with post prandial intestinal gas related
symptoms and no GI diagnoses” - BMC Gastroenterology 2009, 9:85
4. “.....Immunomodulatory and inflammaotry effects …. Lessens the symptoms of arthritis.
….. Safe and effective for patients suffering from Rhuematoid arthritis” - BMC
Complementary and Alternative Medicine 2010, 10:1
5. “The unique characteristics of Lactosporin a novel antimicrobial protein produced by
Bacillus coagulans, including its antimicrobial activity against pathogenic micro-
organisms, indicate that it may have potential for application in foods ...” - J of Appl
Microbi106 (2009) 1370–1377
6. “... results indicate that the administration of B. coagulans …. improves the intestinal
envi ronment , de fecat ion f requency, feca l charac ter i s t ics and dermal
characteristics.[reduction in comedones]” - Microb. Ecol. in Health and Dis. 2002; 14:
4–13
7. “.....results of this toxicological safety assessment indicate that B coagulans does not
demonstrate mutagenic, clastogenic, or genotoxic effects ... and since the suggested
human dose is in the range of 100 x 106 to 3 x 109 CFUs, this gives a safety factor ranging
from 3173 to 95,200 times.” - Food and Chemi.Toxicol. 47 (2009) 1231–1238
77
8. “....the probiotic strain Lactobacillus sporogenes shows a significant stimulation of the cell
mediated immunity and humoral immunity” - Int. J Therap. Appl. Vol.3, 2012, 32 – 38.
9. “.....Probiotic bacteria have a potential role in regulation of intestinal inflammation and
also play a role in intestinal immune homeostasi …. Commensal bacterial strains,
including probiotic bacteria, selected for their properties to activate an epithelial
inhibitory response may be a rational nutritional intervention in IBD patients” - Eur. J
Clin. Nutr. (2002) 56(3):S60–S64
10. “Perturbation of the intestinal microbiota may lead to chronic diseases such as
autoimmune diseases, colon cancers, gastric ulcers, CV disorders, functional bowel
diseases and obesity … but the use of probiotics has led to promising results by restoring
gut flora and alleviating disease” - Gastro. Res. and Pract.Volume 2012
11. “A one-year chronic oral toxicity study combined with a one-generation reproduction
study was conducted to further investigate safety of long-term consumption ….is safe for
chronic human consumption at an accumulative daily intake of 9.38 x 1010 CFUs per
day [human]” - Food and Chem. Toxicol. 49 (2011) 1174–1182
12. “Probiotics will provide a safer way to treat the once before incurable, deleterious ailments
like cancer… It will not be an exaggeration to say that probiotics in the future will be the
reigning therapeutic agents” - J Res. in Biol.(2012) 2: 102-113
13. “.... to aid in the digestion of lactose and fructose could be used to prevent occurrence of
intestinal symptoms in individuals sensitive to these carbohydrates and in patients with
pancreatic exocrine deficiency disorders such as chronic pancreatitis and cystic fibrosis, in
which only small amounts of the normal amount of pancreatic enzymes are excreted B
coagulans could contribute to the digestion of the protein and carbohydrates in their
meal” - Beneficial Microbes, 2010; 1(1): 31-36
14. “The intestinal microflora is a positive health asset that crucially influences the normal
structural and functional development of the mucosal immune system ... The flora has a
collective metabolic activity equal o a virtual organ within an organ, and the mechanisms
underlying the conditioning influence of the bacterial on mucosal homeostasis and
immune responses are beginning to be unraveled for treating ... several infectious,
inflammatory and neoplastic disease processes”- EMBO Reports (2006) 7, 688–693 rld.
15. “ A serious problem of effective pro-biotic usage is the survival of the microorganisms in
the stomach and intestines after administration. If spores are used as probiotics instead of
vegetative cells, the survival and subsequent inoculation levels might be much higher
…..the higher resistance to external factors such as toxic compounds, extremes of
temperature, mechanical force etc....” - TRENDS in Biotech. Vol.21 No.8 Aug. 2003
16. Babar V, Thomas R and Bhaskar M, " Immunomodulatory Activity of Lactobacillus
Sporogenes ( Sporlac )." Int.l Jl of Therap. Appl. Vol.3, 2012, 32 - 38.
17. Yueh-Ting Tsai & Po-Ching Cheng & Tzu-Ming Pan " The immunomodulatory effects of
lactic acid bacteria for improving immune functions and benefits." Appl Microbiol
Biotechnol (2012) 96:853–862.
18. Sharma, J.K , Kapoor, K.K , Mukhija R.D, " Clinical Trial of Sporlac in the Treatment of
Recurrent Apthous Ulceration." U.P State Dental Journal Vol. 11, January 1980, 7-12.
19. Mathur, S.N. et al." Clinical Trials In The Treatment of Apthous Stomatitis & Glossitis."
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Aphtous Ulcerations - A Preliminary Report." Annal Dent Univ Malaya; 1999; 1: 43 _46.
21. Wilson J, " Managing recurrent bacterial vaginosis."Sex Transm Infect 2004;80:8–11.
22. Shirodkar NV, Sankholkar PC, Ghosh S, Nulkar SM " Use of Sporlac In The Treatment of
Non-specific Vaginitis." Indian Pract. 1980;33:207-210.
23. Dr. R.C. Arora , " To Evaluate The Role of Sporlac (Lactobacillus Sporogenes) In Jaundice
Patients." MLB Medical collage Jhansi.
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sporogenes in patients with Dyslipidemias: an open fixed dose trial." Indian Heart J. 1990
Sep-Oct;42(5):361-4.
25. Lay-Gaik Ooi and Min-Tze Liong " Cholesterol-Lowering Effects of Probiotics and
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SPORLAC Family
Diarrhoea AAD Aphthous Ulcers
Gastroenteritis
Bacterial Vaginosis
Powder Tablet Powder / Caps
In Diabetes Zinc & Chromium Deficiency
Acute Infectious / Non Infectious DiarrhoeaImmunity
Powder Tablet Powder
HypercholesterolemiaHyperoxaluria Azotemia
Capsule Capsule Capsule
1
The Leader...
Each tablet of SPORLAC Tablet contains 60 million spores
needing 6 tablets of dose in a day (2 tablets x 3 times).
The launch of SPORLAC DS tablet containing 120 million
spores, facilitates convenience of one tablet 3 times in a day.
Plot No.13, Sagar Society, Road No.2, Banjara Hills, Hyderabad - 500 034, A.P., INDIA.
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