Strains and species of lactic acid bacteria in fermented (yogurts): effect on in vivo lactose digestion14 Margaret C Martini, Eric C Lerebours, Nabil M Berrada, Jean M Antoine, and ABSTRACT tolerated than

Lactose in yogurt lactose in other dairy

activity in vivo.

of microbial To evaluate

fl-galactosidase the ability

oflactic

acid

to digest

mixtures

bacteria

ofstrains

garicus,

lactose

in vivo,

individual

production

similarly

or Bijidobacterium

specific

/3-gal

from plete

was monitored.

improved

lactose

activity.

The

marginal improvement lactose digestion with

that

total

/3-gal

digestion,

was

perhaps

yogurts

not

the

lactose species

(containing

subsp therm ophilus bulgaricus) and fermented of S thermophilus, L bul-

subsp species

bijidus)

in microbial f3-gal activity were produced. fed to healthy people who cannot digest drogen

which digests strains and

salivarius

delbrueckii

L acidop/zilus,

Lin, Susan A Savaiano

with live bacteria is better foods, partly because of the

(f3-gal), of different

ofStreptococcus

and Lactobacillus milks (containing

Wei-Jin Dennis

Selected lactose,

All yogurts

digestion,

varied

dramatically

regardless

response

that

and

oftheir

to fermented

total

milks

strate transport.

or

varied

B bijidus milk to nearly comL bulgaricus milk. The results suggest limiting

factor

in promoting

yogurt

fl-gal.

mental

yogurts

The

lactose

WORDS

erance,

Lactose,

maldigestion,

intol-

deficiency,

Lactobacillus have

acidophilus

been

single

mostly

yogurt

not been

digestion

results,

subsp

thermophilus

bulgaricus,

subsp milk

and

the two lactic acid yogurt. As a result

to produce

digestion, yogurt is a well-tolerated foods for who experience symptoms tose maldigesters) mercially available improve ences exist tose trial

lactose among

Am

digestion

among yogurt digestion (7). (in Minnesota)

J C/in Nuir

bacteria of this alternative of lactose

(1-5). It has been suggested yogurts provide sufficient

strains

strains of yogurt is an important

Lactobacillus

and

in the total

tolerance

(6).

or specific

from

the

species

of lactic

and

lactose

with

acid

milks

bacteria

digestion

of

acid bacteria (eg, Bilactose digestion has

we conducted

France)

lactis,

subsp

the ability

a separate

human

fermented

with

in order

in vivo.

This

an

to evaluate

to provide a basis for formulating novel lactic dairy foods designed for the lactose maldigester.

study

its

was conacid-con-

methods

I

>

In study

1, performed

20 ppm

(1.80

milk five

containing men and

Chinese one

and

X 106

were

g HilL

air)

Taiwanese),

American.

fed blind

between

making

the

after

two Each

in random

the investigator

protocol

of Minnesota,

ingestion

1 8 g lactose. The group two women. Four subjects

three

was Jewish

at the University

aged 23-33 y were classified as lactaseofa rise in breath hydrogen concentration

ofthe

order.

the laboratory refrigerator as necessary the meal labeled with his or her name; contact

used to ferment improved lactose to other intolerance

dairy (lac-

that not microbial

all comfl-gal to

In addition,

differ-

activity

in USA.

Therefore, Rouen,

dairy

including

ducted taming

below)

offl-gal

double-blind.

and

were

of an en-

of subjects inwere oriental

Italian

Americans,

five meals

Meals

(described

were

placed

in

and each subject selected this was done to eliminate the subjects, The

subjects

thus were

effectively not

in-

Station

Project

18-016.

Supported by the National Dairy Board and Syndifrais and GervaisDanone Cie and administered in cooperation with the National Dairy Council. 4 Address reprint requests to DA Savaiano, Department of Food Science and Nutrition, University ofMinnesota, St Paul, MN 55108. Received November 2, 1990. Accepted for publication April 10, 1991. 3

in lacfeeding several

© 1991 American

I From the Department ofFood Science and Nutrition, University of Minnesota, St Paul; the Groupe de physiopathologie digestive nutritionelle, UFR de Medicine, Rouen, France; and the BSN Groupe, Paris. 2 Listed as paper 18 667 ofthe scientific series ofthe Minnesota Agricultural Experiment Station on research conducted under the Minnesota

Experiment

may

cultures to determine whether strain selection factor in facilitating lactose digestion in vivo by Printed

lactis

1). Further,

or other novel lactic bijidus) to improve in vivo

(in

from

delbrueckii

cultures, possibly causing a variation Therefore, we conducted a human to evaluate lactose digestion from

l99l;54:1041-6.

(3, 9-1

experiin vitro

bacteria,

Lactococcus

and

strains

trial

riched cluded

activity of microbial 13-galactosidase (/3-gal) in the gastrointestinal tract (1, 2). The f3-gal activity comes from both Streptococcus salivarius

acid

to improve

to

live bacteria lactose-con-

in part,

the

activity

ability

(one

improved

lactic

unsuccessful

studied.

feeding

and

The

with

seven healthy subjects deficient on the basis

subsp

foods.

to produce off-gal

nonyogurt

produced

Subjects

Introduction

taming

used

foods

Study

lactose

Lactose consumed as part of yogurt containing is better digested than is lactose in milk or other

were

sub-

yogurt, dairy products, Lactobacillus bulgaricus, Streptococcus salivarius subsp Lactobacillus acidophilus, B/ldobacterium b/idus

lactase

delbrueckii thermophilus,

lactose

that

in the production

culture system (7, 8). at improving lactose digestion

Subjects. KEY

strains

varied

in a mixed skim-milk Investigations aimed

individual

with

because ofa limited rate of intracellular Am J Clin Nutr 199 1 ;54: 104 1-6.

K Harlander,

Jidobacterium

products were and breath hy-

milks

Society

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for Clinical

Nutrition

1041

1042

MARTINI

formed

ofthe

to mask

identity

the

The project man

ofeach

flavor

was approved

Subjects

Meals.

test

consisted dry-milk

of whole solids and

yogurts.

Subjects

at the

meals

were

milk four

enriched different

consumed

450 g of the four yogurts. products were formulated Minnesota

dairy

pilot

University

of Minnesota

fed

ucts); yogurt Minneapolis);

made

on the Use of Husubjects.

The

milk

meals

and

a single

herd).

source

Fresh

milk

was

(420

All of (The

homoge-

of pressure at 50-60 #{176}C and pasteurized After vanilla extract and 2% milk solids of this milk was retained and the rest was which strains

were inoculated of Lactobacillus

Products,

THY

culture Northern Regional Research Lab, 1 1842 (American Type Culture Collection, gurt 5, with ST 364 1 plus LB 880 (Marschall

(Marschall

at 40 #{176}C until

cooled to 4.5.

to 10 #{176}C. The Beta-gal activity

tration,

pH,

and

pH reached

-4.6

final pH of the (both total and

cell counts

were

(‘-6-7

measured

ranged lactose

in the

defined

healthy

Prod-

on the

milks h) and

yogurts specific),

is precipitated

was

Labo-

basis

(2.25

milk

containing

quickly

formed

tobacillus

samples ingestion

coffee ples

obtained of the

after an overnight test meals. Only

were allowed

acidophilus.

were

analyzed

for hydrogen

during and

tions by using gas chromatography Analyzer, model 12, Quintron

the 8 h. Breath

carbon

(Microlyzer Instruments,

dioxide

Medical

Gas

Analyzer,

(ppm)

was

calculated

fasting

cell

Gas and

shipped

Biochemical,

lactose

by fl-gal,

Indianapolis).

where and

lactose the

liberated

The

is hydrolyzed

fast

galactose

lactose-gaBoehringer involves

to glucose is combined

a

and gawith

Downloaded from https://academic.oup.com/ajcn/article-abstract/54/6/1041/4715102 by guest on 09 March 2018

Eth-

gave

in-

by single

conThe and strains

B bifidus,

or Lac-

contained 15 g lactose in 260were formulated and processed

fermented 2.3%

milks

were

nonfat

the milk

dry

commercial practices.

made

milk

from

solids

was inoculated

a

were

with

the

were

measured and

in the

and

every

products.

consumed within Lactose absorption

30 mm

samples

for 8 h after

The

products

were

3 d of manufacture.

was assessed by breath obtained after an overnight

ingestion

of the

test

meals.

Breath samples were obtained at the end of a normal expiration and repeated twice to ensure uniformity of hydrogen values. Mean hydrogen values were used in calculations. Breath samples

hydrogen

method

subject

was fermented

bulgaricus,

to which

Hydrogen analysis. hydrogen analysis ofbreath

subtracting

reaction

each

GD 428, Lactobacillus acidophilus GD 753, GD 24 and Lactobacillus bul47. Lactose concentration, f-gal activity, pH, and

to Rouen

(2, 4) is based on a commercial lactose method (Lactose-galactose, Mannheim

the four

pasteurization,

GD

were

coupled

from order

of S thermophilus

counts

values from subsequent test values. The extent and severity of intolerance symptoms were also recorded by subjects. Lactose and fl-gal analysis. Lactose concentrations and a-gal activity were measured in the test meals. The assay for lactose spectrophotometric UV method,

males

by the Human

and

the milk

ofmilk,

After

garicus

concentra-

model

by subtracting

was approved

25

>

in a random

Research Center in Paris, by using and according to good manufacturing

and

source

or a mixture

ments, Fullerton, CA, respectively). ues were corrected for atmospheric

excretion

to

of 1.2% low-fat were

fed

12

GD 086, B bijidus

LBII, Beckman InstruThe observed hydrogen valcontamination of alveolar air by normalization of the observed carbon dioxide to 45 mm Hg, which is the Pco2 in the veins (1). The change in hydrogen Beckman

of pro-

appropriate species and incubated for roughly 3.5 h at 42 #{176}C until a pH of 4. 1-4.2 was reached. The strains used for inoculation were S thermophilus GD 245, Lactobacillus bulgaricus

sam-

Hydrogen Milwaukee,

concentration

subjects

University

Each meal All products

or milk.

yogurt

added. absorption breath

fast and hourly for 8 h after water, black tea, and black

to be consumed

unit mg

.

by Student’s

ingestion was

Lactobacillus

in the Dannon Dannon strains

gurts

All

The project

bulgaricus;

single

the

hydrogen

six meals

of Rouen

of S thermophilus,

prod-

(12). and

were consumed within 2 wk of fermentation. Hydrogen and carbon dioxide analysis. Lactose was assessed by breath-hydrogen analysis ofend-alveolar

One

min

-

at Rouen University, classified as lactase-deficient

air) after

15 g lactose.

Lactobacillus

from 4.1 concen-

methods

pasteurization

acid.

released

Meals. Six test meals were fed to the subjects. The meals sisted of low-fat milk, yogurt, and four fermented milks. yogurt was fermented by using a mixture ofS thermophilus

The

standard

5 d after

was analyzed by the measured is a cellular

consent.

yo-

by using within

o-

All chemicals

‘.

was determined

2, performed 18-26 y were

of the

ical Committee

The

determined

consumed

in A420 was as 1 tmol

Co (St Louis).

0.05)

X 10-6 g HilL Each

counts

were

increase

g yogurF

ONP

of a rise in breath

ppm

294 g yogurt

was

(2, 4). Absorbance

the

was defined -

in

is stoi-

(13, 14). by using a spec-

by trichloroacetic

(P

study aged

and double-blind.

were

yogurt

In subjects

ucts. Four of these yogurts (yogurt 3, yogurt 4, yogurt 5, and yogurt 6) were then chosen as the test meals because they cxhibited a wide range of total and specific 9-gal activities. Cell milk

and

described until

as 1 mol

significance

Cameroon.

LB 880.

incubated

previously measured

that

Subjects.

(Yoplait, of Agri-

enriched

method

activity

2

the increase

the amount of original lactose analyzed for total f3-gal activity

protein

Study

Peoria, IL) plus LB Rockville, MD); yoProducts); yogurt

inoculated,

NADH;

Specific activity of 3-gal in the yogurts method ofLin et al (7) whereby the protein

from with

with

The

and

linear for 10 mm. One unit ofactivity nitrophenol (ONP) released - min’ were obtained from Sigma Chemical

(15).

(New Zealand Dairy Research Institute, New Zealand) plus LB 1 1842; and yogurt 7,

plus

H,

spectrophotometrically

at 37 #{176}C) was

t test

6, with ST TS2B Palmerston North, ST TS2B

nm

acid,

bul-

Miles

3, with strains isolated from Yoplait yogurt yogurt 4, with ST 3641 (US Department

with were

tein’. Statistical

with

Each ofthe yogurts were made as follows: yogurt 1, inoculated

LB-ST (Marschall WI); yogurt 2, with

galactonic

be monitored

trophotometric

425-

of milk

can

chiometric Yogurts

of Minnesota.

to the

to form

NADH

with 2% (by weight) nonfat unsweetened vanilla-flavored

from

dairy

NAD

consent.

Each meal contained 18 g lactose. and processed in the University

garicus and S thermophilus. the inoculation of the milk mix

was

informed

University

divided into seven aliquot samples, one of several mixtures of different

the commercial ratories, Madison,

no attempt

gave

3 1 5 g of enriched

plant

nized at 17.24 MN/m2 at 85 #{176}C for 30 mm. were added, a portion

but

Subjects

by the Committee

in Research

Five

product

or texture.

ET AL

analyzed

matograph

for hydrogen with

Analyzer, manufactured The change in hydrogen fasting

concentrations

a thermal-conductivity

hydrogen

by D Wouterson, excretion (ppm) values

by using

a gas chro-

detector

from

(Stimotron

Wendelstein, FRG). was calculated by subsequent

test values.

Intolerance Lactose

symptoms were not monitored. and 13-gal analysis. The specific

activity

measured

in three

20 g ofthe

were diluted

ofthe

products.

in 200 mL ofa solution

Initially,

of f3-gal was

(0. 1 mol Na2HPO4

products -

2H2O/

DAIRY L, 0. 1 5 mol that

NaCl/L,

would

0.0 1 mol

solubilize

trifugations

(16

the pellet

the

in 0. 1 mol

EDTA/L,

casein

x g,

300

in the

15 mm,

phosphate

BACTERIA 0.04

mol

products.

two

7.0),

the cells

pernatant

2 mL

was

(ONPG)/L, mm.

halted

The measurement method by using

times

containing

in a 37 #{176}C water by the

ONP

addition

method

.

were

used

in study

techniques two-way

as A divided

released

concentrations

standard

phosphate

bath

of 2 mL and ‘

activity

L

-

(16).

analysis

min

1 . Cell

of variance

by the

counts

were

cxto the Co).

were

FIG 1 . Change in concentration ofbreath hydrogen after ingestion of enriched whole milk (0), yogurt 3 (made from Yoplait strains)(O), yogurt 4 (made from ST 3641 + LB 1 l842)(L), yogurt 5 (made from ST 3641 + LB 880) (A), and yogurt 6 (made from ST TS2B + LB 1 1842) (#{149}). I ± SE for seven lactase-deficient subjects.

spectrophotomeasured

statistically

with

analyzed

by

(15).

Results

The after

Study

1

Table

1 shows

specific of the

/3-gal activity yogurts were

other reported ranging from typical mol

the lactose

of the seven approximately

cell counts, yogurts. equal

The and

and total

/3-gal activity . g product’

from 3.3 to 6.7 mol ONP released 3, 4, 5, and 6 were thus selected

min for

ofbreath

was

No

than

significant

duction

approximately it was differences

among

TABLE 1 Characteristics

threefold after

were

after

of any

observed

in peak

S

1 2 3 4 5 6 7

Percentage lactose*

product

(LB-ST) (THY) (Yoplait) (ST 3641 (ST 3641 (ST TS2B (ST TS2B

g lactose/lOO

t mol jmol

+ + + +

meals

from

were

milk

consumption

(P

among