EFFECT OF CONSUMPTION OF AQUEOUS EXTRACT OF HIBISCUS SABDARIFFA AND AZADIRACHTA INIDCA DURING PREGNANCY AND LACTATION ON BREAST MILK CREAMATOCRIT AND EARLY POSTNATAL GROWTH OF OFFSPRING

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ABSTRACT
Extracts of Hibiscus Sabdariffa (HS) and Azadirachta Indica (AI) are widely used for medicinal purposes and have been shown to affect postnatal growth of the offspring through mechanisms not yet defined. Breast milk creamatocrit is the length of the cream column separated from milk by centrifugation and expressed as a percentage of the length of the total milk column. It is a simple accurate   inexpensive technique and useful measure of the lipid concentration and the energy content of   breast milk. No data  has  shown  the  effect  of these two  extracts  on breast milk  creamatocrit  which is a vital factor  for the  early  postnatal  growth of offspring. The  study  was  designed  to  investigate the effects of the two  extract  on breast  milk creamatocrit  and  early  postnatal growth of offspring. 40 pregnant rats weighing 150-200g were used for this study. They were divided into control, HS and AI groups. HS and AI groups were subdivided into two subgroups of low and high dose. Extract administration commenced on day 1 of pregnancy and ended on postnatal day 21. On the day of delivery litter sizes, length and weights were recorded. Litter weight and length were subsequently recorded at postnatal days 7, 14 and 21. Breast milk was collected from 4 dams in each group into a plain sample container at postnatal day 7, 14 and 21 for determination of breast  milk  creamatocrit.  Milk sample volume collected ranged between 0.4ml-0.5ml per dam. The study revealed that the aqueous extract of Hibiscus Sabdariffa and Azadirachta Indica increased consumption of food and fluid during pregnancy and lactation, increased maternal weight gain during pregnancy and lactation and also caused a progressive increase in breast milk creamatocrit during lactation days 7, 14, 21. The progressive increase in breast milk creamatocrit values peaked greatly in the third week of lactation.  The aqueous extract of Hibiscus Sabdariffa and Azadirachta Indica caused a progressive increase in BMI of the extract treated group that became significantly greater than that of the control at weaning. From the results of the present study, it can be concluded that consumption of extracts of HS and AI during pregnancy and lactation increases fluid, food intake and weight gain while accelerating the postnatal growth of the offspring through increased breast milk creamatocrit.

TABLE OF CONTENTS

Title page
Abstract
Table of contents
List of Table
List of Figures

CHAPTER ONE
1.0       Introduction
1.1       Statement of Problem
1.2       Aim   and Objectives
1.2.1    Aim
1.2.2    Objectives
1.3       Operational Definition of Terms

CHAPTER TWO
2.0       Literature Review
2.1       Hibiscus sabdariffa
2.1.1    Botanical Description
2.1.2    Geographical Distribution and Common Names
2.1.3    Composition of Hibiscus sabdariffa
2.1.3.1 Nutritive Value
2.1.3.2 Phytochemical Constituents
2.1.4    Pharmacological Properties of Hibiscus sabdariffa
2.1.4.1 Antioxidant Properties
2.1.4.2 Nephroprotective Properties
2.1.4.3 Delayed Puberty Activity
2.1.5    Uses of Hibiscus sabdariffa
2.1.5.1 Medicinal Use
2.1.5.2 Food Use
2.1.5.3 Non Medicinal Use
2.2       Azadirachta indica
2.2.1    Botanical Description
2.2.2    Geographical Distribution and Common Names
2.2.3    Composition of Azadirachta indica
2.2.3.1 Phytochemical Constituents
2.2.4    Pharmacological Properties of Azadirachta indica
2.2.4.1 Anti-Bacterial Properties
2.2.4.2 Antioxidant Properties
2.2.5 Uses of Azadirachta indica
2.2.5.1 Medicinal Use
2.2.5.2 Non Medicinal Use
2.3 Breast Milk
2.3.1 Development of Breast (Mammary Gland) In Rats
2.3.2 Onset of Lactation in Rats
2.3.3 Stages of Lactation
2.3.4 Composition of Breast Milk
2.3.5 Factors Affecting Breast Milk Composition
2.4 Postnatal Development in Rats
2.4.1 Factors Affecting Early Postnatal Development

CHAPTER THREE
3.0       Material and Methods
3.1       Plant Collection and Identification
3.2       Preparation of Plant Extract
3.3.      Phytochemical Analysis
3.4       Experimental Animals
3.5    Induction of Pregnancy in Rats
3.6       Experimental Design
3.7    Extract Administration
3.8    Maternal Body Weight Measurement
3.9    Measurement of Food and Fluid Intake
3.10     Measurement of Litter Size, Length and Weight
3.11     Breast Milk Extraction
3.12     Breast Milk Creamatocrit Measurement
3.13     Statistical Analysis

CHAPTER FOUR
4.0    Results
4.1    Extract Yield
4.2    Phytochemical Screening
4.3.1 Effect of Consumption of Aqueous Extract of Hibiscus sabdariffa on Food Intake During Pregnancy
4.3.2 Effect of Consumption of Aqueous Extract of Azadirachta indica on Food Intake During Pregnancy
4.3.3 Effect of Consumption of Aqueous Extract of Hibiscus sabdariffa on Fluid Intake During Pregnancy
4.3.4 Effect of Consumption of Aqueous Extract of Azadirachta indica on Fluid   Intake During Pregnancy
4.3.5 Effect of Consumption of Aqueous Extract of Hibiscus sabdariffa on Maternal Weight During Pregnancy
4.3.6 Effect of Consumption of Aqueous Extract of Azadirachta indica on Maternal Weight During  Pregnancy
4.3.7 Effect of Consumption of Aqueous Extract of Hibiscus sabdariffa on Food Intake During Lactation
4.3.8 Effect of Consumption of Aqueous Extract of Azadirachta indica on Food Intake During Lactation
4.3.9  Effect of Consumption of Aqueous Extract of Hibiscus sabdariffa on Fluid Intake During Lactation
4.3.10 Effect of Consumption of Aqueous Extract of Azadirachta indica on Fluid Intake During Lactation
4.3.11 Effect of Consumption of Aqueous Extract of Hibiscus sabdariffa on Maternal Weight During Lactation
4.3.12 Effect of Consumption of Aqueous Extract of Azadirachta indica on Maternal Weight During Lactation
4.3.13 Effect of Consumption of Aqueous Extract of Hibiscus sabdariffa on Creamatocrit Value of the Breast Milk (Cream/Lipid) In Percentage(%)
4.3.14 Effect of consumption of aqueous extract of Azadirachta indica on creamatocrit value of the breast milk (cream/lipid) in percentage (%)
4.3.15  Effect of Consumption of Aqueous Extract of Hibiscus sabdariffa During Lactation on Body Weight of Offspring
4.3.16  Effect of Consumption of Aqueous Extract of Azadirachta indica During Lactation on Body Weight of Offspring
4.3.17  Effect of Consumption of Aqueous Extract of Hibiscus sabdariffa During Lactation on BMI of Offspring
4.3.18  Effect of Consumption of Aqueous Extract of Azadirachta indica During Lactation on BMI of Offspring

CHAPTER FIVE
5.0       Discussion and Conclusion
5.1       Maternal Study
5.1.1    Food and Fluid Intake in Dams That Consumed Hibiscus sabdariffa During Pregnancy and Lactation
5.1.2    Food and Fluid Intake in Dams That Consumed Azadirachta indica During Pregnancy and Lactation
5.1.3 Weight of Dams That Consumed Hibiscus sabdariffa During Pregnancy and Lactation
5.1.4 Weight of Dams That Consumed Azadirachta indica During Pregnancy and Lactation
5.1.5   Breast Milk Creamatocrit of Dams That Consumed Hibiscus Sabdariffa and Azadirachta indica During Lactation
5.2       Offspring Study
5.2.1 Postnatal Growth of Offspring Whose Mother Consumed HS and AI During Lactation
5.3       Conclusion
5.4       Recommendation
            Summary of Findings
            References  


CHAPTER ONE
1.0 INTRODUCTION
          Medicinal plants  over the  years  has  been part  and  parcel of  human society  to  combat  diseases  (Kausik  et al., 2002). These  plants  has  been a good  source of food because they provide essential nutritional values, medicinal properties  and  notable  physiological  effect   to life (Dalziel,  1973) .
          Hibiscus sabdariffa  and  Azadinachta  indica  are  used  as  medicinal and food  ingredients  in  many  parts  of the world including Nigeria. Both plants  are  highly  sourced  as food  vegetable particularly because of their health promoting  and  diseases- preventing  properties  strongly  suspected  to be  due to the presence  of many phytochemicals in them (Ijeomah et al, 2012). These phytochemicals  present  like alkaloid, saponin, glycoside, tannin, phenol, flavonoid, steroid , reducing   sugar,  nimbidin,  sodium  nimbidate, nimbin, gedunin,  delphinidin  3-  sambubioside  and  protocatechuic  acid  in the  two  plants  have been  found  to be  protective and preventive against  many degenerative diseases and pathological process such as in ageing (Burns  et al., 2001). Hibiscus sabdariffa (family: malvaceae) commonly known as zobo in Nigeria, is an annual herbaceous shrub,  cultivated  for its  flowers, leaves  and seeds. It is found in the tropics, subtropics and other parts of the world (Dalziel, 1973) but it is utilized beyond these areas of cultivation globally. In folk medicine this   medicinal herb is used for the treatment of hypertension (Wang et al., 2000; Odigie et al., 2003). The plant is reported to have hepatoprotective, antihypertensive, antihyperlipidemic, anticancer and antioxidant properties (Mahadevan and Shivali, 2009).
          Azadirachta indica which is commonly referred to as Dogonyaro or neem plant is an evergreen robust tree belonging to the family meliaceae. It is mostly  found in tropics  and sub- tropical  areas  of the  world African and Asia  (Ejoba, 2012) .The  tree occurs  in  medium to large  size  and  has  dark  grey  bark  and  a dense  rounded  row  of pinnate  leaves (Ogbuewu, 2008). All parts of the neem tree (leaves, flowers, seeds, fruits, roots and bark) are widely used in traditional medicine.  Azadirachta  indica is used for the treatment  of many  health related  problems  and also  known to  extent  anticancer, antioxidant, wound  - healing  and  antimicrobial properties  (Hybercyclopedia, 2014). According to (Ogbuewu, 2008; Mossini et al., 2004), all parts of this plant are useful and has been used to treat diseases ranging from tooth decay, ulcer, swollen liver, malaria and dysentery.
          Breast  milk  is the normative  standard  for  infant  feeding  and nutrition because it has been associated with a reduction in the  incidence  of gastrointestinal tract  infection, respiratory  tract  infection  and  otitis media (AAP, 2012; WHO,  2013). It is important to note that breast milk is widely accepted to be the optimal source  of nutrition for the newborn infants because it contains appropriate amount  of  protein,  lipid, carbohydrate, micronutrients, trace elements, vitamins, minerals, digestive enzyme and hormones (Chantry  et al., 2011) required for early postnatal  growth  of  offspring, development and immune protection (Ostrom, 1990). However  it is   now  universally  recognized  that  there is no  commercial formula  that  can equal breast milk. The relationship of breast milk feeding with a significant reduction in the incidence of necrotizing enterocolitis (NEC) has been suggested in preterm infants (Sullivan, 2010).
          Additionally there is extensive evidence that offspring who had been breastfed with breast milk show lower risk of same non- communicable diseases later in life (AAP, 2012; WHO, 2013).  Indeed  breast  milk  given to infant at the  early  postnatal  stage  has  been associated  with  lower  risk of obesity, Lower levels of   arterial blood  pressure,  lower  total  and  LDL-Blood  cholesterol  level in adulthood  and  lower   risk of developing  type – 2  diabetes  (AAP,  2012; WHO,  2013). Breast milk is not only associated  with  improved  parameters of growth  in  offspring  but  also  with a better  neuronal-behavioral development  (Mortensen et al.,  2002;  AAP,  2012). Breast milk creamatocrit is the cream  column  separated from the milk by  centrifugation and  expressed as a percentages of the  length  of the  total  milk  column.  It is  a simple  method of  estimating  the calorific  value of  breast  milk ( Lucas,  1978;  Wang  et al., 1999).
          Infant  growth  during  the  early  postnatal period is obviously  the  result  of the  development of complex  functional, structural  and  metabolic  mechanisms, which are however not yet entirely clarified (Patel and Srinivasan, 2010). However  this period is  characterized  by a great plasticity  with  critical windows in which any inadequate  insult or intervention  may  cause   both positively and  adversely  postnatal  growth  and  development (Ostadelova and  Babicky,  2012). This  study is to  determine  the  effect of  consumption of aqueous  extract  of  Hibiscus  sabdariffa  and  Azadirachta  indica  during  pregnancy and lactation on breast milk creamatocrit  and  early  postnatal  growth  of  offspring, considering the fact that the extract is widely used socially.
1.1 STATEMENT OF PROBLEM  
          The  sweetened  aqueous  extract  of  Hibiscus  sabdariffa  is consumed  by both males and females including  pregnant  and lactating  mothers  not necessarily  for medicinal purposes but as a substitute for carbonated drinks (Iyare and Adegoke, 2011) while Azadinachta indica is commonly and  indiscriminately used in traditional  medicine for the treatment of malaria in Nigeria (Ucheya et al., 2011). These  plant  extract  are  consumed  without  regards  to the physiological  state  of the body   (Iyare  and Adegoke, 2011). There  is  paucity of information  on the  effect of these  two plant  extracts on breast  milk  creamatocrit  which is a vital  factor  for the  early  postnatal  growth  of  offspring.......

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